201001_psychiatry

israel journal of

psychiatry

Vol 47 - Number 1 2010

ISSN: 0333-7308

2

Editorial: Etiological Hypotheses of Mental Disorders at the Molecular Level Galila Agam, Dorit Ben-Shachar

4

Glutamatergic theories of schizophrenia Daniel C. Javitt

17

Impaired inhibitory regulation of pyramidal neurons in schizophrenia Stephen I. Deutsch, Richard B. Rosse, Barbara L. Schwartz, John Mastropaolo, Jessica A. Burket, Abraham Weizman

27

Circadian rhythms and clock genes in psychotic disorders Elaine Waddington Lamont, Daniel L. Coutu, Nicolas Cermakian, Diane B. Boivin

36

Intrauterine factors as determinants of depressive disorder Marta Weinstock

46

The biology of tryptophan depletion and mood disorders Lilach Toker, Shirly Amar, Yuly Bersudsky, Jonathan Benjamin, Ehud Klein, Galila Agam

56

Tryptophan–Kynurenine Metabolism in Major Depression Gregory F. Oxenkrug

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Special section:

biochemical and anatomical substrates of depression and sickness behavior Thomas C. Hanff, Stephanie j. Furst, Thomas R. Minor

Etiological Hypotheses of Mental Disorders at the Molecular Level Guest Editors: Galila Agam, Dorit Ben-Shachar

72

Genetics of unipolar major depressive disorder Tanya Goltser–Dubner, Esti Galili-Weisstub, Ronnen H. Segman

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Commentary: etiological hypotheses of mental disorders at the molecular level may not help psychiatry Jonathan Benjamin

israel journal of

psychiatry

The Official Publication of the Israel Psychiatric Association Vol 47 - Number 1 2010

and related sciences EDitor

Special section:

David Greenberg PAst Editor

Etiological Hypotheses of Mental Disorders at the Molecular Level

Eli L. Edelstein

Guest Editors: Galila Agam, Dorit Ben-Shachar

Founding Editor

Heinz Z. Winnik Deputy Editor

Rael Strous Editorial Board

Alean Al-Krenawi Alan Apter Yoram Barak Elliot Gershon Talma Hendler Ehud Klein Ilana Kremer ltzhak Levav Yuval Melamed Shlomo Mendlovic David Roe Ronnen Segman Eliezer Witztum Zvi Zemishlany International Advisory Board

Yoram Bilu Aaron Bodenheimer Carl Eisdorfer Julian Leff Margarete Mitscherlich-Nielsen Peter Neubauer Phyllis Palgi Leo Rangell Melvin Sabshin Robert Wallerstein Myrna Weissman Assistant Editor

Section 1: Psychotic Disorders

2 > Editorial: Etiological Hypotheses of Mental Disorders at the Molecular Level Galila Agam, Dorit Ben-Shachar

46 > The biology of tryptophan depletion and mood disorders Lilach Toker, Shirly Amar, Yuly Bersudsky, Jonathan Benjamin, Ehud Klein, Galila Agam

56 > Tryptophan–Kynurenine Metabolism

as a Common Mediator of Genetic and Environmental Impacts in Major Depressive Disorder: The Serotonin Hypothesis Revisited 40 Years Later

Gregory F. Oxenkrug

4 > Glutamatergic theories of

64 > biochemical and anatomical substrates of depression and sickness behavior

Daniel C. Javitt

Thomas C. Hanff, Stephanie j. Furst, Thomas R. Minor

schizophrenia

17 > Regulation of Intermittent Oscillatory Activity of Pyramidal Cell Neurons

by GABA Inhibitory Interneurons is Impaired in Schizophrenia: Rationale for Pharmacotherapeutic GABAergic Interventions

Stephen I. Deutsch, Richard B. Rosse, Barbara L. Schwartz, John Mastropaolo, Jessica A. Burket, Abraham Weizman

27 > Circadian rhythms and clock genes in psychotic disorders

Elaine Waddington Lamont, Daniel L. Coutu, Nicolas Cermakian, Diane B. Boivin

Section 3: Genetics

72 > Genetics of unipolar major depressive disorder

Tanya Goltser–Dubner, Esti Galili-Weisstub, Ronnen H. Segman

83 > Commentary: etiological hypotheses of mental disorders at the molecular level may not help psychiatry Jonathan Benjamin

87

> Book Reviews

Hebrew Section Section 2: Major Depression

36 > Intrauterine factors as determinants of depressive disorder

87

> News and Notes

90

> Abstracts

Marta Weinstock

Joan Hooper

Marketing: MediaFarm Group

+972-77-3219970 23 Zamenhoff st.

Tel-Aviv 64373, Israel

amir@mediafarm.co.il www.mediafarm.co.il

Self portrait, 2000 Dudi Eldar

Dudi Eldar, a graduate of the Bezalel Acadamy of Arts and Design in ceramic design, is a sculptor and painter. In his work in recent years he has been looking into the justification of the existence of abstract in reality.

Isr J Psychiatry Relat Sci - Vol 47 - No. 1 (2010)

Editorial: Etiological Hypotheses of Mental Disorders at the Molecular Level The last decade has brought up novel methodologies and increased volume of research in the field of translational science in psychiatry. We regard it timely and highly important to bring to the readership of the Israel Journal of Psychiatry an updated review of neurobiological aspects of psychosis and affective disorders written by worldwide experts. Obviously this issue provides just a glance into the wide range of topics currently studied focusing on etiological hypotheses of mental disorders at the molecular level. The review papers in this issue deal with molecular and genetic factors possibly contributing to the propensity for psychosis and affective disorders and to the development of novel treatment strategies.

Three different etiological hypotheses of psychotic disorders are presented by Drs. Javitt, Deutsch et al. and Waddington Lamont et al. Dr. Javitt elaborates on the involvement of the glutamatergic-NMDA neurotransmission system in schizophrenia and its treatment. He starts with an eloquent description of the induction of schizophrenia related symptoms by NMDA receptor blockers, continues with an integration of the glutamate hypothesis with the dopamine and GABA hypotheses of schizophrenia, and finally summarizes results of clinical trials with NMDA agonists (glycine, D-serine, etc.).

Dr. Deutsch and his colleagues present the concept of potential GABAergic intervention in impaired high executive functions in schizophrenia. Their hypothesis is based on evidence that specific inhibitory GABA inter-neurons in the frontal cortex interact with, and affect oscillations of intermittent pyramidal neurons critical for high cortical functions such as working memory. Intermittent unsustained GABAergic intervention is a challenge for future antipsychotic treatment development. Dr. Waddington Lamont et al. focus on molecular clock mechanisms. Sleep disturbances are common in psychiatric disorders, in general, and in psychotic disorders, in particular. They present evidence that prolonged sleep deprivation induces hallucinations and psychotic symptoms reminiscent of schizophrenia, and that a relationship between the sleep-wake cycle and changes in mood is apparently important in bipolar 2

disorder. Indeed, genetic studies found associations of the circadian rhythm genes CLOCK, PER1, PER3, and TIMELESS with schizophrenia and bipolar disorder.

Three different etiological hypotheses of affective disorders are presented by Drs. Weinstock, Klein, Agam and colleagues, Oxenkrug and Hanff and Minor. Dr. Weinstock emphasizes the contribution of environmental factors such as intrauterine exposure to stress in the increased vulnerability to major depressive disorder (MDD). Studies in animal models show that following chronic inescapable stress, prenatal stress or alcohol consumption the excess release of corticotropin-releasing hormone (CRH) and cortisol reduces birth weight and impairs the feedback regulation of the hypothalamic-pituitary-adrenal (HPA) axis and the signaling via the 5-HT1A and 5-HT2A receptors, and induces alterations in sleep and circadian rhythms. These changes, reversed by antidepressants, are reminiscent of symptoms in patients with MDD. The following two papers deal with the serotonergic hypothesis of affective disorders focusing on the serotonin precursor, the essential amino acid tryptophan. Drs. Klein, Agam and colleagues review human studies using the tryptophan depletion paradigm in healthy subjects, in untreated remitted or non-remitted and treated remitted MDD patients and in euthymic as well as manic bipolar patients. While in MDD diagnosed patients tryptophan depletion was generally associated with exacerbation of depression, manic patients benefited from short-term tryptophan depletion. The tryptophan depletion paradigm, which results in reduced brain serotonin, opens a window to the complex relationship between serotonin and mood disorders. Dr. Oxenkrug raises an alternative mechanism for serotonin deficiency in depression.

He argues that metabolism of tryptophan is shunted away from serotonin production towards kynurenine production. Both stress hormones and pro-inflammatory cytokines, implicated in depression, activate the rate-limiting enzymes of kynurenine formation. The neurotropic activity of kynurenines suggests that the upregulation of the tryptophan–kynurenine pathway

Galila Agam and Dorit Ben-Shachar 

not only augments serotonin deficiency but also underlies depression-associated anxiety, psychosis and cognitive decline. Drs. Hanff and Minor present the concept of possible interrelationship between organic malady and mood disorders. In particular, they elaborate on conservation-withdrawal, characterized by lethargy, hypoactivity, decreased libido, anorexia, anhedonia and increased sleep. It accompanies infectious disease, after-reaction to traumatic stress and recuperation from injury, but is also an integral component of major depression and related mood disorders. Drs. Hanff and Minor further review the role of the pro-inflammatory cytokine interleukin-1β (IL-1β) and its ability to recruit adenosine signaling at adenosine A2A receptors in mediating symptoms of conservation-withdrawal in illness as well as depressive behavior.

Although the etiology of psychotic and affective disorders is as yet not unraveled, heritable contribution for their phenotype is strongly supported by twin, adoption, familial and population studies. Dr. Segman and his colleagues review the current literature of genes possibly conferring risk for mental disorders and the genetic methodologies employed for their discovery.

Recent advances in the field of genetics of complex disorders, such as non-hypothesis-driven genome-wide

association studies in large cohorts, is suggested to hopefully overcome the scarcity of replications of risk genes reported thus far. In addition to single nucleotide polymorphism (SNP) variations reported in psychiatric disorders, which contribute a minute relative risk, the authors also discuss recent findings of copy number variations (CNVs) observed in rare cases but holding a very high relative risk for carriers. This special issue closes with a short commentary by Dr. Benjamin that challenges the odds to unravel the biological mechanisms of mental disorders. Regardless, the massive efforts invested so far towards the discovery of the pathophysiology of mental disorders have a heuristic value and will potentially culminate in better understanding and more adequate treatment modalities.

Galila Agam, PhD Psychiatry Research Unit, Faculty of Health Sciences, Ben-Gurion University of the Negev and Mental Health Center, Beer-Sheva, Israel

galila@bgu.ac.il

Dorit Ben-Shachar, PhD Laboratory of Psychobiology, Dept Psychiatry, Rambam Medical Center and B. Rappaport Faculty of Medicine, Technion, Haifa, Israel

shachar@tx.technion.ac.il

guest editors

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Isr J Psychiatry Relat Sci - Vol 47 - No. 1 (2010)

Glutamatergic theories of schizophrenia Daniel C. Javitt, MD, PhD Director, Schizophrenia Research Center, Professor of Psychiatry and Neuroscience, Nathan Kline Institute for Psychiatric Research/ New York University Langone School of Medicine, Orangeburg, New York, U.S.A.

ABSTRACT Schizophrenia is a serious mental disorder that affects up to 1% of the population worldwide. Traditional models of schizophrenia have emphasized dopaminergic dysfunction. Over the last 20 years, however, limitations of the dopamine model have become increasingly apparent, necessitating development of alternative models. Glutamatergic models are based upon the observation that the psychotomimetic agents such as phencyclidine (PCP) and ketamine induce psychotic symptoms and neurocognitive disturbances similar to those of schizophrenia by blocking neurotransmission at N-methyl-D-aspartate (NMDA)-type glutamate receptors. Because glutamate/NMDA receptors are located throughout the brain, glutamatergic models predict widespread cortical dysfunction with particular involvement of NMDA receptors throughout the brain.

Further, NMDA receptors are located on brain circuits that regulate dopamine release, suggesting that dopaminergic deficits in schizophrenia may also be secondary to underlying glutamatergic dysfunction.

Agents that stimulate NMDA receptor-mediated neurotransmission, including glycine-site agonists and glycine transport inhibitors, have shown encouraging results in preclinical studies and are currently undergoing clinical development. Encouraging results have been observed as well with agents such as metabotropic 2/3 agonists that decrease resting glutamate levels, reversing potential disruption in firing patterns within prefrontal cortex and possibly other brain regions.

Overall, these findings suggest that glutamatergic theories may lead to new conceptualizations and treatment approaches that would not be possible based upon dopaminergic models alone.

Introduction Schizophrenia is a serious mental disorder that affects up to 1% of the population worldwide, and is one of the leading causes of chronic disability. Although causes of schizophrenia remain unknown, the disease has been extensively characterized from both a symptomatic and neurocognitive perspective, and much information has accumulated about elements such as genetic causation and longitudinal course. Although schizophrenia was once seen as a disease affecting only a few key brain regions and regionally discrete neurotransmitter systems such as dopamine, more recent findings implicate widespread cortical and subcortical dysfunction, suggesting more generalized etiology. On a neurochemical level, antagonists of N-methyl-D-aspartate (NMDA)type glutamate receptors, such as phencyclidine (PCP) or ketamine, uniquely reproduce the symptomatic, neurocognitive and neurochemical aspects of the disorder, suggesting that regardless of underlying etiology, NMDA dysfunction represents a final common pathway leading from pathogenesis to symptoms.

Clinical Phenomenology of Schizophrenia Symptoms of schizophrenia are typically divided into three main classes termed positive, negative and cognitive. Positive symptoms consist of such items as suspiciousness/persecution, grandiosity, delusions, and unusual thought content and, in general, reflect features of the schizophrenia experience that are not shared by the general population. Negative symptoms, in conConflict of interest: Dr. Javitt holds intellectual property rights for use of glycine, D-serine and glycine transport inhibitors in treatment of schizophrenia and related disorders.

Address for Correspondence: Daniel C. Javitt, MD, PhD, Director, Program in Cognitive Neuroscience and Schizophrenia, Nathan Kline Institute for Psychiatric Research 140 Old Orangeburg Road Orangeburg, New York, 10962 U.S.A.  Javitt@nki.rfmh.org

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Daniel C. Javitt

trast, consist of symptoms such as lack of spontaneity, social/emotional withdrawal, poor rapport and blunted affect, and reflect features of normal experience that are reduced in individuals with schizophrenia. Cognitive symptoms – which are also referred to as disorganized symptoms or autistic preoccupation – consist of such elements as conceptual disorganization, disorientation and poor attention. Dopaminergic models of schizophrenia account well only for positive symptoms of the disease. In contrast, glutamatergic models account much more fully for both negative and cognitive symptoms, and thus may serve as an etiological model for the syndrome as a whole.

Another key component of schizophrenia is neurocognitive dysfunction. When tested on basic IQ tests, such as the WAIS, patients with established schizophrenia typically score about 1 standard deviation, or 15 IQ points, below the population mean. Deficits are typically present at first episode and remain relative constant over the course of the illness, suggesting that cognitive decline precedes the onset of substantial symptoms (1, 2). Prospective, follow-back and crosssectional data all suggest that cognitive functioning may decline during the 3–4 years immediately preceding the onset of schizophrenia symptoms. For example, in one prospective study, poor educational achievement at age 15 was a significant predictor of schizophrenia (3). Two follow-back studies have investigated performance on standardized educational testing (Iowa test) during childhood and adolescence in individuals who subsequently developed schizophrenia. Compared with the general population, such individuals showed only modest deficits even when assessed during 4th and 8th grade, but showed a marked decline in performance between 8th and 11th grade (4, 5).

Similarly, individuals with prodromal schizophrenia who have not yet converted to psychosis show cognitive deficits that are intermediate between those of first-episode and control subjects, and such deficits may predict subsequent conversion to psychosis (6). In a study using the Israeli army database, lower than expected IQ at age 17 – based upon childhood reading and spelling abilities – was a significant risk factor for schizophrenia but not bipolar disorders, such that individuals showing a 10 point or greater discrepancy between expected and actual IQ showed an approximately two-fold elevated risk for developing schizophrenia (7).

Also based upon findings from the army database, it appears that intellectual performance remains rela-

tively constant between age 17 and subsequent illness onset in individuals who go on to develop schizophrenia, suggesting that most of the cognitive decline occurs premorbidly, although further deterioration in some domains may be observed (8). Overall, these findings highlight neurocognitive dysfunction as a key manifestation of schizophrenia that precedes onset of symptoms, and must therefore be considered central to etiological hypotheses.

Neurochemical Models of Schizophrenia The first effective treatments for schizophrenia were discovered fortuitously in the late 1950s, and subsequently shown to mediate their effects at dopamine D2 receptors. Since that time, dopamine has been the primary neurotransmitter implicated in schizophrenia, and the majority of neurochemical studies of schizophrenia continue to focus on dopaminergic mechanisms (9, 10).

Neurochemical models of schizophrenia based upon dopamine have had substantial heuristic value in explaining key symptoms of schizophrenia, in particular, positive symptoms, and in guiding treatment considerations. Nevertheless, significant limitations with regard to the dopamine hypothesis remain. First, no intrinsic deficits have been observed within the dopamine system to account for the presumed hyperdopaminergia associated with schizophrenia. Second, reconceptualizations of the dopamine hypothesis propose that subcortical hyperdopaminergia may coexist with cortical hypodopaminergia, although mechanisms underlying the differential cortical and subcortical abnormalities remain Figure 1. Schematic Model of the NMDA Receptor Complex. NMDA=N-methyl-D-aspartate; GLY=glycine, GSH=glutathione.

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GLUTAMATERGIC THEORIES OF SCHIZOPHRENIA

to be determined. Finally, dopaminergic dysfunction, in general, accounts poorly for symptom classes in schizophrenia other than positive symptoms and for the pattern of neurocognitive dysfunction associated with schizophrenia. Thus, alternative conceptual models of schizophrenia are required.

An alternative to the dopamine model was first proposed in the early 1990s, based upon the observation that phencyclidine (PCP), ketamine and other similarly acting psychotomimetic compounds induced their unique behavioral effects by blocking neurotransmission at N-methyl-D-aspartate (NMDA)-type glutamate receptors (11, 12) (Figure 1). The ability of these compounds to transiently reproduce key symptoms of schizophrenia by blocking NMDA receptors led to the concept that symptoms in schizophrenia may reflect underlying dysfunction or dysregulation of NMDA receptor-mediated neurotransmission. This model has been increasingly adopted and is now considered to be one of the useful models for both etiological conceptualization of schizophrenia and new treatment development (13-17).

NMDA receptors are composed of a combination of distinct subunits termed NR1, NR2 and NR3. Multiple splice variants of the NR1 subunit have been described, along with multiple subforms of the NR2 subunit termed NR2A-D. All functional NMDA receptors possess one or more NR1 subunits. In addition, most receptors contain a combination of NR2 subunits, with NR2A and NR2B subunits dominating in adult brain. Different combinations of subunits confer different properties to the receptors. It has been suggested that NR2A and NR2B subunit-containing receptors may have differential roles in psychogenesis (18, 19), although others have suggested that combined blockade is needed (20).

Because of the lack of subunit specific drugs, it is difficult to determine the involvement of the different subunits types in the pathophysiology of schizophrenia and it remains possible that subtype-selective intervention will prove preferable to generalized modulation across NMDA receptor subtypes.

Symptom Patterns Following NMDA Antagonist Administration

In initial studies with PCP and ketamine in the early 1960s, researchers noted that both agents produced what would now be considered positive, negative and cognitive symptoms of schizophrenia (12). At the time, however, no formal rating scales were used. Recent stud6

ies with ketamine, however, have documented significant increases not only in positive symptoms, but also in negative and cognitive symptoms (21-23). Levels of symptoms during acute ketamine challenge, moreover, tend to show a similar pattern across factors as they do in schizophrenia. When patients with schizophrenia are exposed to ketamine, they also show increases in positive symptoms, as well as negative symptoms (24, 25), suggesting that NMDA antagonists affect a brain system that is already vulnerable in schizophrenia.

Cognitive Deficits Following NMDA Antagonist Treatment

As with symptoms, initial studies conducted with PCP in the early 1960s also showed cognitive deficits that are highly reminiscent of schizophrenia (12). Studies conducted with ketamine over the last 15 years have further confirmed and extended these findings. Deficits have been observed across widespread neuropsychological domains including working memory, response inhibition and executive processing (23, 26, 27). Ketamine infusion also reproduces both the severity and type of thought disorder seen in schizophrenia with both, for example, being associated with high levels of poverty of speech, circumstantiality and loss of goal, and relatively low levels of distractive or stilted speech or paraphasias (28). Given the importance of neurocognitive dysfunction to the conceptualization of schizophrenia, these findings support the etiological involvement of NMDA dysfunction in the pathophysiology of schizophrenia.

As opposed to ketamine, administration of dopaminergic agonists such as amphetamine does not reproduce the pattern of deficit observed in schizophrenia. For example, in one recent study that directly compared effects of amphetamine and ketamine in normal volunteers, both ketamine and amphetamine induced positive symptoms and conceptual disorganization. However, only ketamine produced perceptual changes, concrete ideation or negative symptoms. Further, only ketamine induced schizophrenia-like disruptions in delayed recall. Finally, amphetamine did not induce working memory disturbances, and it significantly reversed ketamine-induced disruptions. These findings suggest that augmentation, rather than blockade, of frontal dopaminergic systems may be beneficial in schizophrenia (26). In schizophrenia, amphetamine treatment does not further impair cognition and may in fact lead to cognitive improvement in schizophrenia (29). These findings therefore suggest greater involvement of NMDA than

Daniel C. Javitt

dopamine receptors in the pathophysiology of cognitive impairment in schizophrenia.

Further evidence for a specific involvement of NMDA receptors in schizophrenia comes from finegrained analysis of patterns of neurocognitive dysfunction. Neurocognitive deficits in schizophrenia appear generalized when viewed at a “molar” level, such as the level of a cognitive “domain.” However, when viewed at a more detailed, “molecular” level, fine-grained differences between different processes do emerge. For example, patients with schizophrenia show reduced ability to learn new information, but intact ability to retain information once it has been learned. This pattern differs from the “amnestic” syndrome that results from bilateral hippocampal damage (2), but is highly similar to effects seen following administration of NMDA antagonists (30). Overall, the pattern of cognitive dysfunction in schizophrenia follows closely the pattern observed following administration of NMDA antagonists across a variety of domains, suggesting that NMDA dysfunction may be seen as a parsimonious model of schizophrenia.

NMDA Dysfunction and Sensory Processing Impairment

Another key difference between dopaminergic and NMDA models of schizophrenia is predicted involvement of sensory processing. NMDA receptors are widely distributed throughout cortex. In contrast, dopaminergic innervation is much more circumscribed, with relatively sparse innervations of primary sensory cortex (31, 32). An important issue, therefore, is whether information processing deficits in schizophrenia are seen only in higher order cortical regions, such as prefrontal cortex, or if they are observed throughout brain and involve even primary sensory regions. Studies have been performed primarily in auditory and visual systems, although schizophrenia is known to affect other sensory processes such as weight discrimination (33) and other somatosensory processes (34).

Auditory deficits in schizophrenia. Deficits in auditory processing have been investigated using both behavioral and neurophysiological measures. Behaviorally, patients show deficits in matching of tones following brief delay (35), suggesting dysfunction of the auditory sensory memory system. This is a heuristically valuable paradigm, as underlying anatomical substrates have been well characterized in primate and human models.

Lesions of auditory sensory cortex, located in superior

temporal lobe, produce increases in tone matching threshold without affecting disruptive effects of distracting stimuli. In contrast, lesions of prefrontal cortex increase distractibility without affecting thresholds (36).

In patients with schizophrenia, increased thresholds are observed with no accompanying increase in susceptibility to either visual (37) or auditory distraction (38, 39).

Further, when equated for performance at short interstimulus interval (<1 s), patients show equivalent decay with increasing interval (39), suggesting normal retention within the sensory memory system. These behavioral findings thus suggest dysfunctional information processing at the level of auditory sensory cortex.

Auditory function in schizophrenia has also been assessed with event-related potentials (ERP). One of the most informative potentials has been mismatch negativity (MMN). MMN is elicited by infrequent changes in nature or pattern of repetitive auditory stimulation.

Deviant stimuli may differ from standards in a number of stimulus dimensions, including pitch, duration, intensity or location. Generators for MMN have been mapped to auditory sensory cortex in the region of Heschl’s gyrus (40). Deficits in MMN generation were first demonstrated in schizophrenia over 10 years ago and currently represent one of the best replicated neurophysiological findings in schizophrenia (41).

Schizophrenia-like deficits in MMN generation can be induced by local infusion of NMDA antagonists into primate auditory cortex (40) and by systemic administration of NMDA antagonists in healthy volunteers (23) , suggesting that such deficits may index NMDA dysfunction at the level of auditory cortex. In contrast, MMN is not modulated via a variety of other psychoactive agents, including the 5-HT2A agonist psilocybin (42) and the D1/D2 agonists bromocriptine and pergolide (43), suggesting relative specificity of the NMDA antagonist psychotomimetic effect.

More recent studies have investigated consequences of elevated tone matching thresholds to more complex forms of information processing dysfunction. Patients with schizophrenia, for example, show well-established deficits in ability to determine emotion based upon vocal modulation (prosody), which are thought to be rate-limiting in terms of functional outcome (44). The etiology of such deficits has been poorly understood, as patients show normal emotional responses to happy or sad events, and show intact internal representation of emotion (45), suggesting that failure to detect emotion may be related to underlying failure to utilize sensory cues.

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GLUTAMATERGIC THEORIES OF SCHIZOPHRENIA

An initial study of prosodic detection in schizophrenia evaluated the relationship between tone matching performance on the one hand and both auditory and visual emotion detection on the other. Deficits in auditory perceptual performance (tone matching) strongly predicted deficits in auditory, but not visual, emotion detection. Further, although patients showed deficits in both auditory and visual emotion detection, the two sets of deficits were statistically unrelated, suggesting that deficits clustered within, rather than across modalities. These results thus strongly supported the hypothesis that deficits in “social cognition” in schizophrenia, rather than reflecting deficits in the conceptualization of emotion instead reflect upward consequences of the effects of underlying disturbances in underlying tone matching ability (46). A subsequent study demonstrated a similar relationship between tone matching ability and ability to detect attitudinal prosody (sarcasm) (47), as well as non-affective prosody such as ability to differentiate questions from statements (semantic prosody) (48). Further, severity of deficit across individuals correlated highly with reduced structural integrity within auditory white matter pathways at the level of auditory cortex (48). When sensory performance has been evaluated as a function of stimulus properties deficits in emotional detection have been found to involve particularly those types of emotional distinctions that depend upon differentiation of pitch (49). Further, in addition to showing deficits in identifying emotions, patients show deficits in differentiating between emotional intensities, also consistent with inability to process changes in pitch that differentiate emotions (49). Taken together, these findings suggest that basic deficits in NMDA receptormediated neurotransmission at the level of auditory sensory cortex lead to sensory level disturbances which, in turn, upward generalize to produce disturbances in high level processes such as ability to interpret toneof-voice.

Visual processing deficits. Similar studies have now been performed investigating consequences of NMDA dysfunction in the early visual system. The early visual system consists of discrete magnocellular and parvocellular pathways that differ in characteristics and function.

The magnocellular pathway provides rapid transmission of low-resolution information to cortex, in order to prime attentional systems and “frame” the overall visual scene. The parvocellular pathway, in contrast, provides slower, higher resolution information to fill in scene details (50). NMDA receptors are located at mul8

tiple levels of the early visual system, including retina, lateral geniculate nucleus (LGN) and primary cortex.

The magnocellular system, in particular, functions in a non-linear gain mode that is dependent upon NMDA receptor-mediated neurotransmission. Administration of NMDA antagonists to cat LGN produces a characteristic reduction in gain that is also observed in schizophrenia (51).

To date, deficits in visual processing have been demonstrated in schizophrenia using both steady-state (51-53) and transient (54-56) visual evoked potential approaches. Further, deficits in early visual processing produce subsequent impairments on higher order processes such as object identification (57), motion processing (58) and reading (59). Further, change in the physical properties of stimuli to make them more tractable to visual analysis leads to significant improvement in performance in such high-level tasks as the AX-version of the continuous performance task (60) or Wisconsin Card Sorting Test (61). Thus, as in the auditory system, basic deficits in NMDA function within subcortical and cortical systems lead to breakdown of basic sensory discrimination abilities, which, in turn, produce complex patterns of higher level cognitive disturbances in schizophrenia.

Glutamate-dopamine Glutamate-GABA Interactions

Finally, NMDA dysfunction may also account for both the impaired dopaminergic regulation and the impaired GABAergic neurotransmission that has been documented in schizophrenia. Dopaminergic dysfunction has been studied most extensively using positron emission (PET) or single photon emission (SPECT) markers of response to amphetamine. In such studies, D2 agonists are tagged with appropriate radionuclides (e.g., [14C], [123I]) and pattern of displacement is evaluated following amphetamine administration. Across cohorts, patients with acute schizophrenia show enhanced striatal dopamine release to amphetamine challenge, consistent with presumed dysregulation of subcortical dopamine circuits (62).

Deficits similar to those observed in schizophrenia are observed in normal volunteers undergoing ketamine infusion (63), and in rodents treated subchronically (64, 65) with NMDA receptor antagonists, suggesting that dopaminergic dysregulation in schizophrenia may be “downstream” of a primary deficit in NMDA function.

Similarly, NMDA antagonists alter the random firing rate of rodent prefrontal neurons while decreasing burst

Daniel C. Javitt

firing (66), also supporting the concept that deficits in NMDA transmission may lead to the widely cited disturbances in prefrontal function in schizophrenia.

Changes in GABAergic neurotransmission have also been increasingly well documented over recent years, with studies showing reduced parvalbumin and GAD67 expression, particularly in prefrontal cortex (67-71) and hippocampus (72-74). Similar effects are seen in both rodents (75-79) and monkeys (80) treated with NMDA antagonists such as PCP, as well as in cell culture (81).

GABAergic dysfunction in PFC may be directly linked to well-documented deficits in working memory function, and may therefore represent an appropriate target of pharmacological intervention (82). Nevertheless, etiologically such abnormalities may reflect downstream effects of primary deficits in NMDA receptor-mediated neurotransmission.

Clinical Studies with NMDA Agonists Given the ability of NMDA receptor antagonists to induce symptoms that closely resemble those of schizophrenia, a critical issue is whether treatment approaches based upon glutamatergic and NMDA models can lead to new treatment approaches. Over the past decade, several new treatment strategies have been proposed. First, direct and indirect approaches have targeted the glycine modulatory site of the NMDA receptor complex. Direct agonists have included treatment with the naturally occurring amino acids glycine and D-serine, which serve as endogenous modulators of NMDA receptors in vivo, as well the anti-tuberculosis drug D-cycloserine, which fortuitously cross-reacts with the NMDA/glycine site (83). These agents have proven effective in several Figure 2. Schematic Model of Synaptic Glycine Regulation by Glycine Transport Inhibitors.

Table 1: Preclinical Paradigms of Relevance to Schizophrenia in Which Glycine Transport Inhibitors Have Proven Effective Test Measure

Reference

Inhibition of phencyclidine (PCP)-induced hyperactivity in vivo

[88, 123, 124]

Inhibition of striatal dopamine release in vitro

[125]

Potentiation of hippocampal NMDA responses in vitro

[126]

Potentiation of prefrontal/hippocampal NMDA responses in vitro

[127-129]

Normalization of prepulse inhibition (PPI) deficits in rodents

[128-130]

Normalization of PCP-induced increases in amphetaminestimulated dopamine release

[84]

Reversal of locomotor hypersensitivity to amphetamine neonatally PCP-treated rats

[129]

Elevation of CSF glycine levels

[131]

preclinical models, including reversal of PCP effects in both rodents (84, 85) and primates (86).

A â&#x20AC;&#x153;second generationâ&#x20AC;? approach to this problem has been the use of glycine type I (GlyT1) transport inhibitors (GTIs). Rather than serving as direct glycine precursors, these compounds increase glycine levels in brain by preventing glycine removal from the synaptic cleft, leading to endogenous increases in CSF glycine levels (87) (Figure 2). An initial study with glycyldodeclamide, a relatively low affinity agent, demonstrated significant reversal of PCP-induced hyperactivity in rodents (88, 89). Since then, high affinity GTIs have been synthesized by several pharmaceutical companies, and have shown to be effective in multiple animal models (Table 1). Several of these compounds are currently in early-stage clinical trials, with results expected over the next several years.

Two other treatment strategies have been proposed.

First, in addition to the glycine modulatory site, NMDA receptors contain a redox-sensitive site that is modulated by the oxidized form of glutathione (GSH) (90, 91) (Figure 1). Schizophrenia has also been shown to be associated with reduced levels of GSH (92-94), leading to potential dysfunction of NMDA receptors (95). Early studies testing this mechanism have utilized N-acetylcysteine, a glutathione precursor, as a potential psychopharmacological agent.

Second, based upon the observation that NMDA blockade leads to rebound increases in glutamate release that may themselves be pathological (96), it has been proposed that compounds that inhibit presynaptic glutamate release may also be therapeutic (97). Examples of such compounds include the anti-epilepsy drug 9

GLUTAMATERGIC THEORIES OF SCHIZOPHRENIA

Results of Clinical Studies

The most studies to date have been performed with NMDA agonists, primarily because several of the agents used have been natural compounds, and so it has not been necessary to wait for structure activity optimization or preclinical toxicity testing. Nevertheless, this approach is also a limitation, as permeability of these agents may be limited, and delivering optimal doses may therefore be impossible. Nevertheless, positive studies with these 10

compounds have provided proof-of-concept for development of compounds with higher affinity and specificity.

Studies with naturally occurring compounds to date have primarily used glycine, administered at a dose of up to 800 mg/kg (approx. 60 g/d) (109-112); D-serine, administered at a dose of 30 mg/kg (approx. 2.1 g/d) (113, 114) or D-alanine administered at a dose of 100 mg/kg (115); and sarcosine, administered at a dose of 30 mg/kg (approx. 2.1 g/d) (116, 117). For glycine, this represents the highest practical dose because of the quantity of amino acid needed to significantly increase brain glycine levels. For other compounds, formal dose findings studies have not been performed, and maximum tolerated doses are presently unknown.

Across all studies utilizing full agonists in combination with either typical or newer atypical antipsychotic drugs, NMDA agonists have been found to produce an approximately 15% improvement in negative symptoms, along with significant changes in positive and cognitive Figure 3. Summary of clinical trials performed to date with full NMDA agonists combined with antipsychotics other than clozapine. Studies were conducted using the amino acid glycine at doses of 0.4-0.8 g/kg (30-60 g/d) unless otherwise indicated.

Further details about individual studies are provided in (83).

CONSIST refers to The Cognitive and Negative Symptoms in Schizophrenia Trial (132). Statistics were calculated as weighted average of % change scores for negative symptoms, across trials.

50

Treated Control

40

Weighted d = .47 (mod), p <.0001

30 20 10

Weighted average

Buchanan et al., 2007 (glycine)

Tsai et al., 2004 (sarcosine) Lane et al., 2008 (sarcosine)

Tsai et al., 2001 (D-serine) Heresco et al., 2005 (D-serine) Tsai et al., 2006 (D-alanine)

-10

Heresco et al., 2005 (glycine)

0 Javitt et al. 1994 (glycine) Heresco et al., 1999 (glycine) Javitt et al., 2001 (glycine)

lamotrigine and agonists of metabotropic glutamate type 2/3 (mGluR2/3) receptors, which are localized to presynaptic glutamate terminals in prefrontal cortex. mGluR2/3 agonists have been shown to be effective in reversing behavioral effects of NMDA antagonists in rodent models (98), supporting the potential efficacy of these compounds as novel antipsychotic agents. In addition, both lamotrigine (99) and mGluR 2/3 agonists (100) have also been shown to reverse clinical effects of ketamine during acute challenge in normal volunteers, further supporting the applicability of basic models to humans. In general, therefore, as the NMDA model reaches its second decade, the base of treatment development based upon glutamatergic theories continues to increase.

Other metabotropic ligands, including mGluR5 (101, 102) and mGluR8 (103) agonists, have also been proposed as potential treatments for schizophrenia, based upon their ability to modulate NMDA receptor-mediated neurotransmission (104). Finally, N-acetylaspartylglutamate (NAAG) may be an endogenous ligand for mGlu2/3 receptors in CNS. NAAG is broken down by NAAG peptidase (glutamate carboxypeptidase II) (105). Compounds that inhibit NAAG peptidase, such as an experimental inhibitor termed ZJ43, would therefore lead to increased mGlu2/3 occupancy. This compound has been tested preclinically and shown to inhibit PCP- and MK-801-induced behaviors in animals, consistent with an effect on NMDA receptor-mediated neurotransmission (106, 107).

Finally, some authors have suggested that NMDA antagonists may be beneficial, based upon concepts that cognitive deficits in schizophrenia may result from hyper-glutamatergic neurotoxicity (13). Examples of compounds that have been considered based upon this hypothesis are AMPA antagonists and the antiAlzheimers disease drug memantine. To date, however, clinical experience with NMDA antagonists has not been encouraging (108).

symptoms in some but not all studies (83) (see figure 3). One study has evaluated effects of glycine in a limited number of individuals showing prodromal symptoms of schizophrenia. In that study, large effect-size improvement was observed, including early remission in three of 10 subjects (118). These data, if confirmed, would indicate that NMDA agonists might have a primary role in the earliest stages of schizophrenia psychosis, with potential impact across symptomatic domains.

Daniel C. Javitt

Studies of other mechanisms also show suggestive findings. Thus, one study of N-acetylcysteine, a precursor of glutathione, produced significant improvement in PANSS total and negative symptoms in schizophrenia (119), along with improvement in generation of MMN, which may serve as a biomarker of NMDA dysfunction (120). Two small studies with lamotrigine showed suggestive results (121, 122), although a subsequent multicenter double-blind study was negative. To date, one phase II study with the oral mGluR2/3 agonist prodrug LY2140023, used as monotherapy in acutely relapsing subjects, showed clinical efficacy similar to that of olanzapine with markedly reduced incidence of metabolic side effects. Although this study requires replication, it is encouraging with regard to overall efficacy of glutamatergic approaches.

Summary Glutamatergic models of schizophrenia were first proposed over two decades ago, based upon the effects of the agents PCP and ketamine, which were shown to induce their unique psychotomimetic effects by blocking neurotransmission at NMDA-type glutamate receptors.

Since that time, glutamatergic models have been strongly supported by NMDA antagonists studies in animals, as well as ketamine challenge studies in humans. Over that time, potential molecular contributors to NMDA dysfunction have been increasingly documented.

New treatment approaches based upon glutamatergic approaches are only now reaching the clinic, and will serve to further elucidate and refine these models over upcoming years. Whether glutamatergic approaches will eventually supplant dopamine antagonists for treatment of positive symptoms remains to be determined.

Nevertheless, glutamatergic approaches offer particular hope for treatment of negative symptoms and cognitive deficits in schizophrenia, and thus for improvement of the clinical situation of thousands of patients in Israel and millions of patients worldwide.

Acknowledgements: Preparation of this manuscript was supported in part by USPHS grants R01 DA03383, R37 MH49334, and the NYC Conte Center for Schizophrenia Research (P50 MH060450).

References 1. Woodberry KA, Giuliano AJ, Seidman LJ. Premorbid IQ in schizophrenia: A meta-analytic review. Am J Psychiatry 2008;165:579-587.

2. Bilder RM, Goldman RS, Robinson D, Reiter G, Bell L, Bates JA, et al. Neuropsychology of first-episode schizophrenia: Initial characterization and clinical correlates.

Am J Psychiatry 2000; 157:549-559.

3. Jones P, Rodgers B, Murray R, Marmot M. Child developmental risk factors for adult schizophrenia in the British 1946 birth cohort. Lancet 1994;344:1398-1402.

4. Fuller R, Nopoulos P, Arndt S, O,Leary D, Ho BC, Andreasen NC. Longitudinal assessment of premorbid cognitive functioning in patients with schizophrenia through examination of standardized scholastic test performance. Am J Psychiatry 2002;159:1183-1189.

5. Ho BC, Andreasen NC, Nopoulos P, Fuller R, Arndt S, Cadoret RJ. Secondary prevention of schizophrenia: Utility of standardized scholastic tests in early identification. Ann Clin Psychiatry 2005;17:11-18.

6. Lencz T, Smith CW, McLaughlin D, Auther A, Nakayama E, Hovey L, et al. Generalized and specific neurocognitive deficits in prodromal schizophrenia. Biol Psychiatry 2006;59:863-871.

7. Reichenberg A, Weiser M, Rapp MA, Rabinowitz J, Caspi A, Schmeidler J, et al. Elaboration on premorbid intellectual performance in schizophrenia: Premorbid intellectual decline and risk for schizophrenia. Arch Gen Psychiatry 2005;62:1297-1304.

8. Caspi A, Reichenberg A, Weiser M, Rabinowitz J, Kaplan Z, Knobler H, et al. Cognitive performance in schizophrenia patients assessed before and following the first psychotic episode. Schizophr Res 2003;65:87-94.

9. Carlsson A. The current status of the dopamine hypothesis of schizophrenia. Neuropsychopharmacology 1988;1:179-186.

10. Davis KL, Kahn RS, Ko G, Davidson M. Dopamine in schizophrenia: A review and reconceptualization. Am J Psychiatry 1991;148:1474-1486.

11. Javitt DC. Negative schizophrenic symptomatology and the PCP (phencyclidine) model of schizophrenia. Hillside J Clin Psychiatry 1987;9:12-35.

12. Javitt DC, Zukin SR. Recent advances in the phencyclidine model of schizophrenia. Am J Psychiatry 1991;148: 1301-1308.

13. Olney JW, Farber NB. Glutamate receptor dysfunction and schizophrenia. Arch Gen Psychiatry 1995;52:998-1007.

14. Coyle JT, Tsai G. The NMDA receptor glycine modulatory site: A therapeutic target for improving cognition and reducing negative symptoms in schizophrenia.

Psychopharmacology (Berl) 2004;174:32-38.

15. Moghaddam B. Bringing order to the glutamate chaos in schizophrenia. Neuron 2003;40:881-884.

16. Heresco-Levy U. Glutamatergic neurotransmission mod11

GLUTAMATERGIC THEORIES OF SCHIZOPHRENIA

ulators as emerging new drugs for schizophrenia. Expert Opin Emerg Drugs 2005;10:827-844.

17. Javitt DC. Phenomenology, aetiology and treatment of schizophrenia. Novartis Foundation Symposium 2008;289:4-16; discussion 7-22, 87-93.

18. Lee J, Rajakumar N. Role of NR2B-containing N-methylD-aspartate receptors in haloperidol-induced c-Fos expression in the striatum and nucleus accumbens.

Neuroscience 2003;122:739-745.

19. Higgins GA, Ballard TM, Huwyler J, Kemp JA, Gill R. Evaluation of the NR2B-selective NMDA receptor antagonist Ro 63-1908 on rodent behaviour: Evidence for an involvement of NR2B NMDA receptors in response inhibition. Neuropharmacology 2003;44:324-341.

20. Spooren W, Mombereau C, Maco M, Gill R, Kemp JA, Ozmen L, et al. Pharmacological and genetic evidence indicates that combined inhibition of NR2A and NR2B subunit containing NMDA receptors is required to disrupt prepulse inhibition. Psychopharmacology (Berl) 2004;175:99-105.

21. Krystal JH, Karper LP, Seibyl JP, Freeman GK, Delaney R, Bremner JD, et al. Subanesthetic effects of the noncompetitive NMDA antagonist, ketamine, in humans.

Psychotomimetic, perceptual, cognitive, and neuroendocrine responses. Arch Gen Psychiatry 1994;51:199-214.

22. Lahti AC, Koffel B, LaPorte D, Tamminga CA. Subanesthetic doses of ketamine stimulate psychosis in schizophrenia.

Neuropsychopharmacology 1995;13:9-19.

23. Umbricht D, Schmid L, Koller R, Vollenweider FX, Hell D, Javitt DC. Ketamine-induced deficits in auditory and visual context-dependent processing in healthy volunteers: Implications for models of cognitive deficits in schizophrenia. Arch Gen Psychiatry 2000;57:1139-1147.

24. Lahti AC, Weiler MA, Tamara Michaelidis BA, Parwani A, Tamminga CA. Effects of ketamine in normal and schizophrenic volunteers. Neuropsychopharmacology 2001;25:455-467.

25. Malhotra AK, Pinals DA, Adler CM, Elman I, Clifton A, Pickar D, et al. Ketamine-induced exacerbation of psychotic symptoms and cognitive impairment in neuroleptic-free schizophrenics. Neuropsychopharmacology 1997;17:141-150.

26. Krystal JH, Perry EB, Jr., Gueorguieva R, Belger A, Madonick SH, Abi-Dargham A, et al. Comparative and interactive human psychopharmacologic effects of ketamine and amphetamine: Implications for glutamatergic and dopaminergic model psychoses and cognitive function. Arch Gen Psychiatry 2005;62:985-994.

27. Malhotra AK, Pinals DA, Weingartner H, Sirocco K, Missar CD, Pickar D, et al. NMDA receptor function and human cognition: The effects of ketamine in healthy vol12

unteers. Neuropsychopharmacology 1996;14:301-307.

28. Adler CM, Malhotra AK, Elman I, Goldberg T, Egan M, Pickar D, et al. Comparison of ketamine-induced thought disorder in healthy volunteers and thought disorder in schizophrenia. Am J Psychiatry 1999;156:1646-1649.

29. Barch DM, Carter CS. Amphetamine improves cognitive function in medicated individuals with schizophrenia and in healthy volunteers. Schizophr Res 20051;77:43-58.

30. Parwani A, Weiler MA, Blaxton TA, Warfel D, Hardin M, Frey K, et al. The effects of a subanesthetic dose of ketamine on verbal memory in normal volunteers.

Psychopharmacology (Berl) 2005;183:265-274.

31. Lewis DA, Campbell MJ, Foote SL, Goldstein M, Morrison JH. The distribution of tyrosine hydroxylase-immunoreaction fibers in primate neocortex is widespread but regionally specific. J Neurosci 1987;7:279-290.

32. Lewis DA, Sesack SR, Levey AI, Rosenberg DR. Dopamine axons in primate prefrontal cortex: Specificity of distribution, synaptic targets, and development. Adv Pharmacol 1998;42:703-706.

33. Javitt DC, Liederman E, Cienfuegos A, Shelley AM. Panmodal processing imprecision as a basis for dysfunction of transient memory storage systems in schizophrenia. Schizophr Bull 1999;25:763-775.

34. Chang BP, Lenzenweger MF. Somatosensory processing and schizophrenia liability: proprioception, exteroceptive sensitivity, and graphesthesia performance in the biological relatives of schizophrenia patients. J Abnorm Psychol 2005;114:85-95.

35. Strous RD, Cowan N, Ritter W, Javitt DC. Auditory sensory (“echoic”) memory dysfunction in schizophrenia.

Am J Psychiatry 1995;152:1517-1519.

36. Rabinowicz EF, Silipo G, Goldman R, Javitt DC. Auditory sensory dysfunction in schizophrenia: Imprecision or distractibility? Arch Gen Psychiatry 2000;57:1149-1155.

37. Javitt DC, Strous RD, Grochowski S, Ritter W, Cowan N. Impaired precision, but normal retention, of auditory sensory (“echoic”) memory information in schizophrenia. J Abnorm Psychol 1997;106:315-324.

38. Rabinowitz J, Reichenberg A, Weiser M, Mark M, Kaplan Z, Davidson M. Cognitive and behavioural functioning in men with schizophrenia both before and shortly after first admission to hospital. Cross-sectional analysis. Br J Psychiatry 2000;177:26-32.

39. March L, Cienfuegos A, Goldbloom L, Ritter W, Cowan N, Javitt DC. Normal time course of auditory recognition in schizophrenia, despite impaired precision of the auditory sensory (“echoic”) memory code. J Abnorm Psychol 1999;108:69-75.

Daniel C. Javitt

40. Javitt DC. Intracortical mechanisms of mismatch negativity dysfunction in schizophrenia. Audiol Neurootol 2000;5:207-215.

A, Bockbrader MA, et al. Steady state visual evoked potential abnormalities in schizophrenia. Clin Neurophysiol 2005;116:614-624.

41. Umbricht D, Krljes S. Mismatch negativity in schizophrenia: A meta-analysis. Schizophr Res 2005;76:1-23.

54. Foxe JJ, Doniger GM, Javitt DC. Early visual processing deficits in schizophrenia: Impaired P1 generation revealed by high-density electrical mapping. Neuroreport 2001;12:3815-3820.

42. Umbricht D, Vollenweider FX, Schmid L, Grubel C, Skrabo A, Huber T, et al. Effects of the 5-HT2A agonist psilocybin on mismatch negativity generation and AXcontinuous performance task: Implications for the neuropharmacology of cognitive deficits in schizophrenia.

Neuropsychopharmacology 2003;28:170-181.

43. Leung S, Croft RJ, Baldeweg T, Nathan PJ. Acute dopamine D(1) and D(2) receptor stimulation does not modulate mismatch negativity (MMN) in healthy human subjects. Psychopharmacology (Berl) 2007;194:443-451.

44. Brekke J, Kay DD, Lee KS, Green MF. Biosocial pathways to functional outcome in schizophrenia. Schizophr Res 2005;80:213-225.

45. Kring AM, Barrett LF, Gard DE. On the broad applicability of the affective circumplex: representations of affective knowledge among schizophrenia patients. Psychol Sci 2003;14:207-214.

46. Leitman DI, Foxe JJ, Butler PD, Saperstein A, Revheim N, Javitt DC. Sensory contributions to impaired prosodic processing in schizophrenia. Biol Psychiatry 2005;58:56-61.

47. Leitman DI, Ziwich R, Pasternak R, Javitt DC. Theory of Mind (ToM) and counterfactuality deficits in schizophrenia: Misperception or misinterpretation? Psychol Med 2006;36:1075-1083.

48. Leitman DI, Hoptman MJ, Foxe JJ, Saccente E, Wylie GR, Nierenberg J, et al. The neural substrates of impaired prosodic detection in schizophrenia and its sensorial antecedents. Am J Psychiatry 2007;164:474-482.

49. Leitman Di, Laukka P, Juslin PN, Saccente E, Butler P, Javitt DC. Getting the cue: Sensory contributions to auditory emotion recognition impairments in schizophrenia.

Schizophr Bull 2008; Sept. 23 [Epub ahead of print].

50. Chen CM, Lakatos P, Shah AS, Mehta AD, Givre SJ, Javitt DC, et al. Functional anatomy and interaction of fast and slow visual pathways in macaque monkeys. Cereb Cortex 2007;17:1561-1569.

51. Butler PD, Zemon V, Schechter I, Saperstein AM, Hoptman MJ, Lim KO, et al. Early-stage visual processing and cortical amplification deficits in schizophrenia. Arch Gen Psychiatry 2005;62:495-504.

52. Butler PD, Schechter I, Zemon V, Schwartz SG, Greenstein VC, Gordon J, et al. Dysfunction of early-stage visual processing in schizophrenia. Am J Psychiatry 2001;158:1126-1133.

53. Krishnan GP, Vohs JL, Hetrick WP, Carroll CA, Shekhar

55. Schechter I, Butler PD, Zemon VM, Revheim N, Saperstein AM, Jalbrzikowski M, et al. Impairments in generation of early-stage transient visual evoked potentials to magnoand parvocellular-selective stimuli in schizophrenia. Clin Neurophysiol 2005; 116:2204-2215.

56. Butler PD, Martinez A, Foxe JJ, Kim D, Zemon V, Silipo G, et al. Subcortical visual dysfunction in schizophrenia drives secondary cortical impairments. Brain 2007;130:417-430.

57. Doniger GM, Foxe JJ, Murray MM, Higgins BA, Javitt DC. Impaired visual object recognition and dorsal/ ventral stream interaction in schizophrenia. Arch Gen Psychiatry 2002;59:1011-1020.

58. Kim D, Wylie G, Pasternak R, Butler PD, Javitt DC. Magnocellular contributions to impaired motion processing in schizophrenia. Schizophr Res 2006;82:1-8.

59. Revheim N, Butler PD, Schechter I, Jalbrzikowski M, Silipo G, Javitt DC. Reading impairment and visual processing deficits in schizophrenia. Schizophr Res 2006;87:238-245.

60. Javitt DC, Rabinowicz E, Silipo G, Dias EC. Encoding vs. retention: Differential effects of cue manipulation on working memory performance in schizophrenia.

Schizophr Res 2007;91:159-168.

61. Kantrowitz JT, Revheim N, Pasternak R, Silipo G, Javitt DC. Itî&#x2C6;&#x20AC;s all in the cards: Effects of stimulus manipulation on WCST performance in schizophrenia. Psychiatry Res 2009; 168: 198-204.

62. Laruelle M, Abi-Dargham A, Gil R, Kegeles L, Innis R. Increased dopamine transmission in schizophrenia: Relationship to illness phases. Biol Psychiatry 1999;46:56-72.

63. Kegeles LS, Abi-Dargham A, Zea-Ponce Y, RodenhiserHill J, Mann JJ, Van Heertum RL, et al. Modulation of amphetamine-induced striatal dopamine release by ketamine in humans: Implications for schizophrenia. Biol Psychiatry 2000;48:627-640.

64. Balla A, Koneru R, Smiley J, Sershen H, Javitt DC. Continuous phencyclidine treatment induces schizophrenia-like hyperreactivity of striatal dopamine release.

Neuropsychopharmacology 2001;25:157-164.

65. Balla A, Sershen H, Serra M, Koneru R, Javitt DC. Subchronic continuous phencyclidine administration potentiates amphetamine-induced frontal cortex dopamine 13

GLUTAMATERGIC THEORIES OF SCHIZOPHRENIA

release. Neuropsychopharmacology 2003;28:34-44.

66. Jackson ME, Homayoun H, Moghaddam B. NMDA receptor hypofunction produces concomitant firing rate potentiation and burst activity reduction in the prefrontal cortex. Proc Natl Acad Sci USA 2004;101:8467-8472.

67. Beasley CL, Reynolds GP. Parvalbumin-immunoreactive neurons are reduced in the prefrontal cortex of schizophrenics. Schizophr Res 1997;24:349-355.

68. Woo TU, Miller JL, Lewis DA. Schizophrenia and the parvalbumin-containing class of cortical local circuit neurons. Am J Psychiatry 1997;154:1013-1015.

69. Reynolds GP, Beasley CL. GABAergic neuronal subtypes in the human frontal cortex-development and deficits in schizophrenia. J Chem Neuroanat 2001;22:95-100.

70. Torrey EF, Barci BM, Webster MJ, Bartko JJ, MeadorWoodruff JH, Knable MB. Neurochemical markers for schizophrenia, bipolar disorder, and major depression in postmortem brains. Biol Psychiatry 2005;57:252-260.

71. Lewis DA, Moghaddam B. Cognitive dysfunction in schizophrenia: convergence of gamma-aminobutyric acid and glutamate alterations. Arch Neurol 2006;63:1372-1376.

72. Zhang ZJ, Reynolds GP. A selective decrease in the relative density of parvalbumin-immunoreactive neurons in the hippocampus in schizophrenia. Schizophr Res 2002;55:1-10.

73. Knable MB, Barci BM, Webster MJ, Meador-Woodruff J, Torrey EF. Molecular abnormalities of the hippocampus in severe psychiatric illness: postmortem findings from the Stanley Neuropathology Consortium. Mol Psychiatry 2004;9:544, 609-620.

74. Braun I, Genius J, Grunze H, Bender A, Moller HJ, Rujescu D. Alterations of hippocampal and prefrontal GABAergic interneurons in an animal model of psychosis induced by NMDA receptor antagonism. Schizophr Res 2007;97:254263.

75. Cunningham MO, Hunt J, Middleton S, LeBeau FE, Gillies MJ, Davies CH, et al. Region-specific reduction in entorhinal gamma oscillations and parvalbumin-immunoreactive neurons in animal models of psychiatric illness. J Neurosci 2006;26:2767-2776.

in the hippocampal expression of parvalbumin, neuronal nitric oxide synthase and cFOS similar to those found in human schizophrenia. Neuroscience 2004;126:591-598.

79. Reynolds GP, Abdul-Monim Z, Neill JC, Zhang ZJ. Calcium binding protein markers of GABA deficits in schizophrenia-postmortem studies and animal models.

Neurotox Res 2004;6:57-61.

80. Morrow BA, Elsworth JD, Roth RH. Repeated phencyclidine in monkeys results in loss of parvalbumin-containing axo-axonic projections in the prefrontal cortex.

Psychopharmacology (Berl) 2007;192:283-290.

81. Kinney JW, Davis CN, Tabarean I, Conti B, Bartfai T, Behrens MM. A specific role for NR2A-containing NMDA receptors in the maintenance of parvalbumin and GAD67 immunoreactivity in cultured interneurons.

J Neurosci 2006;26:1604-1615.

82. Lewis DA, Volk DW, Hashimoto T. Selective alterations in prefrontal cortical GABA neurotransmission in schizophrenia: A novel target for the treatment of working memory dysfunction. Psychopharmacology (Berl) 2004;174:143-150.

83. Javitt DC. Is the glycine site half saturated or half unsaturated? Effects of glutamatergic drugs in schizophrenia patients. Curr Opin Psychiatry 2006;19:151-157.

84. Javitt DC, Balla A, Burch S, Suckow R, Xie S, Sershen H. Reversal of phencyclidine-induced dopaminergic dysregulation by N-methyl-D-aspartate receptor/glycine-site agonists. Neuropsychopharmacology 2004;29:300-307.

85. Contreras PC. D-serine antagonized phencyclidineand MK-801-induced stereotyped behavior and ataxia.

Neuropharmacology 1990;29:291-293.

86. Linn GS, Oî&#x2C6;&#x20AC;Keeffe RT, Lifshitz K, Schroeder C, Javitt DC. Behavioral effects of orally administered glycine in socially housed monkeys chronically treated with phencyclidine. Psychopharmacology (Berl) 2007; 192: 27-38.

87. Atkinson BN, Bell SC, De Vivo M, Kowalski LR, Lechner SM, Ognyanov VI, et al. ALX 5407: a potent, selective inhibitor of the hGlyT1 glycine transporter. Mol Pharmacol 2001;60:1414-1420.

76. Rujescu D, Bender A, Keck M, Hartmann AM, Ohl F, Raeder H, et al. A pharmacological model for psychosis based on N-methyl-D-aspartate receptor hypofunction: Molecular, cellular, functional and behavioral abnormalities. Biol Psychiatry 2006;59:721-729.

88. Javitt DC, Sershen H, Hashim A, Lajtha A. Reversal of phencyclidine-induced hyperactivity by glycine and the glycine uptake inhibitor glycyldodecylamide.

Neuropsychopharmacology 1997;17:202-204.

77. Behrens MM, Ali SS, Dao DN, Lucero J, Shekhtman G, Quick KL, et al. Ketamine-induced loss of phenotype of fast-spiking interneurons is mediated by NADPHoxidase. Science 2007;318:1645-1647.

89. Javitt DC, Frusciante M. Glycyldodecylamide, a phencyclidine behavioral antagonist, blocks cortical glycine uptake: Implications for schizophrenia and substance abuse.

Psychopharmacology (Berl) 1997;129:96-98.

78. Keilhoff G, Becker A, Grecksch G, Wolf G, Bernstein HG. Repeated application of ketamine to rats induces changes

90. Sucher NJ, Lipton SA. Redox modulatory site of the NMDA receptor-channel complex: Regulation by oxi-

14

Daniel C. Javitt

dized glutathione. J Neurosci Res 1991;30:582-591.

91. Lipton SA, Choi YB, Takahashi H, Zhang D, Li W, Godzik A, et al. Cysteine regulation of protein functionâ&#x20AC;&#x201C;as exemplified by NMDA-receptor modulation. Trends Neurosci 2002;25:474-480.

92. Do KQ, Trabesinger AH, Kirsten-Kruger M, Lauer CJ, Dydak U, Hell D, et al. Schizophrenia: Glutathione deficit in cerebrospinal fluid and prefrontal cortex in vivo. Eur J Neurosci 2000;12:3721-3728.

93. Matsuzawa D, Obata T, Shirayama Y, Nonaka H, Kanazawa Y, Yoshitome E, et al. Negative correlation between brain glutathione level and negative symptoms in schizophrenia: a 3T 1H-MRS study. PLoS One 2008;3:e1944.

94. Yao JK, Leonard S, Reddy R. Altered glutathione redox state in schizophrenia. Disease Markers 2006;22:83-93.

95. Steullet P, Neijt HC, Cuenod M, Do KQ. Synaptic plasticity impairment and hypofunction of NMDA receptors induced by glutathione deficit: Relevance to schizophrenia.

Neuroscience 2006;137:807-819.

96. Moghaddam B, Adams B, Verma A, Daly D. Activation of glutamatergic neurotransmission by ketamine: A novel step in the pathway from NMDA receptor blockade to dopaminergic and cognitive disruptions associated with the prefrontal cortex. J Neurosci 1997;17:2921-2927.

97. Moghaddam B. Targeting metabotropic glutamate receptors for treatment of the cognitive symptoms of schizophrenia. Psychopharmacology (Berl) 2004; 174; 39-44.

98. Moghaddam B, Adams BW. Reversal of phencyclidine effects by a group II metabotropic glutamate receptor agonist in rats. Science 1998;281:1349-1352.

99. Anand A, Charney DS, Oren DA, Berman RM, Hu XS, Cappiello A, et al. Attenuation of the neuropsychiatric effects of ketamine with lamotrigine: support for hyperglutamatergic effects of N-methyl-D- aspartate receptor antagonists. Arch Gen Psychiatry 2000;57:270-276.

100. Krystal JH, Abi-Saab W, Perry E, Dî&#x2C6;&#x20AC;Souza DC, Liu N, Gueorguieva R, et al. Preliminary evidence of attenuation of the disruptive effects of the NMDA glutamate receptor antagonist, ketamine, on working memory by pretreatment with the group II metabotropic glutamate receptor agonist, LY354740, in healthy human subjects.

Psychopharmacology (Berl) 2005;179:303-309.

101. Lecourtier L, Homayoun H, Tamagnan G, Moghaddam B. Positive allosteric modulation of metabotropic glutamate 5 (mGlu5) receptors reverses N-Methyl-D-aspartate antagonist-induced alteration of neuronal firing in prefrontal cortex. Biol Psychiatry 2007;62:739-746.

102. Rodriguez AL, Williams R. Recent progress in the development of allosteric modulators of mGluR5. Curr Opin Drug Discov Devel 2007;10:715-722.

103. Robbins MJ, Starr KR, Honey A, Soffin EM, Rourke C, Jones GA, et al. Evaluation of the mGlu8 receptor as a putative therapeutic target in schizophrenia. Brain Res 2007;1152:215-227.

104. Javitt DC. Glutamate as a therapeutic target in psychiatric disorders. Mol Psychiatry 2004;9:979, 984-997.

105. Coyle JT. The nagging question of the function of N-acetylaspartylglutamate. Neurobiol Dis 1997;4:231-238.

106. Olszewski RT, Wegorzewska MM, Monteiro AC, Krolikowski KA, Zhou J, Kozikowski AP, et al.

Phencyclidine and dizocilpine induced behaviors reduced by N-acetylaspartylglutamate peptidase inhibition via metabotropic glutamate receptors. Biol Psychiatry 2008;63:86-91.

107. Olszewski RT, Bukhari N, Zhou J, Kozikowski AP, Wroblewski JT, Shamimi-Noori S, et al. NAAG peptidase inhibition reduces locomotor activity and some stereotypes in the PCP model of schizophrenia via group II mGluR. J Neurochem 2004;89:876-885.

108. Lieberman JA, Papadakis K, Csernansky J, Litman R, Volavka J, Jia XD, et al. A randomized, placebo-controlled study of memantine as adjunctive treatment in patients with schizophrenia. Neuropsychopharmacology 2009; 34:1322-1329.

109. Javitt DC, Zylberman I, Zukin SR, Heresco-Levy U, Lindenmayer JP. Amelioration of negative symptoms in schizophrenia by glycine. Am J Psychiatry 1994;151: 1234-1236.

110. Heresco-Levy U, Javitt DC, Ermilov M, Mordel C, Silipo G, Lichtenstein M. Efficacy of high-dose glycine in the treatment of enduring negative symptoms of schizophrenia. Arch Gen Psychiatry 1999;56:29-36.

111. Javitt DC, Silipo G, Cienfuegos A, Shelley AM, Bark N, Park M, et al. Adjunctive high-dose glycine in the treatment of schizophrenia. Int J Neuropsychopharmacol 2001;4:385-392.

112. Heresco-Levy U, Ermilov M, Lichtenberg P, Bar G, Javitt DC. High-dose glycine added to olanzapine and risperidone for the treatment of schizophrenia. Biol Psychiatry 2004;55:165-171.

113. Tsai G, Yang P, Chung LC, Lange N, Coyle JT. D-serine added to antipsychotics for the treatment of schizophrenia. Biol Psychiatry 1998;44:1081-1089.

114. Heresco-Levy U, Javitt DC, Ebstein R, Vass A, Lichtenberg P, Bar G, et al. D-serine efficacy as add-on pharmacotherapy to risperidone and olanzapine for treatment-refractory schizophrenia. Biol Psychiatry 2005;57:577-585.

115. Tsai GE, Yang P, Chang YC, Chong MY. D-Alanine added to antipsychotics for the treatment of schizophrenia. Biol Psychiatry 2006; 59:230-234.

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GLUTAMATERGIC THEORIES OF SCHIZOPHRENIA

116. Tsai G, Lane HY, Yang P, Chong MY, Lange N. Glycine transporter I inhibitor, N-methylglycine (sarcosine), added to antipsychotics for the treatment of schizophrenia.

Biol Psychiatry 2004;55:452-456.

117. Lane HY, Chang YC, Liu YC, Chiu CC, Tsai GE. Sarcosine or D-serine add-on treatment for acute exacerbation of schizophrenia: a randomized, double-blind, placebo-controlled study. Arch Gen Psychiatry 2005;62:1196-1204.

118. Woods SW, Thomas L, Tully E, Hawkins KA, Miller TJ, Rosen JL, et al. Effects of oral glycine in the schizophrenia prodrome. Schiz Res 2004;70:79.

119. Berk M, Copolov D, Dean O, Lu K, Jeavons S, Schapkaitz I, et al. N-acetyl cysteine as a glutathione precursor for schizophrenia – a double-blind, randomized, placebocontrolled trial. Biol Psychiatry 2008;64:361-368.

120. Lavoie S, Murray MM, Deppen P, Knyazeva MG, Berk M, Boulat O, et al. Glutathione precursor, N-Acetyl-Cysteine, improves mismatch negativity in schizophrenia patients.

Neuropsychopharmacology2008; 33:2187-2199.

121. Tiihonen J, Hallikainen T, Ryynanen OP, Repo-Tiihonen E, Kotilainen I, Eronen M, et al. Lamotrigine in treatmentresistant schizophrenia: A randomized placebo-controlled crossover trial. Biol Psychiatry 2003;54:1241-1248.

122. Kremer I, Vass A, Gorelik I, Bar G, Blanaru M, Javitt DC, et al. Placebo-controlled trial of lamotrigine added to conventional and atypical antipsychotics in schizophrenia. Biol Psychiatry 2004;56:441-446.

123. Javitt DC, Balla A, Sershen H, Lajtha A. A.E. Bennett Research Award. Reversal of phencyclidine-induced effects by glycine and glycine transport inhibitors. Biol Psychiatry 1999;45:668-679.

124. Harsing LG, Jr., Gacsalyi I, Szabo G, Schmidt E, Sziray N, Sebban C, et al. The glycine transporter-1 inhibitors NFPS and Org 24461: A pharmacological study. Pharmacol Biochem Behav 2003;74:811-825.

16

125. Javitt DC, Hashim A, Sershen H. Modulation of striatal dopamine release by glycine transport inhibitors.

Neuropsychopharmacology 2005;30:649-656.

126. Bergeron R, Meyer TM, Coyle JT, Greene RW. Modulation of N-methyl-D-aspartate receptor function by glycine transport. Proc Natl Acad Sci USA 1998;95:15730-15734.

127. Chen L, Muhlhauser M, Yang CR. Glycine tranporter-1 blockade potentiates NMDA-mediated responses in rat prefrontal cortical neurons in vitro and in vivo. J Neurophysiol 2003;89:691-703.

128. Kinney GG, Sur C, Burno M, Mallorga PJ, Williams JB, Figueroa DJ, et al. The glycine transporter type 1 inhibitor N-[3-(4-fluorophenyl)-3-(4-phenylphenoxy)propyl] sarcosine potentiates NMDA receptor-mediated responses in vivo and produces an antipsychotic profile in rodent behavior. J Neurosci 2003;23:7586-7591.

129. Depoortere R, Dargazanli G, Estenne-Bouhtou G, Coste A, Lanneau C, Desvignes C, et al. Neurochemical, electrophysiological and pharmacological profiles of the selective inhibitor of the glycine transporter-1 SSR504734, a potential new type of antipsychotic. Neuropsychopharmacology 2005;30:1963-1985.

130. Le Pen G, Kew J, Alberati D, Borroni E, Heitz MP, Moreau JL. Prepulse inhibition deficits of the startle reflex in neonatal ventral hippocampal-lesioned rats: reversal by glycine and a glycine transporter inhibitor. Biol Psychiatry 2003;54:1162-1170.

131. Walter MW, Hoffman BJ, Gordon K, Johnson K, Love P, Jones M, et al. Discovery and SAR studies of novel GlyT1 inhibitors. Bioorg Med Chem Lett 2007;17:5233-5238.

132. Buchanan RW, Javitt DC, Marder SR, Schooler NR, Gold JM, McMahon RP, et al. The cognitive and negative symptoms in schizophrenia trial (CONSIST): The efficacy of glutamatergic agents for negative symptoms and cognitive impairments. Am J Psychiatry 2007;164:1593-1602.

Stephen I. Deutsch et al.

Regulation of Intermittent Oscillatory Activity of Pyramidal Cell Neurons by GABA Inhibitory Interneurons is Impaired in Schizophrenia: Rationale for Pharmacotherapeutic GABAergic Interventions Stephen I. Deutsch, MD, PhD,1 Richard B. Rosse, MD,2, 3 Barbara L. Schwartz, PhD,2, 3 John Mastropaolo, PhD,2 Jessica A. Burket, BS,1 and Abraham Weizman, MD4 1

Department of Psychiatry and Behavioral Sciences, Eastern Virginia Medical School, Norfolk, Virginia, U.S.A. Mental Health Service Line, Department of Veterans Affairs Medical Center, Washington, D.C., U.S.A. 3 Department of Psychiatry, Georgetown University School of Medicine, Washington, D.C., U.S.A. 4 Department of Psychiatry, Sackler Faculty of Medicine, Tel Aviv University, Ramat Aviv, Israel 2

ABSTRACT GABA, the major inhibitory neurotransmitter in the brain, is synthesized from L-glutamate and packaged within a family of highly differentiated inhibitory interneurons.

Individual GABA inhibitory interneurons in the frontal cortex can make terminal synaptic connections with more than 200 distinct pyramidal neurons, the principal output neuron. Moreover, the sites of these synaptic connections include shafts of dendritic spines, soma, dendritic branches, and initial axon segments.

The phasic activity of GABAergic neurons regulate intermittent oscillations of assemblies of pyramidal cell neurons, which are critical for many higher cortical functions such as working memory. Potentially, there are several viable pharmacotherapeutic strategies for facilitating GABAergic neurotransmission. A major research question is whether tonically-administered, selective GABAergic therapeutic interventions can mimic and correct disruptions of the intermittent oscillatory activity of assemblies of cortical pyramidal cell neurons.

Schizophrenia is a neurodevelopmental disorder of disturbed synaptic connectivity. Cognitive deficits correlate with poor functional outcomes, do not vary with state of illness or medication status, and can often be detected in milder form in unaffected closely-related biological relatives of patients with schizophrenia. Disturbances of GABAergic inhibitory influences in the dorsolateral prefrontal cortex contribute to cognitive symptoms and serve as therapeutic targets in schizophrenia (1-6).

Importantly, a delicate balance must exist between GABAergic inhibitory influences and excitatory influences that are primarily mediated by L-glutamate (7).

GABAergic Dysfunction in Schizophrenia GABA is the major inhibitory neurotransmitter in the brain, mediating fast synaptic neurotransmission on a less than 100-millisecond timeframe via its ability to gate a receptor-associated chloride ion channel, referred to as the GABAA receptor (8-11). GABA is a critical neurotransmitter involved in the control of cortical oscillatory rhythmic activity, during which fundamental cognitive processes occur; these GABA-regulated oscillatory network activities and their associated cognitive processes are often disturbed in schizophrenia (e.g., beta2 and gamma oscillations) (1-5, 9, 12, 13).

Address for Correspondence: Stephen I. Deutsch, MD, PhD, Ann Robinson Endowed Chair in Psychiatry, Professor and Chairman, Department of Psychiatry and Behavioral Sciences, Eastern Virginia Medical School, 825 Fairfax Avenue, Suite 710, Norfolk, Virginia 23507-1912, U.S.A. â&#x20AC;&#x2020;deutscsi@evms.edu

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Abnormalities of GABAergic inhibitory tone in the cortex of patients with schizophrenia were suggested by postmortem studies reporting decreased activity of glutamic acid decarboxylase (GAD), decreased GABA reuptake and a “compensatory” increase of GABA receptor binding (8, 9, 14-18). These earlier studies were corroborated by later gene expression studies showing diminished cortical expression of mRNA transcripts expressed in GABAergic neurons: GAD67, the biosynthetic enzyme for GABA; reelin, a signaling-protein that is critical for the correct lamination of the developing brain and changes in dendritic spine density that serve as morphological correlates of neuronal plasticity; parvalbumin, a calcium ion binding protein; and the GABA membrane transporter (GAT1), which mediates presynaptic reuptake of GABA and termination of its synaptic actions (8, 9, 14-19).

Diminished expression of parvalbumin in GABAergic neurons (e.g., chandelier neurons) may be a compensatory response that facilitates presynaptic GABA release because, ordinarily, parvalbumin is responsible for decreasing the transient elevations of intraneuronal calcium ion concentrations that mediate depolarizationdependent GABA release from GABA nerve terminals (9). Similarly, reduced expression of GAT1 on the nerve terminals of chandelier cells, which would increase synaptic levels of GABA, and increased density of α2-containing GABAA receptors on the axon initial segment of pyramidal neurons may reflect a compensatory, but inadequate, “facilitation” of GABA signaling (9, 15, 16-18).

Isolation-induced rearing of rats after they are weaned leads to a quantifiable deficit in sensorimotor gating (i.e., impaired prepulse inhibition (PPI) of the acoustic startle response) that is also manifest by patients with schizophrenia and their closely-related biological relatives (20). Of possible interest to the GABAergic dysfunction of schizophrenia, which may be widespread throughout the cerebral cortex and hippocampus, is that the immunoreactive protein content of parvalbumin and calbindin, two calcium ion binding proteins expressed in GABA inhibitory interneurons, was significantly reduced in several hippocampal subfields (i.e., dentate gyrus, CA2/3, and CA1 [only calbindin]) of the isolation-reared rats (20). These data are important for several reasons: they suggest that an environmentally-relevant social stressor can selectively alter hippocampal expression of proteins in animals displaying deficits relevant to the pathophysiology of schizophrenia and can cause dysfunction of GABAergic 18

neurons. Presumably, this effect of isolation-rearing is mediated by epigenetic mechanisms that affect expression of parvalbumin and calbindin (20).

Pharmacological Anatomy of the GABAA Receptor Complex The GABAA receptor complex is a member of a superfamily of ligand-gated ion channel receptors that includes nicotinic acetylcholine receptors; the receptor is a pentameric protein complex constructed from five constituent polypeptide subunits (9-11). The individual subunits are integral membrane proteins that share significant sequence homology with each other, as well as a common motif: a large extracellular N-terminal domain, three membrane-spanning hydrophobic domains (M1-M3), an intracellular cytoplasmic loop between M3 and M4, a final membrane-spanning domain (M4), and a C-terminal extracellular end. The M2 domain from each of the five constituent polypeptides align themselves in such a manner that they form a potential pore or channel, whose kinetics of opening is determined by the binding of GABA to the receptor.

The intracellular loop has multiple phosphorylation sites; the state of phosphorylation may be regulated by the second messenger cascades of other neurotransmitter receptors, reflecting “cross-talk” between GABA and these other neurotransmitters. Based on the extent of their sequence homology, at least 16 individual polypeptide subunits, which are encoded by distinct genes, are grouped into families identified by Greek letters: α1-6, β1-3, γ1-3, δ, ε, θ, π, and ρ1-3. Most commonly, functional receptors contain two α subunits, two β subunits, and a γ subunit, the γ subunit is rarely replaced by a subunit from another family (9).

If the constraint of subunit combinations is restricted to receptors containing two α and two β subunits, combinatorial diversity could result in more than 2,000 pharmacologically-distinctive GABAA receptors; however, fewer than 20 of the 2,000 possible combinations are commonly detected in the brain. The α1-containing receptors are most abundant, they are located synaptically on cell bodies and dendrites of principal neurons and extrasynaptically; they are frequently combined with β2 and γ2 subunits, and have a high-affinity for classical benzodiazepines, mediating their sedative, amnestic and anticonvulsant properties (10, 11, 21). α2-Containing receptors are expressed less commonly than α1-containing receptors, are usually combined with

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β3 and γ2 subunits, and have a high-affinity for classical benzodiazepines, mediating primarily their anxiolytic actions (9-11). The localization of α2-containing GABAA receptors on the initial segment of principal neuron axons in the cerebral cortex and hippocampus is consistent with their functional role in controlling the output of these neurons. Mice with a genetically-engineered knockout of the gene for the α3 subunit show a profound deficit of prepulse inhibition of the acoustic startle reflex, reflecting a deficit in sensorimotor gating that models what is frequently seen in patients with schizophrenia (9, 21). α5-Containing GABAA receptors are located extrasynaptically in hippocampus and the cerebral cortex; their location on the dendritic spines of hippocampal pyramidal cells, a site of excitatory N-methyl-D-aspartate (NMDA) receptors, implicates their complementary involvement with NMDA receptors in regulating activity of this principal hippocampal efferent neuron (1, 9, 21). The α5-containing receptor is involved in a variety of hippocampal functions, including spatial and temporal associative memory, spatial learning, fear conditioning and prepulse inhibition. Although not sufficient by itself, the presence of the γ2 subunit contributes to benzodiazepine sensitivity; receptors containing either γ1 or γ3 subunits have reduced affinity for classical benzodiazepines (9, 21).

Clearly, the inhibitory tone of the brain, including phasic inhibition that supports critical oscillatory rhythms, is highly regulated. However, the diversity of pharmacologically-distinctive GABAA receptors presents, at least theoretically, the opportunity for the development of highly-selective therapeutic interventions, such as α3 and α5-selective GABAA receptor agonists (9, 21, 22).

GABA Inhibitory Interneurons Regulate Principal Pyramidal Cell Neuron Output Pyramidal neurons are primarily glutamatergic and serve as the principal output neurons of the dorsolateral prefrontal cortex, integrating excitatory and inhibitory inputs and sending axonal projections to other cortical regions (layers 2 and 3), striatum and subcortical structures (layer 5), and thalamus (layer 6), among other possible projections (1-6). A possible morphological correlate of disturbed synaptic connectivity in schizophrenia is a lowered density of dendritic spines on pyramidal neurons in deep layer 3; also, there may be a reduction of somal area of these pyramidal cell neurons (4, 5, 9).

The mediodorsal thalamus provides excitatory input to

the pyramidal neurons in layers 3 and 4 of the dorsolateral prefrontal cortex (5). Postmortem data show that the cortical neuropil, which is the “space” between neurons comprised of axon terminals, dendritic spines and glial processes, is reduced in the dorsolateral prefrontal cortex of patients with schizophrenia, which is consistent with disruption of synaptic connectivity.

GABAergic inhibitory neurons can be distinguished from each other by shape, laminar locations, projections to principal neurons (e.g., dendritic spines, soma, or initial axon segment) and other GABAergic cells, membrane firing properties, and expression of specific proteins and neuropeptides (e.g., parvalbumin and calcitonin, which are calcium ion binding proteins, and somatostatin and cholecystokinin, which are neuropeptides) (5, 9).

Moreover, GABAergic interneurons show a domainspecific innervation of the principal cells to which they project (e.g., hippocampal pyramidal cells) affecting both the input that these principal cells receive as well as their output, which is regulated by axo-axonic GABAergic interneurons. Importantly, the response of the principal neuron is not only affected by the spatial distribution of GABAergic innervation and the morphological type of interneuron, but also by the type of synaptic and extrasynaptic GABAA receptor (5, 9). For example, there are two types of GABAergic “basket” cells that innervate the cell body of the hippocampal cell: so-called “fast spiking” parvalbumin-containing basket cells that synapse with α1-containing GABAA receptors and “regular spiking” cholecystokinin (CCK)-positive basket cells that synapse with α2-containing GABAA receptors (5, 9). Within the dorsolateral prefrontal cortex, parvalbumin-containing GABAergic neurons are of two types: chandelier neurons that form synapses on the axon initial segments and basket neurons that form synapses on the cell bodies and proximal dendrites of pyramidal neurons (5, 9).

Working memory is very sensitive to disruption of the normal activity of GABAergic inhibitory neurons in the dorsolateral prefrontal cortex (1, 5, 9, 12). Chandelier and basket GABAergic inhibitory neurons subserve the timing mechanism that synchronizes the activity of local populations of pyramidal neurons that they target, which underlie the gamma oscillations in the 30 to 80 Hz band.

The power of the gamma oscillations in the dorsolateral prefrontal cortex are associated with working memory load; both working memory load and the power of gamma oscillatory activity in the dorsolateral prefrontal cortex and elsewhere are diminished in schizophrenia (5, 12).

The synchronous firing of assemblies of cortical pyra19

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midal neurons in prefrontal cortex that is controlled by fast-spiking GABAergic interneurons is intermittent, which is consistent with a role in higher cognitive processes that are activated selectively and according to specific task demands. Cortical GABAergic inhibitory interneurons may establish synapses with more than 200 different pyramidal neurons, which serves as the anatomic substrate for orchestrating the intermittent synchronous firing of these assemblies of pyramidal neurons. As discussed, the output of the principal cortical pyramidal cell neurons (including locally-evoked graded membrane potentials, triggering of action potential that invades the axon, and depolarization-dependent and calcium ion-dependent release of glutamate) is multidetermined and depends on their laminar locations, efferent input of morphologically and biochemically-distinct GABAergic interneurons, and sites of synaptic connections (e.g., proximal dendrite or distal brach, dendritic spine or neck of dendritic spine, cell body, or initial axon segment). Thus, pyramidal cell bodies and proximal dendrites receive innervation from basket cells in cortical layers II, III, V, and VI, axon initial segments receive innervation from chandelier interneurons in middle cortical layers, and double-bouquet and horizontal cells innervate shafts of dendritic branches in layers II/III and layer I, respectively (21).

Biochemical Heterogeneity of GABA Inhibitory Interneurons GABAergic inhibitory interneurons can be biochemically distinguished from each other by whether or not they constitutively express and release reelin, a “signaling” glycoprotein that shares structural analogies with extracellular matrix proteins (19, 23-25). In the developing fetal brain, reelin-signal transduction is critical for the correct positioning of post-mitotic neurons migrating from the ventricular zone, formation of appropriate synaptic connections and cortical lamination, whereas in the fully-developed and mature brain, reelin-signal transduction participates in the mediation of experience-dependent synaptic plasticity (23). A morphological correlate of reelin’s role in experience-dependent synaptic plasticity is the creation and reshaping of dendritic spines, which depend on the stable assembly of correctly-aligned microtubules. Two members of the family of low-density lipoprotein receptors that are expressed in brain (i.e., very low-density lipoprotein [VLDL] receptor and apolipoprotein E 20

[apoE] type 2 receptor) participate in the transduction of the reelin signal, a process that includes several significant “downstream” consequences (23). At least one of these important “downstream” consequences that may be related to changes in the size, shape and density of dendritic spines, a morphological correlate of experience-dependent learning, is the stabilization of microtubules by inhibiting hyper-phosphorylation of the tau protein, an important microtubule-associated protein.

Therefore, it is of interest that GABAergic interneurons regulate both the synchronous intermittent firing of assemblies of cortical pyramidal neurons, which is part of the electrophysiological substrate for gamma oscillations that are necessary for cognitive processes such as working memory, and changes in size, shape and density of dendritic spines, a morphological correlate of experience-dependent learning (5, 12, 23).

There are emerging data suggesting that in at least some patients with schizophrenia, there is diminished expression of GAD 67, the rate-limiting enzyme for GABA biosynthesis with a high-affinity for binding its pyridoxal phosphate cofactor, and reelin in a selective population of GABAergic neurons (9, 14, 19). Because of its high-affinity for pyridoxal phosphate and saturation of the binding site on the enzyme with ordinary tissue levels of this cofactor, the activity of GAD67 is regulated by gene expression; increased activity is related to increased translation of new GAD67 mRNA. Thus, if there is diminished expression of GAD67, there is shrinkage of the neurotransmitter pool of GABA. Diminished expression of both GAD67, which leads to a presynaptic deficiency of GABA in selected neurons, and reelin, which leads to disruption of the morphological expression of synaptic plasticity (i.e., changes in size, shape and density of dendritic spines), can occur as a result of altered epigenetic regulation of the genetic expression of these two proteins in patients with schizophrenia (19).

GABAergic Dysfunction Underlie Deficits of Working Memory in Schizophrenia: Effects of Non-Selective GABAergic Interventions The effects of lorazepam, a GABAA/benzodiazepine receptor agonist, and flumazenil, a GABAA/benzodiazepine antagonist or partial inverse agonist on working memory performance were studied while patients and control subjects underwent functional magnetic resonance imaging (fMRI) (13). The goal of the study was

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to show differential effects of the two benzodiazepine receptor ligands on task performance and fMRI, as well as differences in the modulatory effects of the two ligands between patients and control subjects. Eleven right-handed, male patients with chronic schizophrenia (mean duration of illness +SD = 17.0 + 8.3) stably medicated with antipsychotic medications (10 of whom were receiving clozapine) and showing impairment of episodic memory and verbal fluency and moderate symptom severity were matched for gender, age, handedness and premorbid IQ with 11 control subjects (13). In this placebo-controlled, double-blind study, participants underwent four imaging sessions each of which was separated by two weeks: baseline and then, in counterbalanced, randomized fashion, assigned to lorazepam (2 mg oral capsule), flumazenil (IV bolus of 0.9 mg, followed by constant IV infusion at a rate of 0.0102 mg/min), or matching placebo condition. While the images were obtained, the participants performed an “N-back test of verbal working memory” at three levels of task difficulty. The task required the participants to indicate whether a current visually-presented letter was the same as the one immediately preceding it (N=1) or the same as the one presented two (N=2) or 3 (N=3) letters earlier. Across all participants, task performance worsened with increasing difficulty and cognitive demands of the task, as measured by a discrimination index derived from signal detection theory that corrects for false positive responses. There was also a significant interaction between drugs and task difficulty with the effects of lorazepam and flumazenil most marked at the N=1 and N=2 levels of difficulty. Among the patients with schizophrenia, lorazepam, the GABAA/benzodiazepine receptor agonist, significantly worsened working memory performance, whereas flumazenil, the GABAA/ benzodiazepine receptor antagonist or partial inverse agonist actually improved performance (13). These data are somewhat contrary to what was expected in what may be our limited “linear” conceptualizations of effects of diminished GABAergic tone on, or phasic regulation of, integrated assemblies of principal pyramidal neurons by the parvalbumin-expressing, chandelier GABAergic cortical interneurons. Rather, the exacerbation of the already poorer performance of the patients with schizophrenia relative to controls by lorazepam, a positive allosteric modulator of GABAA receptors, and improvement of performance by flumazenil, which might be expected to dampen the endogenous GABAergic tone or be devoid of any effect suggests that mechanisms of

working memory impairment and disturbed pyramidal oscillatory activity in schizophrenia may be more complex, involving “overinhibition.” The findings were consistent with effects of GABAA receptor-modulating drugs on working memory performance, as opposed to nonspecific effects on response latency (13, 21).

In the control subjects performing the working memory task under the placebo condition, the fMRI data showed that there were areas of brain activation, including the bilateral prefrontal, premotor, parietal and anterior cingulate cortices, and areas of brain deactivation, including the bilateral temporal and posterior cingulate regions, in association with increasing levels of task difficulty (13). The magnitude of the coordinated changes of activation and deactivation was reduced in the patients with schizophrenia, compared to the control subjects. The data suggested that flumazenil had significantly different effects on the fMRI data in the patient group, enhancing deactivation generally and possibly increasing activation in the anterior cingulate cortex. The fMRI data highlight an important role for GABA in coordinating and assuring efficiency of diffuse brain regions while performing complex cognitive tasks. This efficiency appears to be impaired in schizophrenia and may be improved by selective interventions with GABAA receptor-modulating drugs.

Epigenetic Regulation of GABAergic Function: Potential Opportunities for Therapeutic Intervention An epigenetic mechanism that could suppress expression of both GAD67 and reelin is the “hypermethylation” of the 5’ position of the cytosine ring in so-called CpG islands in the promoter regions of the genes for these two proteins.

An enzyme responsible for methylating cytosine rings in promoter regions is DNA-methyltransferase 1 (DNMT1), whose preferential expression may be increased in the same cortical GABAergic interneurons (i.e., there is increased mRNA and protein content for DNMT1 in these neurons) showing decreased expression of GAD67 and reelin (19, 24, 25). If true, it is conceivable that epigenetic therapeutic strategies can be developed to “inhibit” DNMT1 and, thereby, promote expression of GAD67 and reelin. Additionally, strategies to promote chromosomal remodeling that involve inhibiting the deacetylation of histone proteins can lead to promotion of gene expression (26). Thus, in terms of a potential mechanism of pathogenesis of schizophrenia, it is very provocative that both the mRNA and protein content of DNMT1 are increased 21

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in cortical GABAergic interneurons of layers I, II, and IV in postmortem brains obtained from carefully-diagnosed patients with the disorder; interneurons in these same cortical layers show deficient expression of GAD67 and reelin (25).

The hypermethylation hypothesis of specific cytosine residues in the promoter regions of the genes for reelin and GAD67 is an appealing epigenetic mechanism that could account for GABAergic dysfunction.

Unfortunately, when the DNA methylation status of the promoter region for the reelin gene (RELN) was studied in prefrontal cortices of 14 patients with schizophrenia and 13 controls using a different methodology (referred to as the pyrosequencing method), especially focusing on cytosines at positions -139 and -134 that were reported to be hypermethylated in an earlier study (24), the level of DNA methylation in both groups was below the level of detection (27). Thus, this independent study did not confirm that the RELN promoter is hypermethylated in the prefrontal cortex of patients with schizophrenia. There are several possible reasons for the discrepant findings, including differences in brain regions between the studies and the likely possibility that hypermethylation, if it occurs, would be confined to specific subpopulations of GABAergic interneurons (27). Clearly, in view of the reported increase of DNMT1 expression, the possible role of this proposed epigenetic mechanism must be studied further.

Neurotransmitter Receptor-Mediated Regulation of GABA Inhibitory Interneurons: Targets for Therapeutic Intervention Alternative mechanisms for creating and addressing functional deficits of prefrontocortical “fast-spiking” GABAergic interneurons include receptor-mediated regulation of their firing and GABA release by both glutamate and acetylcholine (28-34). Glutamate would act primarily at the N-methyl-D-aspartate (NMDA) receptor, a type of glutamate-gated calcium ion channel receptor on the surface of these “fast-spiking” interneurons, while acetylcholine would act at cell-surface nicotinic acetylcholine receptors containing the α 7 subunit on these same interneurons. The relationship of the firing of the “fast-spiking,” probably parvalbumin-containing, GABAergic interneuron to the firing of the “regularspiking” principal pyramidal output neuron was studied in the medial prefrontal cortex of freely-moving rats 22

(3). Essentially, inhibition of the firing rate of the “fastspiking” interneurons was achieved with a systemic dose of MK-801 (dizocilpine; 0.1 mg/kg, ip), a use-dependent, open-channel, noncompetitive NMDA receptor antagonist that disrupts working memory and set-shifting in these animals, “symptoms” relevant to the cognitive deficits of schizophrenia. While MK-801 inhibited a majority of the “fast-spiking” interneurons, treatment with MK-801 increased the firing rate of the majority of the “regular-spiking” pyramidal neurons. The data show that NMDA receptor-mediated inhibition of GABAergic inhibitory interneurons leads to “disinhibition” of the firing of cortical pyramidal cell neurons. Importantly, there was a lag between the “start/plateau” of MK-801-induced inhibitions of the “fast-spiking” interneurons and its excitations of “regular-spiking” pyramidal neurons, consistent with both models and electrophysiological data of synchronized firing of cortical networks subserving higher-executive functions. In any event, tonic inhibition of NMDA receptors by MK-801 disrupted GABAergic inhibitory control of a synchronized network of “regularspiking” pyramidal neurons (3). a. NMDA Receptor

There are several nonmutually exclusive and nonoverlapping ways of creating “NMDA receptor hypofunction” on the surface of GABAergic inhibitory interneurons, whose end-result would be disinhibition and disruption of the synchronous firing of assemblies of cortical pyramidal cell neurons. These mechanisms can include alterations of genetic expression of specific NMDA receptor polypeptide subunits, post-transcriptional changes in mRNA splice variants of specific receptor subunits, and posttranslational modifications of the phosphorylation state of the intracellular loop of receptor subunits (2, 29, 32). The ability of glutamate to gate the opening of its associated ion channel successfully is also influenced by the extent of saturation of the strychnine-insensitive glycine co-agonist binding site, as well as by the availability of both locally produced and circulating allosteric modulators such as polyamines and neurosteroids (2). The receptor’s function is influenced by the local balance of oxidative and reducing equivalents. Very recently, interest has focused on the regulation of the presynaptic release of glutamate by specific G-protein-coupled metabotropic glutamate receptors in the area of the NMDA receptor synapse that, in turn, affects activation of the NMDA receptor; specifically, strategies for inhibiting enzymatic cleavage of N-acetylaspartate-glutamate, an acidic dipeptide

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that may be the naturally-occurring agonist for specific metabotropic glutamate receptors have been proposed as a therapeutic intervention (35). Thus, NMDA receptor hypofunction can come about by a variety of mechanisms, which also suggest a variety of possible therapeutic interventions. These NMDA receptor-targeted interventions include administration of glycine complete or partial agonists such as D-serine and glycine reuptake inhibitors related to sarcosine; as mentioned, glycine is an obligatory co-agonist whose presence enhances and is necessary for glutamate to be effective in promoting channel opening. Ideally, this would address the GABAergic deficit that results from hypofunction of NMDA receptors on their cell-surface. A provocative therapeutic strategy also includes addressing the downstream consequences of the excessive excitation of pyramidal cell neurons and the possible pathological consequences of the excessive release of glutamate resulting from GABAergic dysfunction (2, 36). This strategy will be facilitated by the development and introduction of AMPA/kainate receptor antagonists into Clinical Medicine (37-39). There has been some preliminary investigation of the therapeutic efficacy of topiramate in schizophrenia, whose properties include kainate receptor antagonism.

fested by many patients with schizophrenia and their closely-related biological relatives (28, 41). There are provocative data showing that schizophrenia, P50 sensory gating abnormalities and the increased prevalence of smoking behavior in this disorder may be linked or associated with abnormal promoter variants for the α7 subunit in the q13-q14 region of chromosome 15 (41). In any event, like NMDA receptor hypofunction, deficient expression of the α7 subunit may result in GABAergic dysfunction and “downstream” disinhibition. The existence of a positive allosteric modulatory site on nicotinic acetylcholine receptors and the fact that galantamine is a positive allosteric modulator, in addition to an inhibitor of acetylcholinesterase, have stimulated thinking about the development of therapeutic interventions (30, 42). In addition to choline, there are anabaseine derivatives that serve as selective agonists for receptors containing the α7 subunit; unfortunately, the receptor undergoes rapid desensitization on exposure to agonist. Nonetheless, targeted therapeutic interventions for the α7-containing nicotinic acetylcholine receptor are under consideration for the treatment of schizophrenia (30).

b. α7-Containing Nicotinic Acetylcholine Receptors

The firing of subpopulations of GABAergic interneurons is regulated, at least in part, in a local “retrograde” manner by endogenous cannabinoids (endocannabinoids), such as anandamide, which are released from principal pyramidal cell neurons and bind to CB1-cannabinoid receptors present on pre-

In addition to the NMDA receptor, nicotinic acetylcholine receptors, which are also ligand-gated ion channel receptors that contain the α7 subunit are expressed on the surface of GABAergic inhibitory interneurons (28, 30). The α7 subunit confers unique electrophysiological and pharmacological properties, including relatively high permeability to calcium ions and rapid desensitization. Also, choline, which is the locally-generated hydrolytic split product of acetylcholine by the catalytically very efficient acetylcholinesterase, is a full agonist that mimics acetylcholine itself (31, 40). Although hydrolytic cleavage of acetylcholine is the major mechanism of its synaptic inactivation, the rapid local generation of choline, a full agonist, and the rapid desensitization kinetics of the receptor suggest that this may be an additional mechanism for locally and rapidly regulating synaptic neurotransmission (31, 40). There are compelling data supporting the regionally-selective diminished expression of the α7-containing nicotinic acetylcholine receptor in the brains of patients with schizophrenia, which may underlie the deficits in sensory impairment (i.e., the P50 auditory-evoked potential abnormality) and voluntary smooth pursuit eye-tracking mani-

c. CB1-Cannabinoid Receptor

Table 1. Potential Therapeutic Strategies for Improving GABAergic Function PRESYNAPTIC a. Metabotropic Glutamate Receptor Agonist Intervention NAAG Peptidase Inhibition GABA INHIBITORY INTERNEURON a. NMDA Receptor Agonist Intervention Glycine Reuptake Inhibitor (e.g., Sarcosine) Glycine Agonist (e.g., D-Serine) Neurosteroids Polyamines b. α7 Nicotinic Acetylcholine Receptor Agonist Intervention Positive Allosteric Modulator (e.g., Galantamine) Selective Agonist (e.g., Choline, Anabaseine Derivative) c. CB1-Cannabinoid Receptor Antagonist d. Epigenetic Intervention Histone Deacetylase Inhibitor DNA Methyltransferase 1 Inhibitor POSTSYNAPTIC a. GABAA Receptor Agonist Intervention Selective Benzodiazepine Ligands Neurosteroids b. Inhibition of Phosphorylation of Tau Protein (Preservation of Assembly, Stability and Alignment of Microtubules)

23

GABAergic Dysfunction in Schizophrenia

synaptic GABA inhibitory interneurons (9, 43). The synthesis and release of endocannabinoids is dependent on the activity of the pyramidal cell neurons; the consequence of the retrograde signal is the transient inhibition of GABA release from terminals containing the CB1-receptors, referred to as depolarizationinduced suppression of inhibition. For example, CB1cannabinoid receptors are present on the terminals of CCK-containing GABAergic basket cells in the hippocampus and amygdala (9).

Figure 1. This fictionalized cartoon depicts sites of potential synaptic innervation of principal pyramidal output neurons by GABA inhibitory interneurons. The output of fast-spiking GABA inhibitory interneurons is controlled, in part, by both NMDA and α7-containing nicotinic acetylcholine receptors on their surface.

The firing of the pyramidal neuron will be determined by the site of the synaptic connection. In addition to controlling the firing of pyramidal cell neurons, GABAergic neurons may also regulate their morphological appearance, especially the density

GABA A.

CONCLUSIONS It is very clear that the phasic inhibitory regulation of the intermittent oscillatory activity of assemblies of pyramidal cell neurons is necessary for the performance of higher cortical functions and may be disturbed in schizophrenia. GABA-gated chloride ion conductance via GABAA receptors is a basic element of this regulatory process; disturbances of GABAergic function can come about by a variety of mechanisms, many of which can of their dendritic spines. The figure also depicts some of the possible therapeutic targets for facilitating GABAergic function and neurotransmission, including the NMDA and α7-containing nicotinic acetylcholine receptors and epigenetic interventions.

Ultimately, pyramidal cell function may depend on the correct alignment of microtubules, and the intermittent oscillatory output of assemblies of these neurons may be critical for higher executive functions, including working memory.

Sites of Synaptic Connection A. Shaft of Dendritic Spine B. Dendritic Branch C. Cell Soma D. Axon Initial Segment

ery Low Density Lipoprotein Receptor Apolipoprotein E Type 2 Receptor - �7nicotinic Acetylcholine Receptor N-Methyl-D-Aspartic Acid Receptor -Acetylated Histone Protein H3- Methylated Cytosine Ring

Legend VLDLR- Very Low Density Lipoprotein Receptor apoER2- Apolipoprotein E Type 2 Receptor �7nAChR- �7nicotinic Acetylcholine Receptor NMDAR- N-Methyl-D-Aspartic Acid Receptor -Acetylated Histone Protein CH3- Methylated Cytosine Ring

REELIN

{ VLDLR apoER2 NMDAR GABA

NMDAR

GABA

B.

GA REE

C.

�7nAChR GAD REELIN �7nAChR

CH3

D. GABA

GABA Neuron

GABA Neuron Aligned microtubules

Dephosphorylated Tau Protein

Output Pyramidal Cell Neuron

24

CH3

ntermittent Synchcronous Oscilations { IWorking Memory

Stephen I. Deutsch et al.

also serve as therapeutic targets. Moreover, because the function of individual GABAA receptors is determined, in large part, by the unique combinations of individual receptor subunits, it is possible to develop subtype selective interventions that target GABAA receptors containing specific subunits (e.g., α3 and α5). Unfortunately, pharmacotherapeutic interventions are usually administered systemically and would facilitate GABAergic neurotransmission widely throughout the brain, whereas the precise pathophysiological disruption may be confined to specific GABA inhibitory interneurons within specific regions and cortical layers. Also, usual pharmacotherapeutic interventions lead to sustained “steady-state” levels of the active medication in blood and other biological compartments, which may not mimic phasic activity and release of GABA. Nonetheless, in view of the current limitations of the pharmacotherapy of schizophrenia that focuses on selective blockade of specific dopamine receptor subtypes with efficacy and effectiveness confined primarily to positive symptoms, the development of alternative and complementary strategies that target cognitive symptoms must be pursued.

Acknowledgement We thank the Mental Illness Research, Education and Clinical Center (MIRECC), VISN5, Department of Veterans Affairs and its Director, Dr. Alan Bellack, for continued support. The authors would also like to thank Mr. Andrew J. White of the Department of Veterans Affairs Medical Center in Washington, D.C., for his outstanding illustration.

References 1. Daskalakis ZJ, Fitzgerald PB, Christensen BK. The role of cortical inhibition in the pathophysiology and treatment of schizophrenia. Brain Res Rev 2007; 56: 427-442.

2. Deutsch SI, Rosse RB, Schwartz BL, Mastropaolo J. A revised excitotoxic hypothesis of schizophrenia: Therapeutic implications. Clin Neuropharmacol 2001; 24: 43-49.

3. Homayoun H, Moghaddam B. NMDA receptor hypofunction produces opposite effects on prefrontal cortex interneurons and pyramidal neurons. J Neurosci 2007; 27: 11496-11500.

4. Lewis DA, Hashimoto T, Volk DW. Cortical inhibitory neurons and schizophrenia. Nat Rev Neurosci 2005; 6: 312-324.

5. Lewis DA, Gonzalez-Burgos G. Neuroplasticity of neocortical circuits in schizophrenia. Neuropsychopharmacol Rev 2008; 33: 141-165.

6. Moghaddam B, Adams B, Verma A, Daly, D. Activation of glutamatergic neurotransmission by ketamine: A novel step in the pathway from NMDA receptor blockade to dopaminergic and cognitive disruptions associated with the

prefrontal cortex. J Neurosci 1997; 17: 2921-2927.

7. Deutsch SI, Park CH, Lukacs LG, Morn C, Koetzner L, Mastropaolo J. MK-801 alters the GABAA receptor complex and potentiates flurazepam’s antiseizure efficacy.

Pharmacology Biochemistry and Behavior 1995; 51: 909-915.

8. Huntsman MM, Tran BV, Potkin SG, Bunney Jr., WE, Jones EG. Altered ratios of alternatively spliced long and short gamma2 subunit mRNAs of the gamma-amino butyrate type A receptor in prefrontal cortex of schizophrenics. Proc Natl Acad Sci USA 1998; 95: 15066-15071.

9. Mohler H. GABAA receptor diversity and pharmacology.

Cell Tissue Res 2006; 326: 505-516.

10. Barnard EA, Skolnick P, Olsen RW, Mohler H, Sieghart W, Biggio G, Braestrup C, Bateson AN, Langer SZ. Subtypes of γ-aminobutyric acidA receptors: Classification on the bases of subunit structure and receptor function. Pharmacol Rev 1998; 50: 291-313.

11. Benson J, Low K, Keist R, Mohler H, Rudolph U. Pharmacology of recombinant GABAA receptors rendered diazepam-insensitive by point-mutated α-subunits. FEBS Lett 1998; 431: 400-404.

12. Howard MW, Rizzuto DS, Caplan JB, Madsen JR, Lisman J, Aschenbrenner-Scheibe R, Schulze-Bonhage, A, Kahana, MJ. Gamma oscillations correlate with working memory load in humans. Cereb Cortex 2003; 13: 1369-1374.

13. Menzies L, Ooi C, Kamath S, Suckling J, McKenna P, Fletcher P, Bullmore E, Stephenson C. Effects of γ-aminobutyric acid-modulating drugs on working memory and brain function in patients with schizophrenia.

Arch Gen Psychiatry 2007; 64: 156-167.

14. Akbarian S, Kim JJ, Potkin SG, Hagman JO, Tafazzoli A, Bunney Jr WE, Jones EG. Gene expression for glutamic acid decarboxylase is reduced without loss of neurons in prefrontal cortex of schizophrenics. Arch Gen Psychiatry 1995; 52: 258-266.

15. Benes FM, Vincent SL, Alsterberg G, Bird ED, SanGiovanni JP. Increased GABAA receptor binding in superficial layers of cingulate cortex in schizophrenics. J Neurosci 1992; 12: 924-929.

16. Hashimoto T, Arion D, Unger T, Maldonado-Aviles JG, Morris HM, Volk DW, Mirnics K, Lewis DA. Alterations in GABA-related transcriptome in the dorsolateral prefrontal cortex of subjects with schizophrenia. Mol Psychiatry 2008; 13: 147-161.

17. Simpson MDC, Slater P, Deakin JFW, Royston MC, Skan WJ. Reduced GABA uptake sites in the temporal lobe in schizophrenia. Neurosci Lett 1989; 107: 211-215.

18. Simpson MD, Slater P, Royston MC, Deakin JF. Regionally selective deficits in uptake sites for glutamate and gammaaminobutyric acid in the basal ganglia in schizophrenia.

Psychiatry Res 1992; 42: 273-282.

19. Guidotti A, Auta J, Davis JM, Dwivedi Y, Gerevini VD, 25

GABAergic Dysfunction in Schizophrenia

Impagnatiello F, Pandey G, Pesold C, Sharma R, Uzunov DP, Costa E. Decrease in reelin and glutamic acid decarboxylase67 (GAD67) expression in schizophrenia and bipolar disorder: A postmortem brain study. Arch Gen Psychiatry 2000; 57: 1061-1069.

20. Harte MK, Powell SB, Swerdlow NR, Geyer MA, Reynolds GP. Deficits in parvalbumin and calbindin immunoreactive cells in the hippocampus of isolation reared rats. J Neural Transm 2007; 114: 893-898.

21. Guidotti A, Auta J, Davis JM, Dong E, Grayson DR, Veldic M, Zhang X, Costa E. GABAergic dysfunction in schizophrenia: New treatment strategies on the horizon.

Psychopharmacology 2005; 180: 191-205.

22. Deutsch SI, Mastropaolo J, Hitri A. GABA-active steroids: Endogenous modulators of GABA-gated chloride ion conductance. Clinical Neuropharmacology 1992; 15: 352-364.

23. Deutsch SI, Rosse RB, Lakshman RM. Dysregulation of tau phosphorylation is a hypothesized point of convergence in the pathogenesis of Alzheimer’s disease, frontotemporal dementia and schizophrenia with therapeutic implications. Prog Neuropsychopharmacol Biol Psychiatry 2006; 30: 1369-1380.

24. Grayson DR, Jia X, Chen Y, Sharma RP, Mitchell CP, Guidotti A, Costa E. Reelin promoter hypermethylation in schizophrenia. Proc Natl Acad Sci USA 2005; 102: 9341-9346.

25. Veldic M, Guidotti A, Maloku E, Davis JM, Costa E. In psychosis, cortical interneurons overexpress DNA-methyltransferase 1. Proc Natl Acad Sci USA 2005; 102: 2152-2157.

26. Deutsch SI, Rosse RB, Mastropaolo J, Long KD, Gaskins BL. Epigenetic therapeutic strategies of neuropsychiatric disorders: Ready for prime time? Clin Neuropharmacol 2008; 31: 104-119.

27. Tochigi M, Iwamoto K, Bundo M, Komori A, Sasaki T, Kato N, Kato T. Methylation status of the reelin promoter region in the brain of schizophrenic patients. Biol Psychiatry 2008; 63: 530-533.

28. Adler LE, Olincy A, Waldo M, Harris JG, Griffith J, Stevens K, Flach K, Nagamoto H, Bickford P, Leonard S, Freedman R. Schizophrenia, sensory gating, and nicotine receptors.

Schizophr Bull 1998; 24: 189-202.

29. Coyle JT. The glutamatergic dysfunction hypothesis for schizophrenia. Harvard Rev Psychiatry 1996; 3: 241-253.

30. Deutsch SI, Rosse RB, Schwartz BL, Weizman A, Chilton M, Arnold DS, Mastropaolo J. Therapeutic implications of a selective α7 nicotinic receptor abnormality in schizophrenia. Isr J Psychiatry 2005; 42: 33-44.

31. Mike A, Castro NG, Albuquerque EX. Choline and acetylcholine have similar kinetic properties of activation and desensitization on the α7 nicotinic receptors in rat hippocampal neurons. Brain Res 2000; 882: 155-168.

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32. Stevens KE, Freedman R, Collins AC, Hall M, Leonard S, Marks MJ, Rose GM. Genetic correlation of hippocampal auditory evoked response and α-bungarotoxin binding in inbred mouse strains. Neuropsychopharmacology 1996; 15: 152-162.

33. Stevens KE, Kem WR, Mahnir VM, Freedman R. Selective α7nicotinic agonists normalize inhibition of auditory response in DBA mice. Psychopharmacology 1998; 136: 320-327.

34. Tamminga CA. Schizophrenia and glutamatergic transmission. Crit Rev Neurobiol 1998; 12: 21-36.

35. Olszewski RT, Wegorzewska MM, Monteiro AC, Krolikowski KA, Zhou J, Kozikowski AP, Long K, Mastropaolo J, Deutsch SI, Neale JH. PCP and MK-801 induced behaviors reduced

by NAAG peptidase inhibition via metabotropic glutamate receptors. Biol Psychiatry 2008; 63: 86-91.

36. Deutsch SI, Schwartz BL, Rosse RB, Mastropaolo J, Marvel CL, Drapalski AL. Adjuvant topiramate administration: A pharmacologic strategy for addressing NMDA receptor hypofunction in schizophrenia. Clin Neuropharmacol 2003; 26: 199-206.

37. Gibbs JW, Sombati S, DeLorenzo RJ, Coulter DA. Cellular actions of topiramate: Blockade of kainate-evoked inward currents in cultured hippocampal neurons. Epilepsia 2000; 41: S10-S16.

38. Shank RP, Gardocki JF, Streeter AJ, Maranoff, BE. An overview of the preclinical aspects of topiramate: Pharmacology, pharmacokinetics, and mechanism of action. Epilepsia 2000; 41: S3-S9.

39. White HS, Brown SD, Woodhead JH, Skeen GA, Wolf HH. Topiramate modulates GABA-evoked currents in murine cortical neurons by a nonbenzodiazepine mechanism.

Epilepsia 2000; 41: S17-S20.

40. Albuquerque EX, Alkondon M, Pereira EFR, Castro NG, Schrattenholz A, Barbosa CTF, Bonfante-Cabarcas R, Aracava Y, Eisenberg HM, Maelicke A. Properties of neuronal nicotinic acetylcholine receptors: Pharmacological characterization and modulation of synaptic function. J Pharmacol Exp Ther 1997; 280: 1117-1136.

41. Leonard S, Gault J, Hopkins J, Logel J, Vianzon R, Short M, Drebing C, Berger R, Venn D, Sirota P, Zerbe G, Olincy A, Ross RG, Adler LE, Freedman R. Association of promoter variants in the α7 nicotinic acetylcholine receptor subunit gene with an inhibitory deficit found in schizophrenia.

Arch Gen Psychiatry 2002; 59: 1085-1096.

42. Rosse RB, Deutsch SI. Adjuvant galantamine administration improves negative symptoms in a patient with treatment-refractory schizophrenia. Clin Neuropharmacol 2002; 25: 272-275.

43. Wilson RI, Nicoll RA. Endogenous cannabinoids mediate retrograde signaling at hippocampal synapses. Nature 2001; 410: 588-592.

Elaine Waddington Lamont et al.

Circadian Rhythms and Clock Genes in Psychotic Disorders Elaine Waddington Lamont, PhD,1,2 Daniel L. Coutu, MSc,3 Nicolas Cermakian, PhD,2 and Diane B. Boivin, MD, PhD1 1

Centre for Study and Treatment of Circadian Rhythms, Douglas Mental Health University Institute, Faculty of Medicine, McGill University, Montreal, QC, Canada 2 Laboratory of Molecular Chronobiology, Douglas Mental Health University Institute, Department of Psychiatry, McGill University, Montreal, QC, Canada 3 Lady Davis Institute for Medical Research, McGill University, Montreal, QC, Canada

ABSTRACT Numerous lines of evidence suggest that a disordered circadian system contributes to the etiology and symptomatology of major psychiatric disorders.

Sleep disturbances, particularly rapid eye movement (REM) sleep, have been observed in bipolar affective disorder (BPD) and schizophrenia. Therapies aimed at altering the timing and duration of sleep and realigning circadian rhythms, including sleep scheduling, wake extension, light therapy and drug therapies that alter sleep and circadian rhythms appear beneficial for affective disorders. Interventional studies aiming to correct sleep and circadian disturbances in schizophrenia are scarce, although exogenous melatonin has been shown to improve both sleep structure and psychotic symptoms. The study of molecular clock mechanisms in psychiatric disorders is also gaining interest. Genetics studies have found associations with CLOCK, PERIOD1, PERIOD3, and TIMELESS in schizophrenia. Most research on BPD has focused on polymorphisms of CLOCK, but the lithium target GSK-3 may also be significant. New research examining the role of circadian rhythms and clock genes in major mental illness is likely to produce rapid advances in circadian-based therapeutics.

Introduction The circadian system plays a fundamental role in overall health and longevity (1). This is also true for mental dis-

orders since misalignment between the endogenous circadian system and the sleep/wake cycle is a critical factor in the clinical status of many psychiatric disorders (2, 3). This review examines the evidence for circadian disturbances in severe psychiatric disorders such as chronic schizophrenia and bipolar affective disorder (BPD), describes circadianrelated interventions that have been used successfully to treat these disorders, and discusses current research on the role of clock genes in mental illness.

Sleep and Circadian Rhythms in Psychotic Disorder Circadian rhythms, sleep and schizophrenia

Kant, Schopenhauer and Hughlings Jackson (4) all underlined the similarity between dreaming experiences and hallucinations. However, it was not until the discovery of rapid-eye movement (REM) sleep by Aserinsky and Kleitman (5) that the study of “dreaming sleep” and, in turn, REM sleep-related disorders became possible.

Early studies examining the effects of sleep deprivation in schizophrenia patients (6, 7) were based on the hypothesis that hallucinations may be caused by REMlike intrusions into wakefulness, and by the observation that prolonged sleep deprivation induced hallucinations and psychotic symptoms reminiscent of schizophrenia (8). These pioneering experiments demonstrated an absence of REM sleep rebound after REM sleep specific deprivation, but no other abnormalities specific to this sleep stage. Later studies did not provide a consensus regarding other sleep parameters, presumably because of sampling variability and medication status of the subjects (see below). However, some abnormal polysomno-

Address for Correspondence: Diane B. Boivin, MD, PhD, Centre for Study and Treatment of Circadian Rhythms, Douglas Mental Health University Institute, 6875 LaSalle Boulevard, suite F-1127, Montréal, Québec, Canada H4H 1R3. diane.boivin@douglas.mcgill.ca

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CIRCADIAN RHYTHMS AND CLOCK GENES IN PSYCHOTIC DISORDERS

graphic measures do emerge when drug-free subjects are analyzed, and, importantly, these abnormalities appear to be also present in large numbers of medicated subjects (2, 9, 10). These include poor sleep initiation and consolidation, impaired sleep homeostasis expressed as low levels of slow-wave sleep (SWS), with many subjects showing a total absence of stage 4 sleep, and shortened REM sleep latency with frequent sleep onset REM periods. Taken together, these observations suggest a deficient homeostatic regulation of sleep, although sleep deprivation (SD) can lead to SWS rebound on recovery nights (10).

The shortened REM sleep latency, aberrant rest-activity cycles, and blunted melatonin secretion, on the other hand, suggest that abnormalities of the circadian system cannot be completely ruled out.

As with sleep, circadian disturbances have been reported in schizophrenia patients, but the results are inconsistent. One study has measured core body temperature (CBT) variation and reported desynchronization of temperature, pulse and blood pressure rhythms, although this study was conducted under ambulatory conditions and the data influenced by masking effects (11). In some experiments, the analysis of melatonin secretion demonstrated blunted circadian variation, although uncontrolled light exposure prior to data collection may have confounded these results (12-14). Others have reported phase advances of body temperature (15), prolactin and melatonin (16). These advanced rhythms are surprising considering that actigraphic recordings have revealed disturbed rest-activity cycles that are often inconsistent with a phase advance, including phase delays, longer periods of activity or, occasionally, 48 hour rest-activity patterns (17-19). Patients with schizophrenia/schizoaffective disorders also show a greater tendency towards eveningness (later wake and bed times and being most alert later in the day) than controls (20).

The observation that schizophrenia is commonly associated with disturbed sleep/wake cycles supports the hypothesis that these pathologies may emerge from a common underlying neuropathology. Interestingly, atypical antipsychotics can relieve both psychotic and sleep-related abnormalities (9). The dopaminergic hypothesis of schizophrenia postulates that hyperdopaminergic activity from the mesocorticolimbic systems causes positive symptoms. However, because the activity of atypical antipsychotics is associated with serotoninergic and cholinergic pathways, a monoaminergiccholinergic imbalance has also been hypothesized (21).

Cholinergic neurons from the basal forebrain, peduncu28

lopontine tegmentum and laterodorsal tegmentum play key roles in both the arousal and REM sleep systems (22).

Importantly, their activity modulates sensory processing and deregulation of these systems (particularly those of the midbrain) could cause hallucinations, as seen in schizophrenia (23). Whereas the role of dopamine in sleep-wake regulation has long been ignored, recent evidence demonstrated that hyperdopaminergic transgenic mice showed REM-like intrusions during wakefulness, an effect that could be blocked by haloperidol (24).

These intrusions were exacerbated when the animals were emotionally challenged. Further studies are needed to elucidate the role of the common neurotransmission pathways possibly involved in both schizophrenia and sleep or circadian regulation of sleep-wake states. The lack of consensus across studies on the sleep-related or circadian abnormalities in schizophrenia calls for better controlled studies with standardized experimental protocols. The fact that sleep-wake disorders may play a role in the development of the disease could also lead to novel therapeutic avenues for schizophrenia, including light therapy (25) and melatonin agonists (14, 26).

Circadian rhythms, sleep and bipolar affective disorder

The rhythmic nature of BPD, which is exaggerated in so-called â&#x20AC;&#x153;rapid cyclersâ&#x20AC;? who show rapid mood changes from depression to mania or hypomania, has long invited speculation that the endogenous circadian system may play a role in the etiology of this disorder (27). Circadian disturbances have been observed in BPD such as a phase advance of the diurnal rhythm of plasma melatonin (28) and plasma cortisol (28, 29), although negative results have also been reported (30). Sleep disturbances are a defining symptom of BPD, with insomnia or hypersomnia, early morning awakening, reduced sleep efficiency, and altered REM sleep latency being the most consistently reported (2). The relationship between the sleep-wake cycle and changes in mood appear to be important in BPD, with the switch from mania or hypomania to depression or euthymia often occurring during or after sleep, while positive changes in mood from depression to hypomania or mania are more likely to occur after a period of wakefulness (31, 32). Sleep duration also appears to be critical, as sleep restriction predicts the onset of mania or hypomania the following day (33), while sleep extension is often associated with depression (33, 34).

As with schizophrenia, BPD patients score higher on eveningness scales than controls, although age could

Elaine Waddington Lamont et al.

play a role in this association (20). Interestingly, a classification of “evening type” was associated with greater severity of BPD including seeking treatment at an earlier age and a greater likelihood of rapid-cycling mood changes. A major limitation of this study is that mood state at the time of chronotype assessment is unknown (20). Interestingly, phase advance of sleep timing has been found to be effective in the treatment of BPD patients during the depressive phases (35, 36). Further studies will be needed to clarify the association between chronotype and BPD and controversies persist since the therapeutic action of lithium in BPD is associated with lengthening of the circadian period (described below) (37-39).

A number of interventions related to sleep and circadian rhythms have been successfully used to treat BPD. Primary interventions are bright light exposure and SD therapy, either partial or total, scheduled during the depressive phases of the illness. The advantage of these therapies is that they are non-invasive, fast acting, and found to be effective in patients with major depressive disorder and BPD (35, 36, 40-42). The main limitations of SD are a short duration of action, a high relapse rate, and the risk of triggering a manic or hypomanic episode (33, 43). Light therapy has been used to treat BPD, most frequently in combination with mood stabilizers and during the depressive phases, but more studies are needed to clarify its therapeutic utility and caution should be exercised because of the possibility of mood instability and relapses into hypomania (43, 44).

While an improved mood score is unambiguous in a patient with unipolar depression, in BPD it may in fact be an early sign of mania or hypomania (40). One way of achieving the benefits of SD while avoiding the drawbacks is to use it in combination with light or pharmacotherapy (45, 46). For example, Colombo and colleagues (45) found that the acute effects of total SD on mood were sustained when combined with lithium or light therapy, but there were no additive effects when all three treatments were combined. Phase advance of sleep timing has also been found to be effective in BPD patients during the depressive phase of the illness (35, 36). While these results seem promising, both SD and light therapy must be used with caution in BPD because of the potential for mood instability and relapses into hypomania or mania (33, 43). Interestingly, extended periods of sleep/darkness have been used to prevent/treat mania (47, 48). These studies suggest that, as part of a judicious sleep hygiene program, chronotherapeutic interventions such as SD and bright light exposure during depressive phases and

sleep/darkness stabilization during manic/hypomanic phases can be used in BPD (49).

Exogenous melatonin and melatonin agonists have received attention recently as potential treatments for depression, and may also be of benefit in BPD. Lower baseline plasma melatonin levels (50), and greater sensitivity to light-induced melatonin suppression, have been reported in BPD patients compared to controls (51), although some controversies persist (50). Lithium, an effective mood stabilizer for BPD, was shown to reduce melatonin suppression by light in healthy controls (52) and changes in melatonin levels may accompany successful treatment in BPD patients. The few studies to date that have tested the therapeutic effects of exogenous melatonin in BPD have been disappointing as melatonin was not effective (51). However, preliminary results of the effect of the melatonin agonist agomelatine, used with lithium or valproate, seem promising (53), but larger studies are needed.

Clock Genes and Psychotic Disorders Basic molecular mechanisms of circadian rhythms

The molecular mechanisms underlying circadian rhythmicity are comprised of positive and negative transcriptional/translational feedback loops and post-transcriptional regulatory elements. The genes Clock and Bmal1 encode the transcription factors CLOCK and BMAL1(54-56), which together activate transcription of three Period (Per) and two Cryptochrome (Cry) genes (57-62), Rorα and Rev-Erbα (63-65). The proteins PER and CRY combine to inhibit their own transcription, while RORα and REV-ERBα act on Bmal1 to activate and inhibit transcription, respectively. Post transcriptional modifications of PERs and CRYs by the enzymes casein kinase I epsilon (CKIε) and delta (CKIδ) (66-68), and possibly the Drosophila SHAGGY homologue glycogen synthase kinase-3 (GSK-3) (69) control the rate of accumulation, association and translocation of PER and CRY (70), which can alter the period and phase of the molecular clock. NPAS2 is an alternate dimerization partner for BMAL1 that does not seem to be abundant in the master of the circadian clock, the suprachiasmatic nucleus (SCN), but may be important for circadian function in the forebrain (71, 72). TIMELESS (TIM) is similar to the fruit fly clock gene tim. Although the latter is essential for the fly clock, its role in the mammalian clock has been subject to controversy. Mammalian TIM appears to have a role in the clock of the SCN (73), but also in early 29

CIRCADIAN RHYTHMS AND CLOCK GENES IN PSYCHOTIC DISORDERS

embryonic development (74) and DNA replication (75).

Mutations of any of these circadian genes can potentially have an impact on the circadian clock, and thus subtly or dramatically alter sleep, mood or behavior in ways that contribute to physical and mental illness.

All these clock genes have been identified in humans, and there is a growing body of literature reporting their expression in human tissues (reviewed in 2).

These studies have enabled the tracking of circadian rhythms in peripheral tissues and extra-SCN brain regions under various conditions, including shift work (76), Alzheimerî&#x2C6;&#x20AC;s disease (77, 78) and various lighting manipulations (2, 79, 80).

Clock genes in non-psychiatric disorders

The first demonstration of the importance of the human molecular clock was related to extreme sleep timing disorders. On one hand, an amino acid substitution was shown to explain the extremely early sleep and wake times (bedtime = 6-9 pm, wake = 1-3 am) of two familial cases of advance sleep phase disorder (FASPD). These mutations were in the PER2 and CKIδ genes (81-84).

On the other hand, delayed sleep phase disorder (DSPD; bedtime = 3-6 am, wake = 1-3 pm) was found to be associated with a polymorphism of human PER3 (85, 86).

PER3 polymorphisms have also been associated with less extreme chronotypes. Individuals with the shorter 4-repeat allele of PER3 were found more frequently in evening types, while the longer 5-repeat was more frequently seen in morning types, although this association was only significant in younger individuals (18-29 years old) (87). Interestingly, recent data suggest that this polymorphism of PER3 might impact sleep homeostasis; individuals with the longer allele PER35/5 showed increased SWS and electroencephalographic (EEG) slowing, but the circadian rhythms of melatonin, cortisol and peripheral PER3 expression were not affected (88). In addition, the rhythm of PER3 expression in white blood cells, but not that of PER2 or BMAL1, is significantly correlated with the timing of the sleep/wake rhythms and rhythms of plasma melatonin and cortisol secretion, indicating PER3 as a potential genetic marker for circadian mechanisms underlying sleep/wake timing (89).

The PER and CKI genes are not the only clock genes that have an influence on chronotype. Morningnesseveningness scores have been found to be associated with a single nucleotide polymorphism (SNP) of the human CLOCK gene (90, 91). Individuals with the Clock 3111C/C and C/T alleles showed increased evening30

ness and reduced morningness, while 3111T/T subjects showed higher morningness scores (90, 91), but conflicting results have been reported (92). This may be related to differences in allelic frequencies in different ethnic groups. In a Korean population, there were no individuals carrying the 3111C/C, but individuals with the C/T allele were more likely to show a diurnal preference for eveningness when they also carried a 825C/T SNP of the GNB3 gene (93). The 3111C/C genotype is also associated with delayed sleep timing and greater daytime sleepiness in a Japanese population (91), but not in Caucasians (92).

There is currently no evidence in support of an association between the 3111C/C geneotype and DSPD or non24 hour sleep phase disorder (92, 94).

Clock genes and schizophrenia

There are few studies demonstrating a link between circadian clock gene polymorphisms or deregulation and schizophrenia. However, the CLOCK 3111C/T polymorphism showed a transmission bias in a sample of 145 Japanese schizophrenic subjects relative to healthy controls (95). The authors suggested that this polymorphism, associated with aberrant dopaminergic transmission to the SCN, may underlie the pathophysiology of schizophrenia. Since dopaminergic signalling through D2 receptors is associated with increased CLOCK:BMAL1 activity (96), this provides an interesting link between the dopaminergic hypothesis of schizophrenia and circadian abnormalities in these patients. Another study used microarray technology to analyze gene expression in the temporal lobe of postmortem brains obtained at autopsy from patients previously diagnosed with schizophrenia (97). The results demonstrated decreased expression of the PER1 mRNA in schizophrenia patients compared with age-matched normal controls. These findings were also confirmed by RT-PCR. Association of PER3 and TIM with schizophrenia/schizoaffective disorder, as well as with bipolar disorder have also been found (98). The association with PER3 is interesting, given the evidence of a relationship between PER3 with delayed sleep phase disorder and evening chronotype. However, the function of Tim in mammals is not entirely clear (73), making it difficult to interpret this finding. Finally, the CRY1 gene was hypothesized to be a candidate gene for schizophrenia based on its location near a linkage hotspot for schizophrenia on chromosome 12q24 (99). The fact that CRY1 is expressed in dopaminergic cells in the retina (100) and that its expression influences the effects of psychoactive drugs (101) lends further supports to this hypothesis.

Elaine Waddington Lamont et al.

Clock genes and bipolar disorder

Of all major mental disorders, the evidence for genetic abnormalities associated with clock genes is strongest in BPD. An analysis of 46 SNPs of eight clock genes (BMAL1, CLOCK, PER 1,2,3, CRY 1,2, TIM) revealed association of BMAL1 and TIM with BPD using familybased samples with BPD or schizophrenia (98). Even though these were modest associations found using very liberal analyses, the association with BMAL1 has been independently confirmed using haplotype analysis (102).

The same study also demonstrated an association with PER3. Studies examining other genes have found negative results: screening for human PER2 mutations at the CKIδ/ε binding site showed no difference in frequency between BPD patients and controls (103), nor was there any evidence for linkage or association of CRY1 with BPD (104). Interestingly, a recent study pointed to a general reduction in the amplitude of clock gene mRNAs in fibroblasts taken from BPD patients (105).

One group has published numerous studies on the association of the CLOCK 3111T/C polymorphism with BPD (106-109). The C/C allele has been associated with greater severity of insomnia during antidepressant treatment (109) and a higher recurrence rate of bipolar episodes (107), reduced need for sleep (108), and a tendency to increased activity during the later part of the day prior to sleep (all C allele carriers,(110). There is even some suggestion for differential activation in the cingulate cortex during a moral decision-making task, dependent on the C/T genotype of major depressive disorder and BPD patients who underwent fMRI and testing during a depressive episode (110).

Support for a role of CLOCK mutation in bipolar disorder has recently come from the animal literature, with evidence that the Clock mutant mouse might constitute an animal model of mania (111).

Recent evidence suggests that the therapeutic action of lithium may be related to direct effects on the circadian clock. For example, lithium has been shown to lengthen the period of circadian rhythms in rodents (39), and can lengthen the period of neuronal firing of cultured SCN neurons in a dose dependent manner (37). A delay in the circadian phase markers, body temperature and REM sleep has also been shown in a BPD patient (38).

One proposed mechanism of the therapeutic action of lithium is via the inhibition of GSK-3 (69, 112). Although this enzyme has a number of functions that could potentially mediate the therapeutic effects of lithium (113), one possibility is via its function as a central regulator of the circadian clock (112). Numerous lines of evidence

support this idea. For example, both lithium and Gsk-3 knockdown produce lengthening of the period of Per2 mRNA transcription rhythms in mouse fibroblasts (114) and GSK-3β affects the entry of Per2 into the nucleus (69). Rev-Erbα protein expression is dramatically reduced by lithium, via the inhibition of GSK-3β, but Rev-Erbα RNA is unaffected (115). This suggests that the inhibition of GSK-3 by lithium has multiple effects on key clock components. Even more interesting are findings that inhibition of GSK-3 may be common to other mood stabilizing agents such as valproate and may even be a target of antidepressant therapies including drugs which target the serotonergic and dopaminergic systems as well as electroconvusive therapy (112). The above-mentioned study examining circadian gene expression in fibroblasts from BPD patients also identified group differences in GSK-3β (105). This makes GSK-3 a promising target for future development of pharmotherapeutic agents.

Conclusion The study of psychiatric populations is difficult and the literature is rife with inconsistencies. Lack of replication, or even contradiction in results between studies may be due to a number of factors, including heterogeneous patient populations, differences in diagnostic criteria, use of medications, insufficient withdrawal periods from medications before testing, and lack of control for variations in light-dark cycles. In spite of these difficulties, well-controlled studies in psychiatric populations must be pursued in order to increase our knowledge of the clinical repercussions of sleep and circadian rhythms disturbances in these populations. Health care professionals should be better trained on the potential diagnostic utility and consequences of changes in sleep-wake patterns for their psychiatric patients. Interventions aimed at correcting disturbed circadian rhythms and/or rest-activity cycles have resulted in effective, well-tolerated therapies used either alone or in conjunction with traditional pharmacotherapies (116).

Finally, advances in the circadian genetics of mental illness are likely to open a new frontier of genetic therapies, as well as guide the development of new pharmaceuticals.

Acknowledgements: Supported by the National Alliance for Research on Schizophrenia and Depression, the Canadian Psychiatric Research Foundation, the Levinschi Foundation, the Canadian Institutes of Health Research, the “Institut de recherche Robert-Sauvé en santé et en sécurité du travail du Québec,” and the “Fonds de la Recherche en Santé du Québec.” We are grateful to Dr. Valérie Mongrain for her editorial comments.

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References 1. Van Someren EJ, Riemersma-Van Der Lek RF. Live to the rhythm, slave to the rhythm. Sleep Med Rev 2007;11:465484.

2. Boivin DB. Influence of sleep-wake and circadian rhythm disturbances in psychiatric disorders. J Psychiatry Neurosci 2000;25:446-458.

3. Lamont EW, James FO, Boivin DB, Cermakian N. From circadian clock gene expression to pathologies. Sleep Med 2007;8:547-556.

4. Gottesmann C. The dreaming sleep stage: A new neurobiological model of schizophrenia? Neuroscience 2006;140:1105-1115.

5. Aserinsky E, Kleitman N. Regularly occurring periods of eye motility, and concomitant phenomena, during sleep.

Science 1953;118:273-274.

6. Zarcone V, Azumi K, Dement W, Gulevich G, Kraemer H, Pivik T. REM phase deprivation and schizophrenia II. Arch Gen Psychiatry 1975;32:1431-1436.

7. Zarcone V, Gulevich G, Pivik T, Dement W. Partial REM phase deprivation and schizophrenia. Arch Gen Psychiatry 1968;18:194-202.

8. Bliss EL, Clark LD, West CD. Studies of sleep deprivation-relationship to schizophrenia. AMA Arch Neurol Psychiatry 1959;81:348-359.

9. Benson KL, Zarcone VP, Jr. Schizophrenia. In: Kryger MH, Roth T, Dement WC, editors. Principles and practice of sleep medicine. 4th ed. Philadelphia, Penn.: Elsevier/ Saunders, 2005.

10. Chouinard S, Poulin J, Stip E, Godbout R. Sleep in untreated patients with schizophrenia: A meta-analysis.

Schizophr Bull 2004;30:957-967.

11. Madjirova NP, Petrova NS, Delchev NK. Daily rhythmicity of temperature, pulse and blood pressure in schizophrenic patients. Schizophr Res 1995;14:183.

16. Rao ML, Gross G, Strebel B, et al. Circadian rhythm of tryptophan, serotonin, melatonin, and pituitary hormones in schizophrenia. Biol Psychiatry 1994;35:151-163.

17. Wirz-Justice A, Cajochen C, Nussbaum P. A schizophrenic patient with an arrhythmic circadian rest-activity cycle.

Psychiatry Res 1997;73:83-90.

18. Wirz-Justice A, Haug HJ, Cajochen C. Disturbed circadian rest-activity cycles in schizophrenia patients: An effect of drugs? Schizophr Bull 2001;27:497-502.

19. Wulff K, Joyce E, Middleton B, Dijk DJ, Foster RG. The suitability of actigraphy, diary data, and urinary melatonin profiles for quantitative assessment of sleep disturbances in schizophrenia: A case report. Chronobiol Int 2006;23:485-495.

20. Mansour HA, Wood J, Chowdari KV, et al. Circadian phase variation in bipolar I disorder. Chronobiol Int 2005;22:571-584.

21. Tandon R. Cholinergic aspects of schizophrenia. Br J Psychiatry Suppl 1999;37:7-11.

22. Jones B. Basic mechanisms of sleep-wake states. In: Kryger MH, Roth T, Dement WC, editors. Principles and practice of sleep medicine. 4th ed. Philadelphia, Penn.: Elsevier/Saunders, 2005.

23. Behrendt RP. Hallucinations: Synchronisation of thalamocortical gamma oscillations underconstrained by sensory input. Conscious Cogn 2003;12:413-451.

24. Dzirasa K, Ribeiro S, Costa R, et al. Dopaminergic control of sleep-wake states. J Neurosci 2006;26:10577-10589.

25. Abbott A. Restless nights, listless days. Nature 2003;425: 896-898.

26. Katz G, Durst R, Knobler HY. Exogenous melatonin, jet lag, and psychosis: Preliminary case results. J Clin Psychopharmacol 2001;21:349-351.

27. Wirz-Justice A. Biological rhythm disturbances in mood disorders. Int Clin Psychopharmacol 2006;21 :S11-15.

12. Bersani G, Mameli M, Garavini A, Pancheri P, Nordio M. Reduction of night/day difference in melatonin blood levels as a possible disease-related index in schizophrenia. Neuro Endocrinol Lett 2003;24:181-184.

28. Beck-Friis J, Ljunggren JG, Thoren M, von Rosen D, Kjellman BF, Wetterberg L. Melatonin, cortisol and ACTH in patients with major depressive disorder and healthy humans with special reference to the outcome of the dexamethasone suppression test. Psychoneuroendocrinology 1985;10:173-186.

13. Monteleone P, Maj M, Fusco M, Kemali D, Reiter RJ. Depressed nocturnal plasma melatonin levels in drug-free paranoid schizophrenics. Schizophr Res 1992;7:77-84.

29. Linkowski P, Kerkhofs M, Van Onderbergen A, et al. The 24-hour profiles of cortisol, prolactin, and growth hormone secretion in mania. Arch Gen Psychiatry 1994;51:616-624.

14. Shamir E, Laudon M, Barak Y, et al. Melatonin improves sleep quality of patients with chronic schizophrenia. J Clin Psychiatry 2000;61:373-377.

30. Cervantes P, Gelber S, Kin FN, Nair VN, Schwartz G. Circadian secretion of cortisol in bipolar disorder. J Psychiatry Neurosci 2001;26:411-416.

15. Morgan R, Cheadle AJ. Circadian body temperature in chronic schizophrenia. Br J Psychiatry 1976;129:350-354.

31. Feldman-Naim S, Turner EH, Leibenluft E. Diurnal variation in the direction of mood switches in patients with

32

Elaine Waddington Lamont et al.

rapid-cycling bipolar disorder. J Clin Psychiatry 1997; 58: 79-84.

depression: Replication of main effects and interaction.

Psychiatry Res 2000;95:43-53.

32. Voderholzer U, Weske G, Ecker S, Riemann D, Gann H, Berger M. Neurobiological findings before and during successful lithium therapy of a patient with 48-hour rapid-cycling bipolar disorder. Neuropsychobiology 2002;45:13-19.

46. Smeraldi E, Benedetti F, Barbini B, Campori E, Colombo C. Sustained antidepressant effect of sleep deprivation combined with pindolol in bipolar depression. A placebo-controlled trial. Neuropsychopharmacology 1999;20:380-385.

33. Leibenluft E, Albert PS, Rosenthal NE, Wehr TA. Relationship between sleep and mood in patients with rapid-cycling bipolar disorder. Psychiatry Res 1996;63:161-168.

34. Wehr TA. Sleep loss: A preventable cause of mania and other excited states. J Clin Psychiatry 1989;50, Suppl:8-16; discussion 45-47.

35. Benedetti F, Barbini B, Campori E, Fulgosi MC, Pontiggia A, Colombo C. Sleep phase advance and lithium to sustain the antidepressant effect of total sleep deprivation in bipolar depression: New findings supporting the internal coincidence model? J Psychiatr Res 2001;35:323-329.

36. Wehr TA, Wirz-Justice A, Goodwin FK, Duncan W, Gillin JC. Phase advance of the circadian sleep-wake cycle as an antidepressant. Science 1979;206:710-713.

37. Abe M, Herzog ED, Block GD. Lithium lengthens the circadian period of individual suprachiasmatic nucleus neurons. Neuroreport 2000;11:3261-3264.

38. Campbell SS, Gillin JC, Kripke DF, Janowsky DS, Risch SC. Lithium delays circadian phase of temperature and REM sleep in a bipolar depressive: A case report.

Psychiatry Res 1989;27:23-29.

47. Barbini B, Benedetti F, Colombo C, et al. Dark therapy for mania: A pilot study. Bipolar Disord 2005;7:98-101.

48. Wehr TA, Turner EH, Shimada JM, Lowe CH, Barker C, Leibenluft E. Treatment of rapidly cycling bipolar patient by using extended bed rest and darkness to stabilize the timing and duration of sleep. Biol Psychiatry 1998;43:822-828.

49. Leibenluft E, Suppes T. Treating bipolar illness: Focus on treatment algorithms and management of the sleep-wake cycle. Am J Psychiatry 1999;156:1976-1981.

50. Lam RW, Berkowitz AL, Berga SL, Clark CM, Kripke DF, Gillin JC. Melatonin suppression in bipolar and unipolar mood disorders. Psychiatry Res 1990;33:129-134.

51. Srinivasan V, Smits M, Spence W, et al. Melatonin in mood disorders. World J Biol Psychiatry 2006;7:138-151.

52. Hallam KT, Olver JS, Horgan JE, McGrath C, Norman TR. Low doses of lithium carbonate reduce melatonin light sensitivity in healthy volunteers. Int J Neuropsychopharmacol 2005;8:255-259.

53. Gao K, Calabrese JR. Newer treatment studies for bipolar depression. Bipolar Disord 2005;7:13-23.

39. LeSauter J, Silver R. Lithium lengthens the period of circadian rhythms in lesioned hamsters bearing SCN grafts.

Biol Psychiatry 1993;34:75-83.

54. Cermakian N, Boivin DB. A molecular perspective of human circadian rhythm disorders. Brain Res Brain Res Rev 2003;42:204-220.

40. Barbini B, Colombo C, Benedetti F, Campori E, Bellodi L, Smeraldi E. The unipolar-bipolar dichotomy and the response to sleep deprivation. Psychiatry Res 1998;79:43-50.

55. Dardente H, Cermakian N. Molecular circadian rhythms in central and peripheral clocks in mammals. Chronobiol Int 2007;24:195-213.

41. Leibenluft E, Wehr TA. Is sleep deprivation useful in the treatment of depression? Am J Psychiatry 1992;149:159-168.

56. Ko CH, Takahashi JS. Molecular components of the mammalian circadian clock. Hum Mol Genet 2006;15:R271-277.

42. Wu JC, Bunney WE. The biological basis of an antidepressant response to sleep deprivation and relapse: Review and hypothesis. Am J Psychiatry 1990;147:14-21.

57. Albrecht U, Sun ZS, Eichele G, Lee CC. A differential response of two putative mammalian circadian regulators, mper1 and mper2, to light. Cell 1997;91:1055-1064.

43. Leibenluft E, Turner EH, Feldman-Naim S, Schwartz PJ, Wehr TA, Rosenthal NE. Light therapy in patients with rapid cycling bipolar disorder: preliminary results.

Psychopharmacol Bull 1995;31:705-710.

58. Albrecht U, Zheng B, Larkin D, Sun ZS, Lee CC. MPer1 and mper2 are essential for normal resetting of the circadian clock. J Biol Rhythms 2001;16:100-104.

44. Sit D, Wisner KL, Hanusa BH, Stull S, Terman M. Light therapy for bipolar disorder: A case series in women.

Bipolar Disord 2007;9:918-927.

45. Colombo C, Lucca A, Benedetti F, Barbini B, Campori E, Smeraldi E. Total sleep deprivation combined with lithium and light therapy in the treatment of bipolar

59. Gekakis N, Staknis D, Nguyen HB, et al. Role of the CLOCK protein in the mammalian circadian mechanism. Science 1998;280:1564-1569.

60. Hogenesch JB, Gu YZ, Jain S, Bradfield CA. The basichelix-loop-helix-PAS orphan MOP3 forms transcriptionally active complexes with circadian and hypoxia factors.

Proc Natl Acad Sci USA 1998;95:5474-5479.

33

CIRCADIAN RHYTHMS AND CLOCK GENES IN PSYCHOTIC DISORDERS

61. Kume K, Zylka MJ, Sriram S, et al. mCRY1 and mCRY2 are essential components of the negative limb of the circadian clock feedback loop. Cell 1999;98:193-205.

62. Shearman LP, Sriram S, Weaver DR, et al. Interacting molecular loops in the mammalian circadian clock. Science 2000;288:1013.

63. Preitner N, Damiola F, Lopez-Molina L, et al. The orphan nuclear receptor REV-ERBalpha controls circadian transcription within the positive limb of the mammalian circadian oscillator. Cell 2002;110:251-260.

64. Sato TK, Panda S, Miraglia LJ, et al. A functional genomics strategy reveals Rora as a component of the mammalian circadian clock. Neuron 2004;43:527-537.

65. Ueda HR, Chen W, Adachi A, et al. A transcription factor response element for gene expression during circadian night. Nature 2002;418:534-539.

66. Akashi M, Tsuchiya Y, Yoshino T, Nishida E. Control of intracellular dynamics of mammalian period proteins by casein kinase I epsilon (CKIepsilon) and CKIdelta in cultured cells. Mol Cell Biol 2002;22:1693-1703.

67. Eide EJ, Woolf MF, Kang H, et al. Control of mammalian circadian rhythm by CKIepsilon-regulated proteasomemediated PER2 degradation. Mol Cell Biol 2005;25:27952807.

68. Vielhaber E, Eide E, Rivers A, Gao ZH, Virshup DM. Nuclear entry of the circadian regulator mPER1 is controlled by mammalian casein kinase I epsilon. Mol Cell Biol 2000;20:4888-4899.

69. Iitaka C, Miyazaki K, Akaike T, Ishida N. A role for glycogen synthase kinase-3beta in the mammalian circadian clock. J Biol Chem 2005;280:29397-29402.

70. Gallego M, Virshup DM. Post-translational modifications regulate the ticking of the circadian clock. Nat Rev Mol Cell Biol 2007;8:139-148.

71. DeBruyne JP, Weaver DR, Reppert SM. CLOCK and NPAS2 have overlapping roles in the suprachiasmatic circadian clock. Nat Neurosci 2007;10:543-545.

72. Reick M, Garcia JA, Dudley C, McKnight SL. NPAS2: an analog of clock operative in the mammalian forebrain.

Science 2001;293:506-509.

73. Barnes JW, Tischkau SA, Barnes JA, et al. Requirement of mammalian Timeless for circadian rhythmicity. Science 2003;302:439-442.

74. Gotter AL, Manganaro T, Weaver DR, et al. A time-less function for mouse timeless. Nat Neurosci 2000;3:755756.

75. Gotter AL. A Timeless debate: resolving TIM᾿s noncircadian roles with possible clock function. Neuroreport 34

2006;17:1229-1233.

76. James FO, Cermakian N, Boivin DB. Circadian rhythms of melatonin, cortisol, and clock gene expression during simulated night shift work. Sleep 2007;30:1427-1436.

77. Ackermann K, Dehghani F, Bux R, Kauert G, Stehle JH. Day-night expression patterns of clock genes in the human pineal gland. J Pineal Res 2007;43:185-194.

78. Wu YH, Fischer DF, Kalsbeek A, et al. Pineal clock gene oscillation is disturbed in Alzheimer᾿s disease, due to functional disconnection from the “master clock.” Faseb J 2006;20:1874-1876.

79. Kusanagi H, Hida A, Satoh K, et al. Expression profiles of 10 circadian clock genes in human peripheral blood mononuclear cells. Neurosci Res 2008;61:136-142.

80. Tsinkalovsky O, Smaaland R, Rosenlund B, et al. Circadian variations in clock gene expression of human bone marrow CD34+ cells. J Biol Rhythms 2007;22:140-150.

81. Jones CR, Campbell SS, Zone SE, et al. Familial advanced sleep-phase syndrome: A short-period circadian rhythm variant in humans. Nat Med 1999;5:1062-1065.

82. Toh KL, Jones CR, He Y, et al. An hPer2 phosphorylation site mutation in familial advanced sleep phase syndrome.

Science 2001;291:1040-1043.

83. Xu Y, Padiath QS, Shapiro RE, et al. Functional consequences of a CKIdelta mutation causing familial advanced sleep phase syndrome. Nature 2005;434:640-644.

84. Xu Y, Toh KL, Jones CR, Shin JY, Fu YH, Ptacek LJ. Modeling of a human circadian mutation yields insights into clock regulation by PER2. Cell 2007;128:59-70.

85. Archer SN, Robilliard DL, Skene DJ, et al. A length polymorphism in the circadian clock gene Per3 is linked to delayed sleep phase syndrome and extreme diurnal preference. Sleep 2003;26:413-415.

86. Ebisawa T, Uchiyama M, Kajimura N, et al. Association of structural polymorphisms in the human period3 gene with delayed sleep phase syndrome. EMBO Rep 2001;2:342346.

87. Jones KH, Ellis J, von Schantz M, Skene DJ, Dijk DJ, Archer SN. Age-related change in the association between a polymorphism in the PER3 gene and preferred timing of sleep and waking activities. J Sleep Res 2007;16:12-16.

88. Viola AU, Archer SN, James LM, et al. PER3 polymorphism predicts sleep structure and waking performance.

Curr Biol 2007;17:613-618.

89. Archer SN, Viola AU, Kyriakopoulou V, von Schantz M, Dijk DJ. Inter-individual differences in habitual sleep timing and entrained phase of endogenous circadian rhythms of BMAL1, PER2 and PER3 mRNA in human leukocytes.

Elaine Waddington Lamont et al.

Sleep 2008;31:608-617.

90. Katzenberg D, Young T, Finn L, et al. A CLOCK polymorphism associated with human diurnal preference. Sleep 1998;21:569-576.

91. Mishima K, Tozawa T, Satoh K, Saitoh H, Mishima Y. The 3111T/C polymorphism of hClock is associated with evening preference and delayed sleep timing in a Japanese population sample. Am J Med Genet B Neuropsychiatr Genet 2005;133:101-104.

92. Robilliard DL, Archer SN, Arendt J, et al. The 3111 Clock gene polymorphism is not associated with sleep and circadian rhythmicity in phenotypically characterized human subjects. J Sleep Res 2002;11:305-312.

93. Lee HJ, Paik JW, Kang SG, Lim SW, Kim L. Allelic variants interaction of CLOCK gene and G-protein beta3 subunit gene with diurnal preference. Chronobiol Int 2007;24:589597.

94. Iwase T, Kajimura N, Uchiyama M, et al. Mutation screening of the human Clock gene in circadian rhythm sleep disorders. Psychiatry Res 2002;109:121-128.

95. Takao T, Tachikawa H, Kawanishi Y, Mizukami K, Asada T. CLOCK gene T3111C polymorphism is associated with Japanese schizophrenics: A preliminary study. Eur Neuropsychopharmacol 2007;17:273-276.

96. Yujnovsky I, Hirayama J, Doi M, Borrelli E, Sassone-Corsi P. Signaling mediated by the dopamine D2 receptor potentiates circadian regulation by CLOCK:BMAL1. Proc Natl Acad Sci USA 2006;103:6386-6391.

97. Aston C, Jiang L, Sokolov BP. Microarray analysis of postmortem temporal cortex from patients with schizophrenia.

J Neurosci Res 2004;77:858-866.

98. Mansour HA, Wood J, Logue T, et al. Association study of eight circadian genes with bipolar I disorder, schizoaffective disorder and schizophrenia. Genes Brain Behav 2006;5:150-157.

99. Peng ZW, Chen XG, Wei Z. Cryptochrome1 maybe a candidate gene of schizophrenia. Med Hypotheses 2007;69: 849-851.

100. Gustincich S, Contini M, Gariboldi M, et al. Gene discovery in genetically labeled single dopaminergic neurons of the retina. Proc Natl Acad Sci USA 2004;101:5069-5074.

101. Uz T, Ahmed R, Akhisaroglu M, et al. Effect of fluoxetine and cocaine on the expression of clock genes in the mouse hippocampus and striatum. Neuroscience 2005;134:13091316.

102. Nievergelt CM, Kripke DF, Barrett TB, et al. Suggestive evidence for association of the circadian genes PERIOD3 and ARNTL with bipolar disorder. Am J Med Genet B Neuropsychiatr Genet 2006;141:234-241.

103. Shiino Y, Nakajima S, Ozeki Y, Isono T, Yamada N. Mutation screening of the human period 2 gene in bipolar disorder. Neurosci Lett 2003;338:82-84.

104. Nievergelt CM, Kripke DF, Remick RA, et al. Examination of the clock gene Cryptochrome 1 in bipolar disorder: mutational analysis and absence of evidence for linkage or association. Psychiatr Genet 2005;15:45-52.

105. Yang S, Van Dongen HP, Wang K, Berrettini W, Bucan M. Assessment of circadian function in fibroblasts of patients with bipolar disorder. Mol Psychiatry 2009;14:143-155.

106. Benedetti F, Dallaspezia S, Fulgosi MC, et al. Actimetric evidence that CLOCK 3111 T/C SNP influences sleep and activity patterns in patients affected by bipolar depression.

Am J Med Genet B Neuropsychiatr Genet 2007;144B:631635.

107. Benedetti F, Serretti A, Colombo C, et al. Influence of CLOCK gene polymorphism on circadian mood fluctuation and illness recurrence in bipolar depression. Am J Med Genet B Neuropsychiatr Genet 2003;123:23-26.

108. Serretti A, Benedetti F, Mandelli L, et al. Genetic dissection of psychopathological symptoms: insomnia in mood disorders and CLOCK gene polymorphism. Am J Med Genet B Neuropsychiatr Genet 2003;121:35-38.

109. Serretti A, Cusin C, Benedetti F, et al. Insomnia improvement during antidepressant treatment and CLOCK gene polymorphism. Am J Med Genet B Neuropsychiatr Genet 2005;137:36-39.

110. Benedetti F, Radaelli D, Bernasconi A, et al. Clock genes beyond the clock: CLOCK genotype biases neural correlates of moral valence decision in depressed patients.

Genes Brain Behav 2008;7:20-25.

111. Roybal K, Theobold D, Graham A, et al. Mania-like behavior induced by disruption of CLOCK. Proc Natl Acad Sci USA 2007;104:6406-6411.

112. Gould TD, Manji HK. Glycogen synthase kinase-3: a putative molecular target for lithium mimetic drugs.

Neuropsychopharmacology 2005;30:1223-1237.

113. Quiroz JA, Gould TD, Manji HK. Molecular effects of lithium. Mol Interv 2004;4:259-272.

114. Kaladchibachi SA, Doble B, Anthopoulos N, Woodgett JR, Manoukian AS. Glycogen synthase kinase 3, circadian rhythms, and bipolar disorder: a molecular link in the therapeutic action of lithium. J Circadian Rhythms 2007;5:3.

115. Yin L, Wang J, Klein PS, Lazar MA. Nuclear receptor Reverbalpha is a critical lithium-sensitive component of the circadian clock. Science 2006;311:1002-1005.

116. Wirz-Justice A, Benedetti F, Berger M, et al.

Chronotherapeutics (light and wake therapy) in affective disorders. Psychol Med 2005;35:939-944.

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Isr J Psychiatry Relat Sci - Vol 47 - No. 1 (2010)

Intrauterine factors as determinants of depressive disorder Marta Weinstock, PhD Department of Pharmacology, The Hebrew University Hadassah Medical Center, Jerusalem, Israel

ABSTRACT Although the etiology of major depressive disorder (MDD) is unknown, it is precipitated in susceptible individuals by adverse events. This review examines the role of intrauterine factors resulting from exposure to stress hormones in the increased vulnerability of the organism to MDD. Severe maternal stress or alcohol intake during the second and third trimesters causes excess release of corticotropin releasing hormone (CRH) and cortisol. These hormones reduce birth weight; impair the feedback regulation of the hypothalamic pituitary adrenal axis (HPA) axis and 5-HT1A and 5-HT2A signaling in key brain areas. Similar changes are seen in patients with MDD and in experimental animals after chronic inescapable stress, prenatal stress or alcohol, which also induce depressive-like behavior in rats, alterations in sleep and circadian rhythms reminiscent of those in humans with MDD. Clinical improvement of MDD by antidepressants is accompanied by normalization of the regulation of the HPA axis and of serotoninergic transmission.

Introduction Major depressive disorder (MDD) remains one of the most frequently seen psychiatric illnesses that still presents a treatment challenge since many patients do not respond adequately to existing therapies (1). MDD is believed to result from a combination of genetic and environmental interactions. A depressive episode may be precipitated in vulnerable individuals by a major stressful life event or

exposure to prolonged periods of distress (2). The ability to cope under conditions of adversity varies considerably among individuals. Failure to do so may lead to impaired regulation of the hypothalamic pituitary adrenal (HPA) axis, prolongation of cortisol release and concomitant changes in serotoninergic and noradrenergic mediated neurotransmission (3). The susceptibility to develop depressive illness in the face of adverse situations is greater in subjects with a family history of depression and in those who had been subjected to sexual or physical abuse during early childhood (4, 5). Depressive illness is also more likely to occur in subjects whose mothers suffered from depression during pregnancy (6) or were exposed to uncontrollable stress (7), infection (8) or alcohol (9).

The developing fetal brain is particularly sensitive to hormones, cytokines and noxious substances reaching it from the maternal circulation that may permanently alter its structure and function. Stress hormones include adrenaline, cortisol (corticosterone [COR] in rodents) and corticotrophin releasing hormone (CRH). This review will focus on the changes in brain morphology, regulation of the HPA axis and brain serotoninergic system associated with depressive-like behavior in the offspring resulting from maternal stress hormones, alcohol abuse and infection and that could lead to a greater vulnerability to develop depressive illness.

Association Between Prenatal Stress and Depression in Human Subjects Although many retrospective studies have linked exposure to adverse events during pregnancy to a higher incidence of schizophrenia (reviewed in 10), there have been only a few reports linking intrauterine factors to depressive illness. One of these reported an incidence of 30% with

Address for Correspondence: Professor Marta Weinstock, Department of Pharmacology, School of Pharmacy, The Hebrew University Medical Center, Ein Kerem, Jerusalem 91120, Israel. â&#x20AC;&#x2020;martar@ekmd.huji.ac.il

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Marta Weinstock

severe depression in the adult offspring of women who were subjected to the severe “hunger winter” in Holland during World War II. The frequency of depression was greatest when stress exposure occurred during the second and third trimesters, and its prevalence was higher in men than in women (11). No differentiation was made between nutritional deficiency and the degree of stress in the etiology of the depression in this study. However, an incidence of severe depression of 13.3% was reported in the 18-yearold offspring of women pregnant during a severe earthquake (7.8 on the Richter scale) compared to 5.5% in those born a year later. More men than women were found with depression, particularly when the stress exposure occurred during the second trimester of pregnancy (7).

Chronic psychological stress during pregnancy has been shown to decrease the length of gestation and birth weight (12, 13). Antenatal stress, as assessed from selfreported high levels of anxiety or depression, was associated with raised circulating levels of CRH and of cortisol in both the 18-20th and 28-30th weeks of gestation (6, 14). In contrast to the inhibition by cortisol of CRH release from the hypothalamus, stress levels of cortisol stimulate CRH release from the placenta (15). Plasma CRH and cortisol were inversely related to birth weight (reviewed in 13). Therefore, other studies have used low birth weight (LBW) as an indicator of maternal stress.

LBW infants have a higher prevalence of emotional problems, anxiety and learning difficulties than those of normal birth weight (reviewed in 16, 17). However, an association between LBW and depression is found in some (18-20) but not other studies (21, 22). The reasons for these discrepancies are not clear but may depend on the age at which the subjects are assessed and whether or not other factors, like a family history of depression or childhood hardship, contribute to the outcome.

Low birth weight may be considered as an indicator of poor intrauterine conditions for growth and development that provoke physiological adjustments that have long term consequences for health and function (23). While such adjustments increase the chances of fetal survival, they could render the individual less able to cope with stressful conditions during later life. A significant association was found between LBW and hypertension (24, 25), and was recently tested in relation to the incidence of childhood depression (18). In the absence of other adverse conditions during childhood, such as violence between parents or physical abuse, the rate of depression in LBW teenage boys and girls did not differ from those of normal birth weight. However, exposure of LBW girls

but not boys to one or two such hardships during childhood resulted in depressive symptoms around puberty in 20% and 60% respectively, compared to 4% and 20% of controls. It is possible that the greater prevalence of depression in girls than in boys in these studies, in contrast to those in adults described above, is due to the younger age at which the assessments were made since depression has an earlier onset in females (26). These data allow the inference that an adverse maternal milieu in girls can interact with hormonal changes associated with puberty and stress and predispose the individual to depressive and other mental health illness in later life. It should be emphasized that in none of these studies was any attempt made to analyze a possible contribution to the outcome of a family history of affective disorder.

Association Between Prenatal Infection and Depression in Human Subjects Epidemiological studies show that maternal exposure to infection increases the release of pro inflammatory cytokines, tumor necrosis factor alpha (TNFα) and interleukin-1beta (IL-1β) from macrophages into the circulation (27). Excess levels of these cytokines could induce premature birth (28), itself a possible risk factor for depression (see preceding section). Exposure of pregnant women to the A2/Singapore influenza epidemic resulted in a significantly higher incidence of major depression in their offspring than in those born six years earlier. Again, the prevalence of depression was greater in men than in women and when the mothers were exposed to the disease during the second trimester of pregnancy (8). Severe stress during pregnancy also increased circulating levels of IL-6 and TNFα (29) and reduced those of progesterone and IL-10 that are involved in the maintenance of pregnancy. It therefore appears that the effect of gestational stress on the developing fetus involves several interacting factors including alterations in cytokines and stress hormones.

Association Between Maternal Alcohol Intake and Depression in Human Subjects Ethyl alcohol can have both direct and indirect effects on the developing fetus. It readily crosses the placenta and can affect the integrity of fetal neurones but can also influence maternal endocrine function resulting in excess release of stress hormones which can also adversely affect the developing fetal brain. Abnormalities in development 37

INTRAUTERINE FACTORS AS DETERMINANTS OF DEPRESSIVE DISORDER

and behavior in children exposed prenatally to alcohol are well documented and some of them resemble those seen after prenatal stress. They include deficits in learning, memory and executive functioning, hyperactivity, impulsivity, aggression and delinquency and poor communication and social skills (30, 31). An incidence of depressive symptoms of 27% was also found in children exposed to high levels of prenatal alcohol (cited by 9).

This may be an important predictor of depression in adulthood since about 44% of adult offspring of mothers drinking large amounts of alcohol during pregnancy were reported to be seriously depressed (32). The interpretation of the findings is complicated since alcohol intake is significantly associated with maternal depression (33). A study made to assess the contribution of maternal depression independently of alcohol intake found that each of these independently increased the likelihood of childhood depression (9). However, the highest incidence of depression occurred, particularly in girls, when maternal depression was accompanied by moderate to heavy alcohol intake (3-5 or more drinks per occasion). The weakness of this study is that it failed to include data on paternal drinking in spite of the fact that paternal alcoholism has been shown to be a major predictor of depression of early onset in the offspring (34). Moreover, women with alcoholic partners are twice as likely as those with non-alcoholic partners to abuse alcohol (35). Neither was consideration given to the probability that living with an alcoholic husband increases the likelihood of psychological stress in the mother with the attendant effects on the developing fetus and later on the child.

In summary, severe maternal stress in humans caused by uncontrollable factors such as prolonged famine, a major earthquake or maternal infection can increase the incidence of depression in adult offspring. The highest incidence is associated with stress occurring in the second trimester of pregnancy when circulating levels of cortical and CRH are elevated and appears to be more prevalent in males. On the other hand, LBW, prenatal maternal anxiety, depression and/or excessive alcohol intake are more likely to induce depressive symptoms in adolescent girls than in boys if this is accompanied by additional hardship during childhood.

Maternal Stress Hormones and Programming of the Fetal Brain Acute stress, whether environmental or psychological, activates the HPA axis and sympathetic nervous system 38

and causes a transient increase in plasma glucocorticoids and catecholamines. The response of the HPA axis to chronic stress depends on whether the organism has developed coping strategies and adapts to the stress. In pregnant women who do not adapt to the adverse circumstances and report high levels of stress, circulating CRH, ACTH and β-endorphin and cortisol are elevated in the second and third trimester of gestation (6, 13, 36).

The fetus is normally protected from excess levels of glucocorticoids by placental hydroxysteroid dehydrogenase (11β-HSD) that converts cortisol to inactive cortisone, and by COR binding globulin (CBG) which sequesters any cortisol released into the circulation. However, in rats both prenatal stress (37) and chronic maternal malnutrition (38) reduce the activity of 11β-HSD, while prenatal stress also decreases the levels of CBG in maternal plasma (39), thereby potentially increasing the levels of circulating free COR. The blood levels of glucocorticoids are also controlled by negative feedback on the HPA axis via glucocorticoid receptors (GR) in the pituitary, hippocampus, hypothalamic CRH neurons and prefrontal cortex (PFC) (40). In the hippocampus, these GR and mineralocorticoid receptors (MR) become desensitized by chronic stress (40). Together, a decrease in MR and GR signaling, a fall in placental 11β-HSD activity and in circulating CBG levels increase the exposure of the fetal brain to glucocorticoids. If these reach the fetal brain in sufficient concentrations at a critical time during development they could alter its structure and function thereby sensitizing it to the effects of subsequent stress exposure.

Effect of Prenatal Stress, Alcohol or Infection on the Regulation of the Offspring HPA Axis Although an association has been found between maternal stress, impaired feedback regulation of the HPA axis and MDD in adult humans (41, 42) it is not known whether this results from altered intrauterine, genetic or postnatal factors, or a combination of them. Some depressed subjects, but not others, show a deficit in dexamethasone-CRH suppression indicating a decrease in GR activation (43). More direct support for a role of intrauterine factors in the alteration of the HPA axis and in the etiology of MDD have come from studies in experimental animals (for detailed reviews see 17, 44). Chronic exposure of the fetal rat brain to COR in the course of prenatal stress does not usually alter resting morning (low) levels of the steroid in

Marta Weinstock

the adult offspring, but may increase the total output over 24 hrs (13) and reduce hippocampal MR and GR (44, 45). However, on exposure to stress, plasma COR increases more in prenatally-stressed (PS) rats (44, 46, 47) and monkeys (48, 49) than in controls. The duration of COR elevation is also longer in PS rats (17), indicating impaired regulation of the HPA axis.

Similar alterations in the regulation of the HPA axis are seen in the offspring of rodents or humans exposed prenatally to alcohol (50, 51) or infection (52, 53). Maternal adrenalectomy prevents the changes in the offspring HPA axis induced by stress (54) or alcohol intake (55). Injection of COR to mimic the plasma levels induced by stress reinstates the changes in the offspring HPA axis, thereby confirming its role as a mediator of the altered programming. In general, the HPA axis in the female offspring of rats exposed to stress, alcohol or cytokines is more sensitive than that of males (45, 53, 56). There do not appear to be any reports of studies on the effects of prenatal infection on the regulation of the HPA axis in humans.

Depressive-like Behavior Induced in Rats by Prenatal Stress While depression cannot be diagnosed and assessed in experimental animals, one can discern some of its distinctive features and the appropriate physiological changes in PS rats. These include a phase shift in circadian rhythms of plasma cortisol and body temperature (57) and a disturbance in the normal sleep pattern (58).

These phenomena can be reproduced in rats by maternal stress (59, 60). Moreover, the changes in rapid-eye movement and slow wave sleep in PS rats were found to be correlated with the magnitude of the increase in plasma COR in response to restraint stress (59). Prenatal stress in some rat strains (61-64) but not others (65) induces a form of learned helplessness in the forced swim test more readily than in controls. This behavior is accompanied by an increased response of the HPA axis to stress like in depressed human subjects. It therefore appears that intrauterine and genetic factors play a role in rats in the ultimate effects of brain programming by maternal stress.

Depressive-like Behavior Induced in Rats by Alcohol or Maternal Infection Daily administration of alcohol to pregnant rats also induces depressive-like behavior (66) and enhances the response of the HPA axis to stress particularly in

the female offspring (67-69). On the other hand, there do not appear to be any studies on the behavior of rats subjected in utero to infection. Injection of lipopolysaccharide into pregnant mice on day 17 of gestation to mimic bacterial infection results in anxiety and reduces social interaction in the offspring (70) like that seen in PS mice (71), but its effect on depressive-like behavior has not been investigated.

Depression, HPA Axis Regulation and the Brain Serotonin System The brain serotonin (5-HT) system has been strongly implicated in anxiety and depression (72), and drugs that prolong the action of 5-HT by inhibiting its inactivation by monoamine oxidase or its neuronal uptake are effective antidepressants (73). Although there are many types of 5-HT receptors particular attention has been focused on the 5-HT1A and 5-HT2A/C subtypes in relation to affective disorders (72). In humans, 5-HT1A receptors (5-HT1AR) are expressed presynaptically in 5-HT cell bodies in the raphĂŠ nuclei. They are also found postsynaptically on pyramidal cells in the hippocampus, hypothalamus and frontal cortex (74) where their activation inhibits glutamate-mediated depolarization (75). Hippocampal 5-HT1AR are believed to maintain adaptive behaviors in the face of aversive stimuli, and a decrease in their activation can lead to learned helplessness in rats and depression in humans. 5-HT is also involved in the regulation of the HPA axis. Stimulation of 5-HT2AR on CRH neurons in the hypothalamus (76), and of 5-HT1AR and 5-HT2AR in the anterior pituitary (77) increases the release of ACTH. a. Chronic stress in rats and 5-HT receptors

Adrenal steroids and chronic stress inhibit the expression of postsynaptic 5-HT1AR in the hippocampus and other brain regions of rats (78, 79). Adrenal steroids produce their effect on 5-HT1AR mainly via MR but GR also play a role (80). Chronic treatment of repeatedly-stressed adult rats with different types of antidepressants prevents the down regulation of postsynaptic hippocampal 5-HT1AR and restores MR and GR density to pre-stress levels (78, 81). In contrast to their effect on 5-HT1AR, glucocorticoids and chronic stress in rats increase the expression of 5-HT2AR in the FC and hippocampus (81-83). b. 5-HT receptors in humans with MDD

Like chronically stressed rats, subjects with MDD showed a significant decrease in 5-HT1AR binding in 39

INTRAUTERINE FACTORS AS DETERMINANTS OF DEPRESSIVE DISORDER

the frontal, temporal and limbic cortices, the hippocampus-amygdala region and in raphĂŠ nuclei, as measured by neuroimaging with PET using [carbonyl-11C] WAY100635 (84). However, unlike the finding in chronicallystressed rats, this was not reversed by treatment with antidepressants (85, 86). An increase in 5-HT2AR was found in the pre-FC of young suicide victims presumed to have suffered from MDD (87). c. Prenatal stress and 5-HT receptors

So far it appears that only two studies have examined the effect of prenatal stress on 5-HT receptor immunoreactivity or expression in rats. As in humans with depression, a decrease was found in PS males in 5-HT1AR binding in the ventral hippocampus, an area in the rat primarily linked to emotional processing (88).

Surprisingly, others reported an increase in mRNA of 5-HT1ARs in the prefrontal cortex (PFC) (63), and a reduction in hippocampal MR and GRs together with depressive-like behavior. Although GRs are present in the PFC it is not clear why their down-regulation should be accompanied by an increase in 5-HT1AR in this region. Chronic treatment with different classes of antidepressants prevented the development of learned helplessness in PS rats, restored GR receptors and normalized the regulation of the HPA axis (17, 64). d. Effect of prenatal alcohol administration in rats on 5-HT receptors

Prenatal alcohol exposure in rats has been shown to cause depressive-like behavior in the offspring (66), but 5-HTRs were not measured in this study. Others found that the increased activity of the HPA axis in rats exposed prenatally to alcohol was associated with an alteration in the balance between 5-HT1AR and 5-HT2AR at different levels of the HPA axis in females but not in males (89). Thus, female rats exposed prenatally to alcohol showed a blunted release of ACTH in response to a 5-HT1A agonist, consistent with a reduction in this receptor signaling, together with an enhanced response to a 5-HT2A agonist. Prenatal alcohol exposure also increased the 5-HT transporter in cortical layers 5 and 6, hippocampal CA layers 2 and 3, lateral nucleus of the amygdala and in the dorsal raphĂŠ nucleus, thereby possibly reducing synaptic levels of 5-HT (90). There do not seem to be any reports on the effects of antidepressant treatment on depressive-like behavior, HPA axis regulation or 5-HT receptors in rats exposed prenatally to alcohol. The disparate findings in the effects of prena40

tal stress or alcohol on 5-HT receptors in rats that have evidence of dysregulation of the HPA axis may be due to the methods of assessment of 5-HT receptor activity, the different brain regions examined and the time of examination in relation to the measures of depressive behavior and HPA axis reactivity.

Changes in Brain Structure in Human Subjects with Depression Post mortem studies in patients with MDD indicate a reduction in neuronal size in the orbito-frontal cortex (OFC) and dorsolateral prefrontal cortex (DLPFC) (91).

Magnetic resonance imaging also detected a decrease in blood flow in the DLPFC that may produce psychomotor retardation and apathy (69). An increase in blood flow and cerebral glucose metabolism (CMG) was found in the ventromedial and lateral OFC which may explain the enhanced sensitivity to pain, anxiety and depressive thoughts in such patients (92). Successful treatment with antidepressants normalized CGM in the ventromedial and lateral OFC of MDD subjects (92).

The amygdala is bidirectionally connected to the PFC and hippocampus and plays a crucial role in the regulation of mood and affect (93, 94). CGM is increased in the left amygdala of untreated subjects with MDD. In those showing a persistent positive treatment response to antidepressants there is also a reduction in the level of CGM towards the level in control subjects (84).

Several imaging studies, but not others (reviewed in 91, 95) found a significant reduction in left and right hippocampal volume in patients with MDD compared with controls (96, 97). The degree of hippocampal reduction appears to be directly proportional to the number and the duration of untreated depressive episodes suggesting they may be the result rather than the cause of the depression (98). In patients treated with antidepressants (no details of which are given) the hippocampal volume loss no longer increased with time, suggesting that the drugs may have had some beneficial effect. One can really only determine whether antidepressants can restore hippocampal structure if measurements are made before and after treatment in the same subjects. Nevertheless, other data suggest that a reduction in hippocampal function and volume is more likely to occur in subjects with a significant history of prepubertal physical or sexual abuse than in those with no abuse (99, 100). It is not known whether hippocampal volume is also reduced in human subjects in whom

Marta Weinstock

depression is associated with prenatal stress or infection. However, hippocampal volume was smaller in PS Rhesus monkeys that showed an abnormal dexamethasone suppression test, indicating alterations in the control of the HPA axis (101). It is also not clear from studies in humans with MDD whether antidepressant therapy alters the structural and functional changes in different brain regions associated with this condition.

However, chronic treatment with paroxetine of young subjects with obsessive compulsive disorder significantly reduced the enlarged volume of the left amygdala that is also observed in MDD. The extent of the reduction in amygdala volume was correlated with the total dose of paroxetine (102).

of dendritic spines and spine length. In one study in which only males were examined PS rats showed a 32% decrease in the density of synapses in the CA3 region of the hippocampus (106). Another study found that young female but not male PS rats showed a significant reduction in the total number of hippocampal neurons (107). However, no assessments were made of their behavior or of the reaction of the HPA axis to stress.

In PS male mice which showed an increase in the HPA axis response to stress, a 19-22% reduction was found in the density of synapses and number of dendritic spines in hippocampal CA3 pyramidal cells. These changes were reversed by administration of the antidepressant fluoxetine from the age of 1-3 weeks (108).

Changes in Brain Structure in Human Subjects Exposed Prenatally to Alcohol Structural abnormalities in the cerebellum, basal ganglia and corpus callosum have been reported in humans exposed prenatally to high levels of alcohol (103). Brain imaging and analytic techniques in such subjects have indicated specific alterations including displacements in the corpus callosum, increased gray matter density in the perisylvian regions, altered gray matter asymmetry, and disproportionate reductions in the frontal lobes (31). Apart from the greater changes in the PFC there was no clear resemblance between the structural changes induced by prenatal alcohol and those in subjects with MDD. The lack of such a similarity could be due to a direct toxic effect of alcohol on the developing brain which may mask smaller changes associated with deficits in the regulation of the HPA axis.

Conclusions Data from retrospective and prospective studies on the etiology of depression in humans support a role of stress hormones, cytokines and alcohol in the intrauterine environment. Stress hormones like cortisol that reach the fetal brain during a critical time of development alter its programming and sensitize the organism to the effects of stress in childhood and early adulthood.

Such sensitization is manifested by alterations in the feedback regulation of the HPA axis to stress via MR and GR, and in the actions of 5-HT via presynaptic and postynaptic 5-HT1AR and postsynaptic 5-HT2AR, all of which can increase the likelihood to develop depression. Successful treatment by antidepressants is associated with restoration of the regulation of the HPA axis and possibly also of 5-HT activity.

Structural Changes in the Brain of Animals Subjected to Prenatal Stress The anterior cingulate (AC) and OFC are known to be implicated in the regulation of emotional behavior (104). Prenatal stress on days 17-21 of gestation produced a significant reduction in frequencies of dendritic spines on layer II/III pyramidal neurons of the AC and OFC in both males and females aged 23 days.

PS males, but not females, also showed a decrease in the length and complexity of pyramidal apical dendrites in both cortical regions (105). This agrees with the finding in depressed patients of a decrease in size of the OFC (91) which may also have been partly due to a loss

References 1. Nemeroff CB. The burden of severe depression: A review of diagnostic challenges and treatment alternatives. J Psychiatr Res 2007;41:189-206.

2. Kendler KS, Karkowski-Shuman L. Stressful life events and genetic liability to major depression: Genetic control of exposure to the environment? Psychol Med 1997;27:539-547.

3. Weinstock M. Convergence of antidementia and antidepressant pharmacology. In: Sun M-K, editor. Cognition and mood interactions. New York: Nova Biomedical Books, 2005: pp. 185-225.

4. Bifulco A, Brown GW, Adler Z. Early sexual abuse and clinical depression in adult life. Br J Psychiatry 1991;159:115-122.

5. Phillips NK, Hammen CL, Brennan PA, et al. Early adversity and the prospective prediction of depressive and 41

INTRAUTERINE FACTORS AS DETERMINANTS OF DEPRESSIVE DISORDER

anxiety disorders in adolescents. J Abnorm Child Psychol 2005;33:13-24.

6. Field T, Diego M. Cortisol: The culprit prenatal stress variable. Int J Neurosci 2008;118:1181.

7. Watson JB, Mednick SA, Huttunen M, Wang X. Prenatal teratogens and the development of adult mental illness.

Dev Psychopathol 1999;11:457-466.

8. Machon RA, Mednick SA, Huttunen MO. Adult major affective disorder after prenatal exposure to an influenza epidemic. Arch Gen Psychiatry 1997;54:322-328.

9. O'Connor MJ, Kasari C. Prenatal alcohol exposure and depressive features in children. Alcohol Clin Exp Res 2000;24:1084-1092.

10. Weinstock M. Alterations induced by gestational stress in brain morphology and behaviour of the offspring. Prog Neurobiol 2001;65:427-451.

11. Brown AS, van Os J, Driessens C, et al. Further evidence of relation between prenatal famine and major affective disorder. Am J Psychiatry 2000;157:190-195.

12. Hedegaard M, Henriksen TB, Secher NJ, et al. Do stressful life events affect duration of gestation and risk of preterm delivery? Epidemiology 1996;7:339-345.

13. Weinstock M. The potential influence of maternal stress hormones on development and mental health of the offspring. Brain Behav Immun 2005;19:296-308.

14. Mancuso RA, Schetter CD, Rini CM, et al. Maternal prenatal anxiety and corticotropin-releasing hormone associated with timing of delivery. Psychosom Med 2004;66:762-769.

21. Inskip HM, Dunn N, Godfrey KM, et al. Is birth weight associated with risk of depressive symptoms in young women? Evidence from the Southampton Women's Survey. Am J Epidemiol 2008;167:164-168.

22. Vasiliadis HM, Gilman SE, Buka SL. Fetal growth restriction and the development of major depression. Acta Psychiatr Scand 2008;117:306-312.

23. Bateson P, Barker D, Clutton-Brock T, et al. Developmental plasticity and human health. Nature 2004;430:419-421.

24. Barker DJ. The developmental origins of adult disease.

Eur J Epidemiol 2003;18:733-736.

25. Bellingham-Young DA, Adamson-Macedo EN. Foetal origins theory: Links with adult depression and general self-efficacy. Neuro Endocrinol Lett 2003;24:412-416.

26. Kornstein SG, Schatzberg AF, Thase ME, et al. Gender differences in chronic major and double depression. J Affect Disord 2000;60:1-11.

27. Besedovsky HO, del Rey A. Immune-neuroendocrine circuits: Integrative role of cytokines. Front Neuroendocrinol 1992;13:61-94.

28. Gomez R, Ghezzi F, Romero R, et al. Premature labor and intra-amniotic infection. Clinical aspects and role of the cytokines in diagnosis and pathophysiology. Clin Perinatol 1995;22:281-342.

29. Coussons-Read ME, Okun ML, Schmitt MP, Giese S. Prenatal stress alters cytokine levels in a manner that may endanger human pregnancy. Psychosom Med 2005;67:625-631.

30. Mattson SN, Riley EP. A review of the neurobehavioral deficits in children with fetal alcohol syndrome or prenatal exposure to alcohol. Alcohol Clin Exp Res 1998;22:279-294.

15. Sandman CA, Glynn L, Schetter CD, et al. Elevated maternal cortisol early in pregnancy predicts third trimester levels of placental corticotropin releasing hormone (CRH): Priming the placental clock. Peptides 2006;27:1457-1463.

31. Niccols A. Fetal alcohol syndrome and the developing socio-emotional brain. Brain Cogn 2007;65:135-142.

16. Rice F, Jones I, Thapar A. The impact of gestational stress and prenatal growth on emotional problems in offspring: A review. Acta Psychiatr Scand 2007;115:171-183.

32. Famy C, Streissguth AP, Unis AS. Mental illness in adults with fetal alcohol syndrome or fetal alcohol effects. Am J Psychiatry 1998;155:552-554.

17. Weinstock M. The long-term behavioural consequences of prenatal stress. Neurosci Biobehav Rev 2008;32:1073-1083.

33. Zuckerman B, Amaro H, Bauchner H, Cabral H. Depressive symptoms during pregnancy: Relationship to poor health behaviors. Am J Obstet Gynecol 1989;160:1107-1111.

18. Costello EJ, Worthman C, Erkanli A, Angold A. Prediction from low birth weight to female adolescent depression: A test of competing hypotheses. Arch Gen Psychiatry 2007;64:338-344.

34. Todd RD, Geller B, Neuman R, et al. Increased prevalence of alcoholism in relatives of depressed and bipolar children.

J Am Acad Child Adolesc Psychiatry 1996;35:716-724.

19. Gale CR, Martyn CN. Birth weight and later risk of depression in a national birth cohort. Br J Psychiatry 2004;184:28-33.

35. Windle M, Windle RC, Scheidt DM, Miller GB. Physical and sexual abuse and associated mental disorders among alcoholic inpatients. Am J Psychiatry 1995;152:1322-1328.

20. Thompson C, Syddall H, Rodin I, et al. Birth weight and the risk of depressive disorder in late life. Br J Psychiatry 2001;179:450-455.

36. Wadhwa PD, Dunkel-Schetter C, Chicz-DeMet A, et al.

Prenatal psychosocial factors and the neuroendocrine axis in human pregnancy. Psychosom Med 1996;58:432-446.

42

Marta Weinstock

37. Mairesse J, Lesage J, Breton C, et al. Maternal stress alters endocrine function of the feto-placental unit in rats. Am J Physiol Endocrinol Metab 2007;292:E1526-1533.

38. Whorwood CB, Firth KM, Budge H, Symonds ME. Maternal undernutrition during early to midgestation programs tissue-specific alterations in the expression of the glucocorticoid receptor, 11beta-hydroxysteroid dehydrogenase isoforms, and type 1 angiotensin ii receptor in neonatal sheep. Endocrinology 2001;142:2854-2864.

39. Takahashi LK, Turner JG, Kalin NH. Prolonged stressinduced elevation in plasma corticosterone during pregnancy in the rat: Implications for prenatal stress studies.

Psychoneuroendocrinology 1998;23:571-581.

Psychoneuroendocrinology 2002;27:285-298.

50. Jacobson SW, Bihun JT, Chiodo LM. Effects of prenatal alcohol and cocaine exposure on infant cortisol levels.

Dev Psychopathol 1999;11:195-208.

51. Lee S, Imaki T, Vale W, Rivier C. Effect of prenatal exposure to ethanol on the activity of the hypothalamic-pituitary-adrenal axis of the offspring: Importance of the time of exposure to ethanol and possible modulating mechanisms. Mol Cell Neurosci 1990;1:168-177.

52. Reul JM, Stec I, Wiegers GJ, et al. Prenatal immune challenge alters the hypothalamic-pituitary-adrenocortical axis in adult rats. J Clin Invest 1994;93:2600-2607.

40. De Kloet ER, Vreugdenhil E, Oitzl MS, Joels M. Brain corticosteroid receptor balance in health and disease.

Endocr Rev 1998;19:269-301.

53. Samuelsson AM, Ohrn I, Dahlgren J, et al. Prenatal exposure to interleukin-6 results in hypertension and increased hypothalamic-pituitary-adrenal axis activity in adult rats. Endocrinology 2004;145:4897-4911.

41. Heim C, Newport DJ, Heit S, et al. Pituitary-adrenal and autonomic responses to stress in women after sexual and physical abuse in childhood. Jama 2000;284:592-597.

54. Barbazanges A, Piazza PV, Le Moal M, Maccari S. Maternal glucocorticoid secretion mediates long-term effects of prenatal stress. J Neurosci 1996;16:3943-3949.

42. Modell S, Yassouridis A, Huber J, Holsboer F. Corticosteroid receptor function is decreased in depressed patients. Neuroendocrinology 1997;65:216-222.

55. Slone JL, Redei EE. Maternal alcohol and adrenalectomy: Asynchrony of stress response and forced swim behavior.

Neurotoxicol Teratol 2002;24:173-178.

43. Kunzel HE, Binder EB, Nickel T, et al. Pharmacological and nonpharmacological factors influencing hypothalamic-pituitary-adrenocortical axis reactivity in acutely depressed psychiatric in-patients, measured by the Dex-CRH test.

Neuropsychopharmacology 2003;28:2169-2178.

56. McCormick CM, Smythe JW, Sharma S, Meaney MJ. Sexspecific effects of prenatal stress on hypothalamic-pituitary-adrenal responses to stress and brain glucocorticoid receptor density in adult rats. Brain Res Dev Brain Res 1995;84:55-61.

44. Maccari S, Darnaudery M, Morley-Fletcher S, et al.

Prenatal stress and long-term consequences: Implications of glucocorticoid hormones. Neurosci Biobehav Rev 2003;27:119-127.

57. Goodwin FK, Wirz-Justice A, Wehr TA. Evidence that the pathophysiology of depression and the mechanism of action of antidepressant drugs both involve alterations in circadian rhythms. Adv Biochem Psychopharmacol 1982;32:1-11.

45. Weinstock M, Matlina E, Maor GI, et al. Prenatal stress selectively alters the reactivity of the hypothalamic-pituitary adrenal system in the female rat. Brain Res 1992;595:195-200.

46. Koenig JI, Elmer GI, Shepard PD, et al. Prenatal exposure to a repeated variable stress paradigm elicits behavioral and neuroendocrinological changes in the adult offspring: potential relevance to schizophrenia. Behav Brain Res 2005;156:251-261.

47. Weinstock M, Poltyrev T, Schorer-Apelbaum D, et al.

Effect of prenatal stress on plasma corticosterone and catecholamines in response to footshock in rats. Physiol Behav 1998;64:439-444.

48. Clarke AS, Wittwer DJ, Abbott DH, Schneider ML. Longterm effects of prenatal stress on HPA axis activity in juvenile rhesus monkeys. Dev Psychobiol 1994;27:257-269.

49. Schneider ML, Moore CF, Kraemer GW, et al. The impact of prenatal stress, fetal alcohol exposure, or both on development: perspectives from a primate model.

58. Kupfer DJ. Sleep research in depressive illness: Clinical implications â&#x20AC;&#x201C; a tasting menu. Biol Psychiatry 1995;38:391-403.

59. Dugovic C, Maccari S, Weibel L, et al. High corticosterone levels in prenatally stressed rats predict persistent paradoxical sleep alterations. J Neurosci 1999;19:8656-8664.

60. Koehl M, Darnaudery M, Dulluc J, et al. Prenatal stress alters circadian activity of hypothalamo-pituitary-adrenal axis and hippocampal corticosteroid receptors in adult rats of both genders. J Neurobiol 1999;40:302-315.

61. Abe H, Hidaka N, Kawagoe C, et al. Prenatal psychological stress causes higher emotionality, depression-like behavior, and elevated activity in the hypothalamo-pituitary-adrenal axis. Neurosci Res 2007;59:145-151.

62. Morley-Fletcher S, Darnaudery M, Koehl M, et al.

Prenatal stress in rats predicts immobility behavior in the forced swim test. Effects of a chronic treatment with tianeptine. Brain Res 2003;989:246-251.

43

INTRAUTERINE FACTORS AS DETERMINANTS OF DEPRESSIVE DISORDER

63. Morley-Fletcher S, Darnaudery M, Mocaer E, et al.

Chronic treatment with imipramine reverses immobility behaviour, hippocampal corticosteroid receptors and cortical 5-HT(1A) receptor mRNA in prenatally stressed rats. Neuropharmacology 2004;47:841-847.

64. Poltyrev T, Gorodetsky E, Bejar C, et al. Effect of chronic treatment with ladostigil (TV-3326) on anxiogenic and depressive-like behaviour and on activity of the hypothalamicpituitary-adrenal axis in male and female prenatally stressed rats. Psychopharmacology (Berl) 2005;181:118-125.

65. Van den Hove DL, Blanco CE, Aendekerk B, et al. Prenatal restraint stress and long-term affective consequences.

Dev Neurosci 2005;27:313-320.

66. Carneiro LM, Diogenes JP, Vasconcelos SM, et al.

Behavioral and neurochemical effects on rat offspring after prenatal exposure to ethanol. Neurotoxicol Teratol 2005;27:585-592.

67. Weinberg J. Prenatal ethanol exposure alters adrenocortical response to predictable and unpredictable stressors.

Alcohol 1992;9:427-432.

68. Weinberg J. Hyperresponsiveness to stress: Differential effects of prenatal ethanol on males and females. Alcohol Clin Exp Res 1988;12:647-652.

69. Maletic V, Robinson M, Oakes T, et al. Neurobiology of depression: an integrated view of key findings. Int J Clin Pract 2007;61:2030-2040.

70. Golan H, Stilman M, Lev V, Huleihel M. Normal aging of offspring mice of mothers with induced inflammation during pregnancy. Neuroscience 2006;141:1909-1918.

71. Pallares ME, Scacchi Bernasconi PA, Feleder C, Cutrera RA. Effects of prenatal stress on motor performance and anxiety behavior in Swiss mice. Physiol Behav 2007;92:951-956.

72. Graeff FG, Guimaraes FS, De Andrade TG, Deakin JF. Role of 5-HT in stress, anxiety, and depression. Pharmacol Biochem Behav 1996;54:129-141.

73. Maes M, Meltzer H. The serotonin hypothesis of major depression. In: Bloom FE, Kupfer DJ, editors.

Psychophamacology: The fourth generation of progress.

New York: Raven, 1995: pp. 933-944.

74. Chaouloff F. Physiopharmacological interactions between stress hormones and central serotonergic systems. Brain Res Brain Res Rev 1993;18:1-32.

75. Sprouse JS, Aghajanian GK. Responses of hippocampal pyramidal cells to putative serotonin 5-HT1A and 5-HT1B agonists: A comparative study with dorsal raphe neurons. Neuropharmacology 1988;27:707-715.

76. Van de Kar LD, Javed A, Zhang Y, et al. 5-HT2A receptors stimulate ACTH, corticosterone, oxytocin, renin, and prolactin release and activate hypothalamic CRF and 44

oxytocin-expressing cells. J Neurosci 2001;21:3572-3579.

77. Calogero AE, Bagdy G, Szemeredi K, et al. Mechanisms of serotonin receptor agonist-induced activation of the hypothalamic-pituitary-adrenal axis in the rat.

Endocrinology 1990;126:1888-1894.

78. Lopez JF, Chalmers DT, Little KY, Watson SJ. A.E. Bennett Research Award. Regulation of serotonin1A, glucocorticoid, and mineralocorticoid receptor in rat and human hippocampus: implications for the neurobiology of depression. Biol Psychiatry 1998;43:547-573.

79. Meijer OC, De Kloet ER. Corticosterone suppresses the expression of 5-HT1A receptor mRNA in rat dentate gyrus. Eur J Pharmacol 1994;266:255-261.

80. Meijer OC, Van Oosten RV, De Kloet ER. Elevated basal trough levels of corticosterone suppress hippocampal 5-hydroxytryptamine(1A) receptor expression in adrenally intact rats: Implication for the pathogenesis of depression. Neuroscience 1997;80:419-426.

81. Watanabe Y, Sakai RR, McEwen BS, Mendelson S. Stress and antidepressant effects on hippocampal and cortical 5-HT1A and 5-HT2 receptors and transport sites for serotonin. Brain Res 1993;615:87-94.

82. Dwivedi Y, Mondal AC, Payappagoudar GV, Rizavi HS. Differential regulation of serotonin (5HT)2A receptor mRNA and protein levels after single and repeated stress in rat brain: Role in learned helplessness behavior.

Neuropharmacology 2005;48:204-214.

83. Joels M, Van Riel E. Mineralocorticoid and glucocorticoid receptor-mediated effects on serotonergic transmission in health and disease. Ann N Y Acad Sci 2004;1032:301-303.

84. Drevets WC. Prefrontal cortical-amygdalar metabolism in major depression. Ann N Y Acad Sci 1999;877:614-637.

85. Drevets WC. Orbitofrontal cortex function and structure in depression. Ann N Y Acad Sci 2007;1121:499-527.

86. Sargent PA, Kjaer KH, Bench CJ, et al. Brain serotonin1A receptor binding measured by positron emission tomography with [11C]WAY-100635: Effects of depression and antidepressant treatment. Arch Gen Psychiatry 2000;57:174-180.

87. Pandey GN, Dwivedi Y, Rizavi HS, et al. Higher expression of serotonin 5-HT(2A) receptors in the postmortem brains of teenage suicide victims. Am J Psychiatry 2002;159:419-429.

88. Van den Hove DL, Lauder JM, Scheepens A, et al. Prenatal stress in the rat alters 5-HT1A receptor binding in the ventral hippocampus. Brain Res 2006;1090:29-34.

89. Hofmann CE, Ellis L, Yu WK, Weinberg J. Hypothalamicpituitary-adrenal responses to 5-HT1A and 5-HT2A/C agonists are differentially altered in female and male rats prenatally exposed to ethanol. Alcohol Clin Exp Res 2007;31:345-355.

Marta Weinstock

90. Zafar H, Shelat SG, Redei E, Tejani-Butt S. Fetal alcohol exposure alters serotonin transporter sites in rat brain.

Brain Res 2000;856:184-192.

91. Konarski JZ, McIntyre RS, Kennedy SH, et al. Volumetric neuroimaging investigations in mood disorders: bipolar disorder versus major depressive disorder. Bipolar Disord 2008;10:1-37.

92. Drevets WC, Thase ME, Moses-Kolko EL, et al. Serotonin1A receptor imaging in recurrent depression: Replication and literature review. Nucl Med Biol 2007;34:865-877.

93. Aggleton JP. The contribution of the amygdala to normal and abnormal emotional states. Trends Neurosci 1993;16:328-333.

94. Davis M. The role of the amygdala in fear and anxiety.

Ann Rev Neurosci 1992;15:353-375.

95. Campbell S, Macqueen G. The role of the hippocampus in the pathophysiology of major depression. J Psychiatry Neurosci 2004;29:417-426.

96. Sheline YI, Wang PW, Gado MH, et al. Hippocampal atrophy in recurrent major depression. Proc Natl Acad Sci USA 1996;93:3908-3913.

97. Videbech P, Ravnkilde B. Hippocampal volume and depression: A meta-analysis of MRI studies. Am J Psychiatry 2004;161:1957-1966.

98. Sheline YI, Gado MH, Kraemer HC. Untreated depression and hippocampal volume loss. Am J Psychiatry 2003;160:1516-1518.

99. Teicher MH, Andersen SL, Polcari A, et al. Developmental neurobiology of childhood stress and trauma. Psychiatr Clin North Am 2002;25:397-426, vii-viii.

100. Vythilingam M, Heim C, Newport J, et al. Childhood trauma associated with smaller hippocampal volume in women with major depression. Am J Psychiatry 2002;159:2072-2080.

101. Coe CL, Kramer M, Czeh B, et al. Prenatal stress diminishes neurogenesis in the dentate gyrus of juvenile rhesus monkeys. Biol Psychiatry 2003;54:1025-1034.

102. Szeszko PR, MacMillan S, McMeniman M, et al. Amygdala volume reductions in pediatric patients with obsessivecompulsive disorder treated with paroxetine: preliminary findings. Neuropsychopharmacology 2004;29:826-832.

103. McGee CL, Riley EP. Brain imaging and fetal alcohol spectrum disorders. Ann Ist Super Sanita 2006;42:46-52.

104. Dalley JW, Cardinal RN, Robbins TW. Prefrontal executive and cognitive functions in rodents: Neural and neurochemical substrates. Neurosci Biobehav Rev 2004;28:771-784.

105. Murmu MS, Salomon S, Biala Y, et al. Changes of spine density and dendritic complexity in the prefrontal cortex in offspring of mothers exposed to stress during pregnancy. Eur J Neurosci 2006;24:1477-1487.

106. Hayashi A, Nagaoka M, Yamada K, et al. Maternal stress induces synaptic loss and developmental disabilities of offspring. Int J Dev Neurosci 1998;16:209-216.

107. Zhu Z, Li X, Chen W, et al. Prenatal stress causes genderdependent neuronal loss and oxidative stress in rat hippocampus. J Neurosci Res 2004;78:837-844.

108. Ishiwata H, Shiga T, Okado N. Selective serotonin reuptake inhibitor treatment of early postnatal mice reverses their prenatal stress-induced brain dysfunction.

Neuroscience 2005;133:893-901.

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Isr J Psychiatry Relat Sci - Vol 47 - No. 1 (2010)

The Biology of Tryptophan Depletion and Mood Disorders Lilach Toker, BSc, 1* Shirly Amar, MSc,1* Yuly Bersudsky, MD, PhD,1 Jonathan Benjamin, MD,2 Ehud Klein, MD,3 and Galila Agam, PhD1 1

Psychiatry Research Unit, Faculty of Health Sciences, Ben-Gurion University of the Negev, and Mental Health Center, Beersheva, Israel Professor Emeritus, Clalit Sick Funds and Ben-Gurion University of the Negev, Beersheva, Israel 3 Department of Psychiatry, Rambam Medical Center and Bruce Rappaport Faculty of Medicine, Haifa, Israel 2

*These authors equally contributed to this work

ABSTRACT The involvement of the serotonergic system in the pathophysiology and treatment of affective disorders has been strongly implicated. The tryptophan depletion paradigm is widely used to study the effect of lowering serotonin levels. However, the effects observed in such studies are inconsistent and sometimes contradictory.

The present review summarizes and discusses these discrepancies, emphasizing the importance of methodological details such as acute vs. chronic tryptophan depletion, patientâ&#x20AC;&#x2122;s diagnosis and disease state (euthymic vs. acute phase) and previous drug treatment. Acute tryptophan depletion as a predictive test for personalized antidepressant treatment is suggested.

Introduction Tryptophan is an essential amino acid. Plasma tryptophan levels are determined by the balance between the dietary intake and its removal from plasma by protein synthesis.

Protein molar rate of tryptophan is 1.1% compared to ~ 5% of other amino acids, making it the rarest amino acid found in proteins. Tryptophan is transported across the blood-brain barrier (BBB) by a specific carrier for which tryptophan and all other large neutral amino acids (LNAAs) (valine, leucine, isoleucine, phenylalanine and tyrosine) also compete. The differential affinity ratio of

8:1 in favor of tryptophan secures its transport through the BBB into the brain (1). Some studies have shown that the level of brain tryptophan depends on the ratio between plasma concentrations of free tryptophan and other LNAAs (2) while others (3) showed that the ratio of total (free + albumin-bound) tryptophan and other LNAAs in serum is a better predicting parameter for brain tryptophan levels.

Tryptophan is the only precursor of serotonin (5-hydroxytryptamine, 5-HT). The serotonergic cells of the raphe possess a pacemaker-like activity that is modified by 5-HT autoreceptors and noradrenergic receptors.

Production of serotonin takes place both in the periphery and in the CNS. After entering the brain tryptophan is converted into serotonin in a two-step synthesis process.

L-tryptophan is first converted into 5-hydroxytryptophan by the enzyme tryptophan hydroxylase (Tph).

5-hydroxytryptophan is then decarboxylated by another enzyme, aromatic amino acid decarboxylase (5-hydroxyl-tryptophan decarboxylase) forming serotonin. There are two isoforms of tryptophan hydroxylase: Tph1 and Tph2.

Only the Tph2 isoform is expressed in the raphe neurons (4). Tph is the rate-limiting enzyme in the synthesis of serotonin (Fig. 1) since the Km of Tph for tryptophan is an order of magnitude higher than the Km of 5-HTP decarboxylase and 5-HTP is almost immediately decarboxylated to serotonin (5).

Serotonin modulates a number of developmental events such as neuronal migration, cell differentiation, cell division and synaptogenesis. 5-HT is involved in a wide array of CNS functions including the control of appetite, sleep, memory and learning, temperature

Address for Corrsepondence: Ehud Klein, MD,Chairman, Department of Psychiatry, Rambam Health-Care Campus, Faculty of Medicine - Technion IIT, Haifa, 31096 Israel. â&#x20AC;&#x2020;e_klein@rambam.health.gov.il

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regulation, mood, behavior (including sexual), perception, cardiovascular function, muscle contraction and endocrine regulation.

Multiple serotonin receptors have been cloned, designated 5HT1 through 5HT7. The 5HT1 group includes the subtypes 5HT1A, 5HT1B, 5HT1C, 5HT1D, 5HT1E, and 5HT1F. There are three 5HT2 subtypes, 5HT2A, 5HT2B, and 5HT2C as well as two 5HT5 subtypes, 5HT5A and 5HT5B. Most of these receptors are coupled to G-proteins that affect the activities of either adenylate cyclase or phospholipases. The 5HT3 class of receptors are ion channels (6).

The major pathway for serotonin degradation is reuptake into the neuron by a specific transporter, followed by degradation by monoamine oxidase (MAO) to yield 5-hydroxyindole acetaldehyde. This intermediate is metabolized by an aldehyde reductase or dehydrogenase to yield 5-hydroxytryptophol or 5-hydroxyindole acetic acid (5-HIAA) (Fig. 2).

Fig. 1: Serotonin degradation

Fig. 2: Serotonin synthesis

N HO

Serotonin (5-HT. 5-hydroxyptamine)

N

MAO

N HO

O

5-hydroxyindoleacetaldehyde (5-HIA) aldehyde dehydrogenase (ADH) N

+1H N 3 2

coo1-

Tryptophan Hyrdoxylase

C2 H CH2

HO coo1+1H N 3

HO

O2

N

CH2

HO

N

H +1H N 3 2 C2 H CH2

5-hydroxy-

N

serotonin

H

L-tryptophan +

HO

H

OH

5-hydroxyindoleacetic acid (5-HIAA)

C2 H

H

L-tryptohan

O

CO2

5-hydroxy-L-tryptophan Decarboxylase

Tryptophan depletion as a research paradigm for the study of mood regulation and the neurobiology of depression

Since the rate of 5-HT synthesis is dependent on plasma tryptophan, on how much of it crosses the BBB, and on the activity of brain tryptophan hydroxylase, depletion of plasma tryptophan resulting from an appropriate low-tryptophan diet, or, alternatively, inhibition of tryptophan hydroxylase, may modulate brain activity

and mood. The common paradigm to study tryptophan depletion involves an intervention day and a control day one week apart. During the 24 hours preceding each of the days subjects follow a low tryptophan diet (about 160 mg/day) and then remain in fasting overnight and throughout the next day. On the intervention day the subjects consume a tryptophan-free drink containing a 100 gm load of 15 amino acid mixture, including the LNAAs. On the control day subjects consume a nutritionally balanced drink that does contain tryptophan.

The intervention has two effects: it stimulates protein synthesis in the liver, which uses up free plasma tryptophan, and the LNAA amino acid family competes with tryptophan for the transport through the active protein shuttle across the BBB. This approach has been shown to produce maximal brain tryptophan depletion with maximal effects seen within 5-7 hrs after the depleting drink (7). The validity of acute tryptophan depletion by dietary amino acid loading as a method to diminish brain serotonin is supported by reports that ingestion 47

THE BIOLOGY OF TRYPTOPHAN DEPLETION AND MOOD DISORDERS

of tryptophan-free amino acid mixture in laboratory animals leads to depletion in plasma tryptophan levels and brain serotonin content (8). In humans ingestion of tryptophan-free amino acid mixture has been shown to produce a 70-90% reduction in plasma tryptophan 5-7 hours following administration (9-12). In healthy volunteers the tryptophan depletion paradigm resulted in 80 - 90% reduced CSF tryptophan levels after 7-10 hrs and 24-40% reduced 5-HIAA after 12-14 hrs (13).

Tryptophan depletion was also found to reduce the rate of serotonin synthesis by 90% of baseline values (14).

There are alternative ways to obtain serotonin depletion. One is the administration of serotonin synthesis inhibitors, such as the potent tryptophan hydroxylase inhibitor para-chlorophenylalanine (pCPA) (15, 16).

Another way is the administration of high doses of amphetamine analogs, e.g., para-chloroamphetamine (pCA) (17) and d-fenfluramine (d-FEN) (18) which elicit long-lasting decrease in serotonin levels by indirectly inhibiting tryptophan hydroxylase activity (17, 19). More invasive ways include electrolytic (20) or neurotoxic (20, 21) lesions of the median and dorsal raphe nuclei. The obvious advantages of the tryptophan depletion paradigm are that it is highly reliable and easily accomplished and that there is a clear cause and effect relationship. However, it also exhibits several disadvantages. Compounds other than serotonin may also be affected by alteration in tryptophan levels; e.g., plasma melatonin levels are altered by tryptophan depletion (22) and the levels of the tryptophan metabolites tryptamine, kynurenine and quinolinic acid also change. The last may be potentially psychoactive (23).

Tryptophan-deficient amino acid mixture induces decline in food intake in rats (24) but not in human subjects (12, 25), suggesting species difference in the response to the imbalanced amino acid mixture. In human subjects it is difficult and unfavorable to repeat the tryptophan depletion paradigm too often. From the practical point of view, the taste of the drink is aversive, and might induce involuntary vomiting. But there is also a health concern. Both forced feeding and ad-libitum tryptophan deficient diet compared to control diet resulted in weight loss in rats (26). Early animal studies have shown that prolonged treatment with tryptophan depleted diet caused pathological changes in eyes, liver (27), blood (28), pancreas and spine (29). It is however notable that: 1) The extreme behavioral pattern in tryptophan depleted animals reported in these early studies (27) is absent from more recent studies. 2) The harm48

ful effect of tryptophan depletion could be reversed by seven-day treatment with control diet (27). 3) Some of the pathological findings such as blood changes (28) and eye pathologies (30) were seen after more than 40 days of tryptophan depletion, a period length never applied to human subjects. 4) The experimental animals used in these studies were at their developmental stage which may render higher susceptibility to the depletion of an essential amino acid. In humans, the tryptophan depletion paradigm may obviously only be employed for a relatively short duration, inducing temporary biochemical changes.

The effects of chronic lowered brain serotonin on mood and cognition may differ both quantitatively and qualitatively from those produced by acute tryptophan depletion. Bortolato et al. (31) have shown different effects of acute and chronic tryptophan depletion on prepulse inhibition of the acoustic startle response in rats. It is possible that 5-HT1A and 5-HT2A regulation by acute vs. chronic tryptophan depletion (32) is responsible for the different response. Similar differences are also known to exist between acute vs. chronic administration of serotonin selective reuptake inhibitors (SSRI) (33-36).

In summary, tryptophan depletion provides a useful and interesting tool to investigate the role of serotonin in healthy volunteers, in the pathophysiology of psychiatric disorders, primarily affective disorders, and in understanding the involvement of serotonin in the mechanism of action of drugs such as SSRIs, MAO inhibitors (MAOI), lithium and non-pharmacological treatments such as ECT and sleep deprivation.

Effect of tryptophan depletion on mood in healthy subjects

Numerous studies but not all (37) have shown that tryptophan depletion does not significantly affect mood in healthy male subjects (7, 10, 13, 38, 39). Healthy women, however, were more susceptible than healthy men to mood lowering following tryptophan depletion (40).

This might be related to the fact that tryptophan depletion causes a more significant decrease in the rate of serotonin synthesis in women (41). The gender-related differences in serotonin synthesis could be related to early serotonergic events in brain organization or to effects of circulating gonadal hormones (42-44). People at an elevated genetic risk for mood disorders, such as those with a multigenerational family history of affective disorders, showed an increased vulnerability to

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mood alteration during tryptophan depletion (10, 45, 46). However, a subsequent study did not replicate these findings (47).

Tryptophan depletion in major depression

Serotonin has strongly been implicated in the pathophysiology of depressive syndromes and in the mechanism of antidepressant drug action. Studies of untreated depressed patients suggest that serotonin function is reduced in depression, and studies of treated patients indicate that the mechanism of action of antidepressants is mediated by the enhancement of serotonin and/or noradrenaline neurotransmission.

Tryptophan depletion in untreated depressed patients

A study of the effect of tryptophan depletion on untreated depressed patients reported no mood change during the depletion day; however, on the next day there was a bimodal response, with 23% of the patients describing a clinically significant worsening of depression, 37% reporting an improvement in symptoms and the remaining 40% reporting no change in symptoms, all compared to a placebo group (48). The lack of effect on the day of the depletion may have resulted from a floor effect due to maximally reduced serotonin function in these patients. The possibility that the lack of response to tryptophan depletion on the depletion day was caused by a floor effect is supported by the following: out of the initially-studied patients 15 underwent an additional tryptophan depletion session while being euthymic following successful antidepressant treatment.

Nine of them did experience relapse of depressive symptoms on the depletion day (9). An alternative suggestion for the lack of response to tryptophan depletion on the depletion day is that depression is not caused by direct aberrant serotonin function but, rather, by dysfunctional serotonin-regulated brain circuits. If so, inducing further decrease in brain 5-HT would not necessarily result in an immediate effect.

In Delgado et al.ážżs study (48), participants showed strong correlation between the effect obtained on the day after tryptophan depletion and further response to antidepressants. Antidepressant-responders were more likely to improve in depression score following tryptophan depletion, while nonresponders were those whose depressive symptoms worsened following tryptophan depletion. Such bimodal mood changes on the day after tryptophan depletion and their relationship to treatment response should be interpreted with caution. Exclusion

criteria for participants did not include personality disorder and 11 out of the 43 depressed subjects studied were concomitantly diagnosed as having a DSM-III-R personality disorder. These subjects were both more likely not to respond to antidepressant treatment and to exhibit increased Hamilton-depression (Ham-D) score.

In addition, the authors do not report the exclusion of bipolar depressed subjects. Such a diversity in diagnosis may have led to the different response to tryptophan depletion. Tryptophan depletion directly affects serotonin levels while antidepressants do not achieve their effect just by increasing serotonin levels. If that would have been the case the beneficial effect of the drugs should have been observed earlier than after 2-3 weeks of treatment.

Improvement in depressive symptoms observed in some of the patients on the day after the administration of tryptophan depletion (48), when 5-HT levels are already restored, could be due to regulation of selected serotonin receptors. Binding potential of 5-HT1A autoreceptors but not 5-HT1A postsynaptic receptors was shown to be reduced in rats three hours after acute tryptophan depletion (32). 5-HT2C receptors, shown to exhibit inhibitory effect on dopamine transmission (49, 50), show paradoxical regulation such as down-regulation in response to their blockade (51-53). Receptor modulation could also occur through pre-mRNA editing. Conversion of adenosine to inosine at specific sites of 5-HT2C pre-mRNA can result in protein isoforms with different efficiency to activate G-protein in response to agonist stimulation (54). The editing enzymes recognize five closely spaced adenosine residues (A-E) located within sequences encoding the intracellular domain of the receptor and convert the adenosine residues to inosines. The result of the editing in the C and C' site is expression of 5-HT2c isoforms that are less efficient in activating G-proteins (55). The C' site editing is significantly increased, D site editing is significantly decreased and the C site shows a trend toward increased editing in the dorsal prefrontal cortex of suicide victims with a history of major depression.

In contrast, mice treated chronically with fluoxetine showed opposite alterations in 5-HT2C pre-mRNA editing from those detected in suicide victims. The changes in editing-site in response to chronic fluoxetine treatment in mice are paradoxically similar to those detected in serotonin-depleted mice (56). Possible explanation for this phenomenon is that fluoxetine-induced alteration in 5-HT2c pre-mRNA editing is not a result of 49

THE BIOLOGY OF TRYPTOPHAN DEPLETION AND MOOD DISORDERS

simply increasing synaptic serotonin, but rather of receptor blockade by the drug (57). This is further supported by the fact that treatment with a 5-HT2c agonist increased the pre-mRNA editing at the C' site, resulting in expression of less active isoforms of 5-HT2c. It may be suggested that in subjects in whom the regulation of the serotonergic system functions normally, tryptophan depletion would result in increased sensitivity of the system. Namely, serotonin repletion would result in transient improvement of the symptoms. Similarly, depressive patients with functioning regulation of the system would also be expected to respond to serotonergic antidepressants. On the other hand, it is possible that patients whose condition worsened the day after tryptophan depletion (48) may be the ones with primary dysfunction of the regulation of the serotonergic system. In such subjects response to serotonergic antidepressants may not occur. Tryptophan depletion and repletion non-responders possibly exhibit dysfunction of a non-serotonergic system.

Animal studies of chronic serotonin depletion, as well as animal and human studies of acute and chronic serotonin reuptake blockade, and of direct serotonin antagonists, can complement the tryptophan depletion paradigm and support its implications or suggest an alternative mechanism of action of antidepressant drugs.

Tryptophan depletion in remitted untreated patients with

role of serotonin in the mechanism of action of antidepressants. Tryptophan depletion produced relapse in patients responding to SSRIs, MAOIs and phototherapy (61, 64, 65, 70-73), but did not reverse the beneficial effect of tricyclics (9, 70, 71), ECT (74), sleep deprivation (75) or lithium (9, 76, 77). Since SSRIs and MAOIs intervene specifically with the serotonergic system while the tricyclics, sleep deprivation and lithium affect multiple neurotransmitter systems, e.g., adrenergic and dopaminergic, it is conceivable that tryptophan depletion would counteract the effect of SSRIs and MAOIs but not the other treatment modalities. Higher rates of relapse were observed in patients on serotonergic than on adrenergic drugs (71). The rate of transient exacerbation of depressive symptoms in patients on SSRIs or MAOIs was higher (80%) in patients recently remitted than those remitted for an average of 45 weeks (30%) (64). Similarly, recently remitted seasonal affective disorder (SAD) patients after light therapy relapsed following tryptophan depletion (72, 75), while for long-term remitted patients, one study resulted in a relapse rate of 73% (61), but another showed no effect of tryptophan depletion on symptoms (63). It may be that tryptophan depletion induced relapse in the more recently remitted patients because increasing synaptic serotonin is a primary step in a cascade of events that results in adaptive changes in serotonergic neurons, while at the adaptive stage an interruption to serotonin function is less adversive.

major depression

In remitted untreated subjects with a past history of depression transient reappearance of depressive symptoms following tryptophan depletion was reported in some studies (58-61) but not in others (62, 63). A positron emission tomography (PET) study found that relapse of depressive symptoms in remitted patients following tryptophan depletion is associated with decreased metabolic activity in brain areas implicated in the pathogenesis of depression (64, 65).

Tryptophan depletion and depressive symptoms in SSRItreated remitted patients

Acute tryptophan depletion resulted in reduced binding potential of 5-HT1A autoreceptors in rats (32), but not in SSRI-treated remitted patients (66). It is well established that chronic SSRI treatment reduces 5-HT1A autoreceptor function (67-69). It is possible that the lack of effect of tryptophan depletion on 5-HT1A autoreceptors under chronic SSRI treatment reflects a floor effect.

Tryptophan depletion was also used to examine the 50

Tryptophan depletion in bipolar disorder

Tryptophan depletion in recovered bipolar patients (77), lithium-maintained patients (74, 78) and recently remitted bipolar patients (79) did not result in relapse of bipolar symptoms. In addition, tryptophan depletion did not produce noticeable effects on the neuropsychological performance of euthymic bipolar patients (80). Shopsin et al. (81) studied five depressed patients out of whom two had bipolar depression. All patients responded to the MAO inhibitor antidepressant, tranylcypromine, and relapsed when the tryptophan hydroxylase inhibitor pCPA was added to the treatment. One of the bipolar patients became manic after four weeks of tranylcypromine treatment while a week of combined treatment with pCPA brought the patient to a euthymic state. This kind of cycling was repeated in this subject but the pattern was not observed in the other bipolar patient.

Recently, a new therapeutic use for tryptophan depletion has been evaluated in acutely manic patients (82).

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The results from the double blind placebo controlled study of 17 manic patients showed a clinically and statistically significant difference in the Young Mania Scale between nine patients who received the tryptophan free amino acid drink and the eight patients who received the placebo drink. In the latter study the composition of the amino acid mixture used to cause tryptophan depletion was the same as previously reported (77, 80) but the regime of seven-day administration of the tryptophan depleted mixture did not include pretreatment with low tryptophan diet. Since plasma tryptophan levels were not assayed, Applebaum et al.'s (82) study lacks an objective proof of the degree of tryptophan depletion obtained.

The recruited patients were in a manic state and received valproic acid during the trial while euthymic bipolar subjects were studied previously (74, 77, 78). It is possible that reduction in brain serotonin affects differently remitted vs. manic patients, as seems to be the case in depressed vs. remitted unipolar patients.

Tryptophan depletion and genetics

The response to tryptophan depletion could depend on genetic factors. Polymorphisms in the promoter region of the serotonergic transporter (5HTTLRP) have been found to be associated with unipolar depression (83-87).

Moreno et al. reported an increased risk of drug-treated remitted major depressive patients with the l/l genotype of 5-HTTLRP for depressive mood response to tryptophan depletion (88). In healthy subjects, however, Neumeister et al. (89, 90) found an inverse association.

Summary Despite several decades of studies of the effect of tryptophan depletion using different paradigms, there are yet multiple open issues. Booij et al. (91) reported that high but not moderate tryptophan depletion induced depressive symptoms in patients remitted from depression. Information regarding different regulation pattern of serotonin receptors by acute vs. chronic tryptophan depletion, and discrepancies among studies suggest that methodology details might be critical for the treatment outcome (92). However, given the potential risk in applying chronic tryptophan depletion to human beings (93, 94) further studies of this paradigm should be used for basic research, as a tool towards the understanding of the relationship between serotonin and mood disorders. Potentially, if the finding that depressive patients who responded to acute tryptophan depletion

and repletion were more likely to be antidepressant responders would be substantiated, response to acute tryptophan depletion may be found of predictive value to personalized antidepressant treatment.

References 1. McAllister-Williams RH, Massey AE, Rugg MD. Effects of tryptophan depletion on brain potential correlates of episodic memory retrieval. Psychopharmacology 2002;160:434-442.

2. Perez-Cruet J, Chase TN, Murphy DL. Dietary regulation of brain tryptophan metabolism by plasma ratio of free tryptophan and neutral amino acids in humans. Nature 1974;248:693-695.

3. Fernstrom JD, Hirsch MJ, Faller DV. Tryptophan concentrations in rat brain. Failure to correlate with free serum tryptophan or its ratio to the sum of other serum neutral amino acids. Biochem J 1976;160:589-595.

4. Walther DJ, Bader M. A unique central tryptophan hydroxylase isoform. Biochem Pharmacol 2003;66:1673-1680.

5. Cooper JR, Bloom FE, Roth RH. The biochemical basis of neuropharmacology. Eighth Edition. New York: Oxford University, 2003.

6. Smith CUM. Elements of molecular neurobiology. Second Edition. New York: Wiley, 1996.

7. Reilly JG, McTavish SF, Young AH. Rapid depletion of plasma tryptophan: A review of studies and experimental methodology. J Psychopharmacol 1997;11:381-392.

8. Lieben CK, Blokland A, Westerink B, Deutz NE. Acute tryptophan and serotonin depletion using an optimized tryptophan-free protein-carbohydrate mixture in the adult rat. Neurochem Int 2004;44:9-16.

9. Delgado PL, Charney DS, Price LH, Aghajanian GK, Landis H, Heninger GR. Serotonin function and the mechanism of antidepressant action. Reversal of antidepressant-induced remission by rapid depletion of plasma tryptophan.

Arch Gen Psychiatry 1990;47:411-418.

10. Benkelfat C, Ellenbogen MA, Dean P, Palmour RM, Young SN. Mood-lowering effect of tryptophan depletion. Enhanced susceptibility in young men at genetic risk for major affective disorders. Arch Gen Psychiatry 1994;51:687-697.

11. Young SN, Smith SE, Pihl RO, Ervin FR. Tryptophan depletion causes a rapid lowering of mood in normal males.

Psychopharmacology 1985;87:173-177.

12. Young SN, Tourjman SV, Teff KL, Pihl RO, Anderson GH. The effect of lowering plasma tryptophan on food selection in normal males. Pharmacol Biochem Behav 1988;31:149-152.

51

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13. Carpenter LL, Anderson GM, Pelton GH, Gudin JA, Kirwin PD, Price LH, et al. Tryptophan depletion during continuous CSF sampling in healthy human subjects.

Neuropsychopharmacology 1998;19:26-35.

14. Young SN, Leyton M, Benkelfat C. Pet studies of serotonin synthesis in the human brain. Adv Exp Med Biol 1999;467:11-18.

15. Miczek KA, Altman JL, Appel JB, Boggan WO. Parachlorophenylalanine, serotonin and killing behavior.

Pharmacol Biochem Behav 1975;3:355-361.

16. Herges S, Taylor DA. Modulatory effect of p-chlorophenylalanine microinjected into the dorsal and median raphe nuclei on cocaine-induced behaviour in the rat. Eur J Pharmacol 1999;374:329-340.

17. Sanders-Bush E, Bushing JA, Sulser F. p-Chloroamphetamine-inhibition of cerebral tryptophan hydroxylase.

Biochem Pharmacol 1972;21:1501-1510.

18. Harvey JA, McMaster SE. Fenfluramine: evidence for a neurotoxic action on midbrain and a long-term depletion of serotonin. Psychopharmacol Commun 1975;1:217-228.

19. Fuller RW. Effects of p-chloroamphetamine on brain serotonin neurons. Neurochem Res 1992;17:449-456.

20. Andrade TG, Graeff FG. Effect of electrolytic and neurotoxic lesions of the median raphe nucleus on anxiety and stress. Pharmacol Biochem Behav 2001;70:1-14.

21. Richter-Levin G, Segal M. The effects of serotonin depletion and raphe grafts on hippocampal electrophysiology and behavior. J Neurosci 1991;11:1585-1596.

22. Zimmermann RC, McDougle CJ, Schumacher M, Olcese J, Mason JW, Heninger GR, et al. Effects of acute tryptophan depletion on nocturnal melatonin secretion in humans. J Clin Endocrinol Metab 1993;76:1160-1164.

23. Oxenkrug GF. Tryptophan–kynurenine metabolism as a common mediator of genetic and environmental impacts in major depressive disorder: “The serotonin hypothesis” revisited 40 years later. Isr J Psychiatry and Relat Sci 2010; 47: 56-63.

24. Harper AE, Benevenga NJ, Wohlhueter RM. Effects of ingestion of disproportionate amounts of amino acids.

Physiol Rev 1970;50:428-558.

25. Weltzin TE, Fernstrom MH, Fernstrom JD, Neuberger SK, Kaye WH. Acute tryptophan depletion and increased food intake and irritability in bulimia nervosa. Am J Psychiatry 1995;152:1668-1671.

26. Lee YS, Naito H, Kametaka M. The effect of force feeding a tryptophan-deficient diet on arterio-venous difference of plasma amino acids across skeletal muscle of rats. J Nutr 1977;109:119-128.

27. Spector H, Adamstone FB. Tryptophan deficiency in the rat induced by forced feeding of an acid hydrolyzed casein 52

diet. J Nutr 1950;40:213-229.

28. Albanese AA, Emmet Holt E, Kajdi CN, Frankston JE. Observation on tryptophane deficiency in rats chemical and morphological changes in the blood. J Biol Chem 1943;148:299-309.

29. Sidransky H. Tryptophan: Biochemical and health implications, chapter 2: Findings in deficiency states, pp. 8-18.

Boca Raton, Florida: CRC, 2001.

30. Vrensen GF, van Marle J, Jonges R, Voorhout W, Breipohl W, Wegener AR. Tryptophan deficiency arrests chromatin breakdown in secondary lens fibers of rats. Exp Eye Res 2004;78:661-672.

31. Bortolato M, Frau R, Orru M, Collu M, Mereu G, Carta M, et al. Effects of tryptophan deficiency on prepulse inhibition of the acoustic startle in rats. Psychopharmacology (Berl) 2008;198:191-200.

32. Cahir M, Ardis T, Reynolds GP, Cooper SJ. Acute and chronic tryptophan depletion differentially regulate central 5-HT1A and 5-HT 2A receptor binding in the rat.

Psychopharmacology (Berl) 2007;190:497-506.

33. Jensen JB, Jessop DS, Harbuz MS, Mork A, Sanchez C, Mikkelsen JD. Acute and long-term treatments with the selective serotonin reuptake inhibitor citalopram modulate the HPA axis activity at different levels in male rats. J Neuroendocrinol 1999;11:465-471.

34. Burghardt NS, Sullivan GM, McEwen BS, Gorman JM, LeDoux JE. The selective serotonin reuptake inhibitor citalopram increases fear after acute treatment but reduces fear with chronic treatment: a comparison with tianeptine. Biol Psychiatry 2004;55:1171-1178.

35. Shishkina GT, Kalinina TS, Dygalo NN. Up-regulation of tryptophan hydroxylase-2 mRNA in the rat brain by chronic fluoxetine treatment correlates with its antidepressant effect. Neuroscience 2007;150:404-412.

36. Prisco S, Esposito E. Differential effects of acute and chronic fluoxetine administration on the spontaneous activity of dopaminergic neurones in the ventral tegmental area. Br J Pharmacol 1995;116:1923-1931.

37. Young SN, Smith SE, Pihl RO, Ervin FR. Tryptophan depletion causes a rapid lowering of mood in normal males.

Psychopharmacology (Berl) 1985;87:173-177.

38. Moore P, Landolt HP, Seifritz E, Clark C, Bhatti T, Kelsoe J, et al. Clinical and physiological consequences of rapid tryptophan depletion. Neuropsychopharmacology 2000;23:601-622.

39. Knott VJ, Howson AL, Perugini M, Ravindran AV, Young SN. The effect of acute tryptophan depletion and fenfluramine on quantitative EEG and mood in healthy male subjects. Biol Psychiatry 1999;46:229-238.

40. Ellenbogen MA, Young SN, Dean P, Palmour RM,

Lilach Toker ET AL.

Benkelfat C. Mood response to acute tryptophan depletion in healthy volunteers: Sex differences and temporal stability. Neuropsychopharmacology 1996;15:465-474.

41. Nishizawa S, Benkelfat C, Young SN, Leyton M, Mzengeza S, de Montigny C, et al. Differences between males and females in rates of serotonin synthesis in human brain. Proc Natl Acad Sci USA 1997;94:5308-5313.

42. Rubinow DR, Schmidt PJ, Roca CA. Estrogen-serotonin interactions: Implications for affective regulation. Biol Psychiatry 1998;44:839-850.

43. Epperson CN, Amin Z, Naftolin F, Cappiello A, Czarkowski KA, Stiklus S, et al. The resistance to depressive relapse in menopausal women undergoing tryptophan depletion: Preliminary findings. J Psychopharmacol 2007;21:414-420.

44. Jans LA, Lieben CK, Blokland A. Influence of sex and estrous cycle on the effects of acute tryptophan depletion induced by a gelatin-based mixture in adult Wistar rats.

Neuroscience 2007;147:304-317.

45. Quintin P, Benkelfat C, Launay JM, Arnulf I, PointereauBellenger A, Barbault S, et al. Clinical and neurochemical effect of acute tryptophan depletion in unaffected relatives of patients with bipolar affective disorder. Biol Psychiatry 2001;50:184-190.

46. Klaassen T, Riedel WJ, van Someren A, Deutz NE, Honig A, van Praag HM. Mood effects of 24-hour tryptophan depletion in healthy first-degree relatives of patients with affective disorders. Biol Psychiatry 1999;46:489-497.

47. Ellenbogen MA, Young SN, Dean P, Palmour RM, Benkelfat C. Acute tryptophan depletion in healthy young women with a family history of major affective disorder. Psychol Med 1999;29:35-46.

48. Delgado PL, Price LH, Miller HL, Salomon RM, Aghajanian GK, Heninger GR, et al. Serotonin and the neurobiology of depression. Effects of tryptophan depletion in drug-free depressed patients. Arch Gen Psychiatry 1994;51:865-874.

49. Alex KD, Yavanian GJ, McFarlane HG, Pluto CP, Pehek EA. Modulation of dopamine release by striatal 5-HT2C receptors. Synapse 2005;55:242-251.

50. Porras G, Di Matteo V, Fracasso C, Lucas G, De Deurwaerdere P, Caccia S, et al. 5-HT2A and 5-HT2C/2B receptor subtypes modulate dopamine release induced in vivo by amphetamine and morphine in both the rat nucleus accumbens and striatum. Neuropsychopharmacology 2002;26:311-324.

51. Leysen JE, Van Gompel P, Gommeren W, Woestenborghs R, Janssen PA. Down regulation of serotonin-S2 receptor sites in rat brain by chronic treatment with the serotonin-S2 antagonists: Ritanserin and setoperone.

Psychopharmacology 1986;88:434-444.

52. Pranzatelli MR. Regulation of 5-HT2 receptors in rat cor-

tex. Studies with a putative selective agonist and an antagonist. Biochem Pharmacol 1991;42:1099-1105.

53. Pranzatelli MR, Murthy JN, Tailor PT. Novel regulation of 5-HT1C receptors: Down-regulation induced both by 5-HT1C/2 receptor agonists and antagonists. Eur J Pharmacol 1993;244:1-5.

54. Burns CM, Chu H, Rueter SM, Hutchinson LK, Canton H, Sanders-Bush E, et al. Regulation of serotonin-2C receptor G-protein coupling by RNA editing. Nature 1997;387:303-308.

55. Gurevich I, Tamir H, Arango V, Dwork AJ, Mann JJ, Schmauss C. Altered editing of serotonin 2C receptor premRNA in the prefrontal cortex of depressed suicide victims. Neuron 2002;34:349-356.

56. Gurevich I, Englander MT, Adlersberg M, Siegal NB, Schmauss C. Modulation of serotonin 2C receptor editing by sustained changes in serotonergic neurotransmission. J Neurosci 2002;22:10529-10532.

57. Ni YG, Miledi R. Blockage of 5HT2C serotonin receptors by fluoxetine (Prozac). Proc Natl Acad Sci USA 1997;94:2036-2040.

58. Smith KA, Fairburn CG, Cowen PJ. Relapse of depression after rapid depletion of tryptophan. Lancet 1997;349:915-919.

59. Moreno FA, Gelenberg AJ, Heninger GR, Potter RL, McKnight KM, Allen J, et al. Tryptophan depletion and depressive vulnerability. Biol Psychiatry 1999;46:498-505.

60. Leyton M, Ghadirian AM, Young SN, Palmour RM, Blier P, Helmers KF, et al. Depressive relapse following acute tryptophan depletion in patients with major depressive disorder. J Psychopharmacol 2000;14:284-287.

61. Neumeister A, Praschak-Rieder N, Hesselmann B, Vitouch O, Rauh M, Barocka A, et al. Effects of tryptophan depletion in fully remitted patients with seasonal affective disorder during summer. Psychol Med 1998;28:257-264.

62. Leyton M, Young SN, Benkelfat C. Relapse of depression after rapid depletion of tryptophan. Lancet 1997;349:1840-1841.

63. Lam RW, Bowering TA, Tam EM, Grewal A, Yatham LN, Shiah IS, et al. Effects of rapid tryptophan depletion in patients with seasonal affective disorder in natural summer remission. Psychol Med 2000;30:79-87.

64. Bremner JD, Innis RB, Salomon RM, Staib LH, Ng CK, Miller HL, et al. Positron emission tomography measurement of cerebral metabolic correlates of tryptophan depletion-induced depressive relapse. Arch Gen Psychiatry 1997;54:364-374.

65. Smith KA, Morris JS, Friston KJ, Cowen PJ, Dolan RJ. Brain mechanisms associated with depressive relapse and associated cognitive impairment following acute tryptophan depletion. Br J Psychiatry 1999;174:525-529.

66. Praschak-Rieder N, Hussey D, Wilson AA, Carella A, Lee 53

THE BIOLOGY OF TRYPTOPHAN DEPLETION AND MOOD DISORDERS

M, Dunn E, et al. Tryptophan depletion and serotonin loss in selective serotonin reuptake inhibitor-treated depression: An [(18)F] MPPF positron emission tomography study. Biol Psychiatry 2004;56:587-591.

67. Cornelisse LN, Van der Harst JE, Lodder JC, Baarendse PJ, Timmerman AJ, Mansvelder HD, et al. Reduced 5-HT1Aand GABAB receptor function in dorsal raphe neurons upon chronic fluoxetine treatment of socially stressed rats.

J Neurophysiol 2007;98:196-204.

68. McGehee DS. GIRK channels as a target for SSRIs. Focus on â&#x20AC;&#x153;reduced 5-HT1A- and GABAB receptor function in dorsal raphe neurons upon chronic fluoxetine treatment of socially stressed rats.â&#x20AC;? J Neurophysiol 2007;98:1-2.

69. Rossi DV, Burke TF, McCasland M, Hensler JG. Serotonin1A receptor function in the dorsal raphe nucleus following chronic administration of the selective serotonin reuptake inhibitor sertraline. J Neurochem 2008;105:1091-1099.

70. Delgado PL, Price LH, Miller HL, Salomon RM, Licinio J, Krystal JH, et al. Rapid serotonin depletion as a provocative challenge test for patients with major depression: Relevance to antidepressant action and the neurobiology of depression. Psychopharmacol Bull 1991;27:321-330.

71. Delgado PL, Miller HL, Salomon RM, Licinio J, Krystal JH, Moreno FA, et al. Tryptophan-depletion challenge in depressed patients treated with desipramine or fluoxetine: Implications for the role of serotonin in the mechanism of antidepressant action. Biol Psychiatry 1999;46:212-220.

72. Lam RW, Zis AP, Grewal A, Delgado PL, Charney DS, Krystal JH. Effects of rapid tryptophan depletion in patients with seasonal affective disorder in remission after light therapy. Arch Gen Psychiatry 1996;53:41-44.

73. Neumeister A, Praschak-Rieder N, Besselmann B, Rao ML, Gluck J, Kasper S. Effects of tryptophan depletion on drug-free patients with seasonal affective disorder during a stable response to bright light therapy. Arch Gen Psychiatry 1997;54:133-138.

74. Hughes JH, Dunne F, Young AH. Effects of acute tryptophan depletion on mood and suicidal ideation in bipolar patients symptomatically stable on lithium. Br J Psychiatry 2000;177:447-451.

75. Neumeister A, Praschak-Rieder N, Hesselmann B, Vitouch O, Rauh M, Barocka A, et al. Effects of tryptophan depletion in drug-free depressed patients who responded to total sleep deprivation. Arch Gen Psychiatry 1998;55:167-172.

76. Benkelfat C, Seletti B, Palmour RM, Hillel J, Ellenbogen M, Young SN. Tryptophan depletion in stable lithium-treated patients with bipolar disorder in remission. Arch Gen Psychiatry 1995;52:154-156.

77. Cassidy F, Murry E, Carroll BJ. Tryptophan depletion in recently manic patients treated with lithium. Biol Psychiatry 54

1998;43:230-232.

78. Johnson L, El-Khoury A, Aberg-Wistedt A, StainMalmgren R, Mathe AA. Tryptophan depletion in lithium-stabilized patients with affective disorder. Int J Neuropsychopharmacol 2001;4:329-336.

79. Cappiello A, Sernyak MJ, Malison RT, McDougle CJ, Heninger GR, Price LH. Effects of acute tryptophan depletion in lithium-remitted manic patients: A pilot study. Biol Psychiatry 1997;42:1076-1078.

80. Hughes JH, Gallagher P, Young AH. Effects of acute tryptophan depletion on cognitive function in euthymic bipolar patients. Eur Neuropsychopharmacol 2002;12:123-128.

81. Shopsin B, Friedman E, Gershon S. Parachlorophenylalanine reversal of tranylcypromine effects in depressed patients.

Arch Gen Psychiatry 1976;33:311-319.

82. Applebaum J, Bersudsky Y, Klein E. Rapid tryptophan depletion as a treatment for acute mania: A double-blind, pilot-controlled study. Bipolar Disord 2007 ;9:884-887.

83. Minov C, Baghai TC, Schule C, Zwanzger P, Schwarz MJ, Zill P, et al. Serotonin-2A-receptor and -transporter polymorphisms: Lack of association in patients with major depression. Neurosci Lett 2001;303:119-122.

84. Mann JJ, Huang YY, Underwood MD, Kassir SA, Oppenheim S, Kelly TM, et al. A serotonin transporter gene promoter polymorphism (5-HTTLPR) and prefrontal cortical binding in major depression and suicide. Arch Gen Psychiatry 2000;57:729-738.

85. Lotrich FE, Pollock BG, Ferrell RE. Polymorphism of the serotonin transporter: implications for the use of selective serotonin reuptake inhibitors. Am J Pharmacogenomics 2001;1:153-164.

86. Frodl T, Meisenzahl EM, Zill P, Baghai T, Rujescu D, Leinsinger G, et al. Reduced hippocampal volumes associated with the long variant of the serotonin transporter polymorphism in major depression. Arch Gen Psychiatry 2004;61:177-183.

87. Lesch KP, Mossner R. Genetically driven variation in serotonin uptake: Is there a link to affective spectrum, neurodevelopmental, and neurodegenerative disorders? Biol Psychiatry 1998;44:179-192.

88. Moreno FA, Rowe DC, Kaiser B, Chase D, Michaels T, Gelernter J, et al. Association between a serotonin transporter promoter region polymorphism and mood response during tryptophan depletion. Mol Psychiatry 2002;7:213-216.

89. Neumeister A, Konstantinidis A, Stastny J, Schwarz MJ, Vitouch O, Willeit M, et al. Association between serotonin transporter gene promoter polymorphism (5HTTLPR) and behavioral responses to tryptophan depletion in healthy women with and without family history of depression. Arch Gen Psychiatry 2002;59:613-620.

Lilach Toker ET AL.

90. Neumeister A, Hu XZ, Luckenbaugh DA, Schwarz M, Nugent AC, Bonne O, et al. Differential effects of 5-HTTLPR genotypes on the behavioral and neural responses to tryptophan depletion in patients with major depression and controls. Arch Gen Psychiatry 2006;63:978-986.

92. Merens W, van der Does W. Low-dose tryptophan depletion.

Biol Psychiatry 2007;62:542-543; author reply 543-544.

91. Booij L, Van der Does AJ, Haffmans PM, Riedel WJ, Fekkes D, Blom MJ. The effects of high-dose and low-dose tryptophan depletion on mood and cognitive functions of remitted depressed patients. J Psychopharmacol 2005;19:267-275.

94. Zepf FD, Wรถckel L, Herpertz-Dahlmann B, Poustka F, Freitag CM. Tryptophan depletion in bipolar mania: Reduction of metabolic imbalance by administration according to body weight? Bipolar Disord 2009;11:557-558.

93. Young SN. Rapid tryptophan depletion as a treatment for acute mania: Safety and mechanism of the therapeutic effect. Bipolar Disord 2008;10:850-851.

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Isr J Psychiatry Relat Sci - Vol 47 - No. 1 (2010)

Tryptophan–Kynurenine Metabolism as a Common Mediator of Genetic and Environmental Impacts in Major Depressive Disorder: The Serotonin Hypothesis Revisited 40 Years Later Gregory F. Oxenkrug, MD, PhD Department of Psychiatry, Tufts University School of Medicine and Tufts Medical Center, Boston, Massachusetts, U.S.A.

ABSTRACT The original 1969 Lancet paper proposed, “ in depression the activity of liver tryptophan-pyrrolase is stimulated by raised blood corticosteroids levels, and metabolism of tryptophan is shunted away from serotonin production, and towards kynurenine production.” Discovery of neurotropic activity of kynurenines suggested that up-regulation of the tryptophan-kynurenine pathway not only augmented serotonin deficiency but also underlined depressionassociated anxiety, psychosis and cognitive decline.

The present review of genetic and hormonal factors regulating kynurenine pathway of tryptophan metabolism suggests that this pathway mediates both genetic and environmental mechanisms of depression.

Rate-limiting enzymes of kynurenine formation, tryptophan 2,3-dioxygenase (TDO) and indoleamine 2,3-dioxygenase (IDO) are activated by stress hormones (TDO) and/or by pro-inflammatory cytokines (IDO).

Simultaneous presence of high producers alleles of proinflammatory cytokines genes (e.g., interferon-gamma and tumor necrosis factor-alpha) determines the genetic predisposition to depression via up-regulation of IDO while impact of environmental stresses is mediated via hormonal activation of TDO. Tryptophankynurenine pathway represents a major meeting point of gene-environment interaction in depression and a new target for pharmacological intervention.

Although often referred to as “serotonin hypothesis,” the 1969 Lancet paper proposed the disturbances of tryptophan (TRY) metabolism, i.e., the shunt of TRY from serotonin (5-HT) synthesis to kynurenine (KYN) formation, as a major etiological factor of depression (1).

It suggested the formation of “vicious cycle” perpetuating the increase of KYN and decrease of 5-HT production in depression due to a) stress hormones – induced activation of tryptophan 2,3-dioxygenase (TDO), the rate-limiting enzyme of TRY – KYN pathway; b) diminished availability of TRY as an initial substrate of 5-HT biosynthesis due to increased formation of KYN from TRY; and c) increased production of cortisol due to weakening of 5-HT inhibitory effect on amygdaloidal complex (2) (Fig. 1).

Figure 1. Shunt of TRY metabolism from 5-HT to KYN production in depression (1) Abbreviations: TRY – tryptophan; 5-HT – serotonin; TDO – tryptophan 2,3-dioxygenase TRY

5-HT (brain)

TDO (Liver)

cortisol (adrenals)

5-HT deficiency was thought as a major consequence of the shift of TRY metabolism to KYN formation, and “intensification of the central 5-HT-ergic processes” was suggested as “a possible determinant of the thymoleptic (mood-elevating) effect” (1). Introduction and wide

Address for Correspondence: Gregory F. Oxenkrug, MD, PhD, Director, Psychiatry and Inflammation Program, Tufts University School of Medicine and Tufts Medical Center, Boston, Massachusetts,02111 U.S.A. goxenkrug@tuftsmedicalcenter.org

56

Gregory F. Oxenkrug

use of selective 5-HT uptake inhibitors as antidepressant drugs contributed to almost 40 years of continued interest in the “serotonin hypothesis.”

Another important consequence of so-called “serotonin hypothesis” was stimulation of research of biological and neurotropic activity of KYN and its derivatives (summarily called “kynurenines”) (3-7) and of factors regulating KYN pathway of TRY metabolism (8).

This review offers analysis of the current status of the serotonin hypothesis with special consideration of the discovery of indoleamine 2,3-dioxygenase (IDO) (9), the other rate-limiting enzyme of TRY - KYN pathway, different from TDO in substrate specificity, localization and regulatory mechanisms.

Tryptophan Metabolism In humans TRY is an essential amino acid with two non-protein metabolic pathways: methoxyindoles and KYN (Fig. 2).

Figure 2. Methoxyindoles and kynurenine pathways of tryptophan metabolism Abbreviations: TRY – tryptophan; IDO – indoleamine 2,3-dioxygenase; TDO- tryptophan 2,3-dioxygenase; TPH – tryptophan hydroxylase; 5-HT – serotonin; NAT – N-acetyltransferase; NAS – N-acetylserotonin; NAD nicotinamide adenine dinucleotide TRY

IDO/ TDO

TPH

KYNURENINE

5-HT

NAT

NAS

melatonin

KYN 3-hydroxylase aminotransferase 3-hydroxykynurenine

kynurenic acid

kynureninase 3-hydroxyanthranilic acid picolinic acid

quinolinic acid

NAD

The Methoxyindoles Pathway

Availability of TRY as a substrate is one of the ratelimiting factors of methoxyindoles pathway of 5-HT biosynthesis since less than 5% of TRY metabolized along this pathway (10). The other rate-limiting step is hydroxylation of TRY catalyzed by TRY-hydroxylase with the formation of 5-hydroxytryptophan. The subsequent decarboxylation results in the formation of 5-HT, a substrate for melatonin synthesis. The rate-limiting step of melatonin synthesis is 5-HT-N-acetylation resulting in the formation of N-acetyl-serotonin (NAS) with subsequent O-methylation into 5-methoxy-Nacetyltryptamine (melatonin) (11) (Fig. 2).

5-HT (not competitively) and NAS and melatonin (competitively) inhibit liver TDO (12). Deficient production of 5-HT, NAS and melatonin contribute to depressed mood (13), and disturbances of sleep (14) and circadian rhythms (15).

The Kynurenine Pathway

About 95% of TRY is metabolized via the KYN pathway (10, 16). There are two steps of the TRY–KYN pathway: a) formation of KYN from TRY, and b) post-KYN metabolism via two routes competing for KYN as their initial substrate. a. Tryptophan conversion into Kynurenine Unlike the methoxyindoles pathway that does not affect

the indole ring of TRY, the KYN pathway begins by the cleavage of the indole ring of TRY which results in the formation of N-formylkynurenine followed by kynurenine in an ensuing step (10). The rate-limiting enzymes of KYN formation from TRY are IDO (9) in astrocytes, microglia, microvascular endothelial cells and macrophages and TDO in liver, kidney and brain (10).

KYN inhibits TRY transport via the blood-brain barrier (4), stimulates IDO activity (10), and exerts anxiogenic activity in animal models of anxiety (4). b. Post-Kynurenine metabolism Kynurenine is further metabolized along the two distinct routes competing for KYN as a substrate: KYN–kynurenic acid (KYNA) pathway, and KYN–nicotinamide adenine dinucleotide (NAD) pathway. b1. The KYN –KYNA pathway The KYN-KYNA pathway is regulated by KYN aminotransferases, the major biosynthetic enzymes of KYNA formation in the brain (17).

KYNA, the only known endogenous antagonist to N-methyl-D-aspartate (NMDA) receptors, might, similarly to the exogenous NMDA antagonists, ketamine and MK-801, exert antidepressant (18) and psychotomimetic (19) effects. KYNA may contribute to cross 57

TRYPTOPHAN–KYNURENINE METABOLISM AS A COMMON MEDIATOR

talk between the melatonin and kynurenine pathways by inhibiting 5-HT-N-acetylation, the rate-limiting step of melatonin biosynthesis (20).

KYNA has higher affinity to alpha-7-nicotinic acetylcholine than to NMDA receptors, and as such might contribute to cognitive impairment observed in depression, schizophrenia, dementia, and Down’s and Crohn’s syndromes (21, 22). b2. The KYN-NAD pathway The KYN–NAD pathway produces NMDA agonists (quinolinic and picolinic acids) and free radical generators (3-hydroxykynurenine and 3-hydroxyanthranilic acid) (16). Increased formation of NMDA agonists might result in hyperglutamatergic status suggested to be associated with depression (21).

Quinolinic and picolinic acids exerted an anxiogenic effect in experimental models (4). Quinolinic and picolinic acids stimulate inducible nitric oxide synthase (iNOS) and together with 3-hydroxykynurenine and 3-hydroxyanthranilic acids might increase lipid peroxidation, and trigger arachidonic acid cascade resulting in the increased production of inflammatory factors: prostaglandines, via activation of cycloxygenase (COX) and leucotrienes, via activation of arachidonate 5-lipoxygenase (5-LO) (16, 23, 24). COX-2 is of particular interest since its inhibitors blocked KYNA production (25) and exerted antidepressant and antipsychotics effects (21) while 5-LO was suggested as a link between depression and atherosclerosis (26-28) (Fig. 3).

Figure 3. Kynurenines and psychiatric and vascular complications Abbreviations: TRY – tryptophan; IDO – indoleamine 2,3-dioxygenase; TDO – tryptophan 2,3-dioxygenase; BBB – blood brain barrier; KYN - kynurenine, 3HKYN 3-hydroxyKYN; KYNA – kynurenic acid; QUIN – quinolinic acid; iNOS – inducible nitric oxide synthase; NO - nitric oxide; PLA – phospholipase; AA – arachidonic acid; COX - cycloxygenase; 5-LO - arachidonate 5-lipoxygenase; PGE – prostaglandines.

TRY

IDO/ TDO

3-HKYN, QUIN (anxiety)

BBB serotonin (depression)

58

KYN

NAS

KYNA (psychosis, cognitive impairment)

iNOS NO melatonin (insomnia, circadian dysrhythmia)

PGE PLA

COX-2 AA 5-LO

leucotrienes (atherosclerosis)

Regulation of rate-limiting enzymes of KYN pathway.

Regulation of TDO

a. Substrate activation. TDO is activated by its substrate (TRY) (10). Because KYN competes for cerebral transport and cellular uptake of TRY, and because of substrate inhibition on TRY hydroxylase, the rate-limiting enzyme of 5-HT biosynthesis, excessive TRY doses may decrease 5-HT production (29). b. Hormonal activation. Cortisol activates TDO and increased KYN production (30). Literature data regarding the effect of estrogens and testosterone on TDO are controversial: both adrenalectomy and ovariectomy reduced TDO activity in homogenates of liver from mature rats. However, administration of estrogens and testosterone had no effect on TDO (31, 32).

Hormonal activation of TDO and consequent shift of TRY metabolism from 5-HT to KYN formation was suggested as an etiological factor in depression (1) (Fig.1).

Regulation of IDO

a. Cytokines and IDO a1. IFN-gamma. Pro-inflammatory cytokines (including, most notably, interferons) transcriptionally induce IDO in a variety of immune cells (e.g., monocytederived macrophages and microglia) (33). IFNG is the strongest known inducer of IDO (34). Therefore, the shift of TRY metabolism from 5-HT to KYN formation Figure 4. Cytokines and regulation of IDO Abbreviations: IFNG – interferone-gamma; IFN-alpha – interferone-alpha; TNF-alpha – tumor necrosis factor-alpha; IDO – indoleamine 2, 3-oxygenase IFNG TNF-alpha

TRYPTOPHAN

IFN-alpha

IDO

KYNURENINE

might be caused not only by TDO activation by cortisol but by IFNG-induced IDO activation as well. The shift of TRY towards formation of kynurenines might be further augmented by IFNG-induced stimulation of the enzymes of KYN – NAD route: 3-hydroxylase and kynureninase (35). a2. IFN-alpha. Systemically administrated IFN-alpha

Gregory F. Oxenkrug

passes the blood-brain barrier and reaches effective concentrations acting on microglial cells as well as macrophage receptors (36). IFN-alpha has much weaker direct IDO stimulating effect than IFNG but might increase IDO activity by stimulating the production of IFNG and TNFalpha (37) (Fig. 4). Administration of IFN-alpha to patients with hepatitis C is associated with depression attributed to increased formation of kynurenines (38-40). a3. TNF-alpha. TNF-alpha stimulates IDO activity and enhances (up to 300%) IFNG-induced IDO expression (41). The induction of IDO by the bacterial endotoxin lipopolysaccharide was IFNG-independent and might be mediated by toll-like receptors (42). a4. Other proinflammatory molecules such as IL-1, IL-12, Il-18, PGE2 synergistically with IFNG induce IDO activity (43, 44).

Experimental and clinical data demonstrated that IFNG and TNF-alpha trigger depression (and depressivelike symptoms) via stimulation of IDO and consequent increase of kynurenines formation from TRY (39, 45).

Cytokine gene polymorphism and IDO Cytokine genes are polymorphic and certain SNPs located within coding/regulatory regions affect the overall expression and secretion of cytokines (46).

IFNG production is encoded by polymorphic IFNG (+874) gene with high (T) and low (A) producer alleles (46). Mean concentration of IFNG cytokine released by stimulated peripheral blood mononuclear cells was higher in healthy carriers of T than in carriers A allele (47). High producer T allele was associated with increased IDO activity (i.e., elevated plasma kynurenine levels and kynurenine/tryptophan ratios) in healthy females (48).

These results suggest that IFNG genotype influences TRY catabolism via regulation of IDO activity.

TNF-alpha production is encoded by the TNF-alpha (-308 A/G) polymorphic gene. Since TNF-alpha stimulates IDO and potentiates IFNG-induced stimulation of IDO (see above), a (TNF-alpha (–308) high producer (A) allele might strength the association between IFNG (+874) high producer (T) and IDO up-regulation (16).

Increased frequency of the TNF-alpha -308A allele (high producer) was reported in Korean subjects with major depression (49), in Korean bipolar I patients (50), and in Polish subjects with bipolar affective disorder and positive family history (51). It suggested that (–308) TNF-alpha gene polymorphism might be involved in genetic susceptibility to mood disorders.

IDO–TDO interaction

a. Hormonal activation of IDO. Although IDO is mainly induced by cytokines, experimental data suggested that expression of IFNG gene may be subject to direct hormonal control since receptors of prolactin and IFNG share their structure and signal transduction pathway. Prolactin by itself has little or no effect on IDO but potentiates INFG-induced IDO activation in CD14positive cells (52). 17beta-estradiol increased the activity of the IFNG promoter in lymphoid cells (53).

Hydrocortisone and dexamethasone induced IDO in human astrocytoma cells and in native human astrocytes (54). b. Cytokines and the HPA axis. Proinflammatory cytokines may cause hypothalamic-pituitary-axis (HPA) hyperactivity (that is frequently observed in depression) by disturbing the negative feedback inhibition of circulating corticosteroids on the HPA axis (55, 56). c. Aging as a merging point of IDO-TDO interaction.

Aging is characterized by elevated cortisol production due to disinhibition of the HPA axis (57-59), and by increased IFNG and TNF-alpha production (60). It is noteworthy that high producer allele (T) of the IFNG +874 gene (61) and high IDO activity predicted high lethality in elderly subjects (62). In the same vein, Drosophila Melanogaster mutants with impaired KYN production have longer life span than wild type flies (63). These results suggest that activation of both IDO and TDO might contribute to high risk of depression in the elderly.

Hypothesis The original 1969 hypothesis of up-regulation of the KYN pathway of TRY metabolism as an etiological factor in depression suggested that “in depression the activity of liver TRY-pyrrolase (TDO) is stimulated by raised blood corticosteroids levels” that resulted in 5-HT deficiency due to the shift of TRY metabolism from 5-HT to KYN formation (1). The discovery of neurotropic activity of kynurenines (37) emphasized the increased formation of kynurenines as etiological factor in depression (in addition to 5-HT deficiency) (39). IDO, the other enzyme catalyzing TRY conversion into KYN, is transcriptionally activated by pro-inflammatory cytokines (mainly, IFNG and TNF-alpha). The polymorphisms of genes impacting the production of pro-inflammatory cytokines provides 59

TRYPTOPHAN–KYNURENINE METABOLISM AS A COMMON MEDIATOR

Figure 5. Genetic and environmental impacts on tryptophan metabolism Abbreviations: NAS – N-acetylserotonin; IFNG – interferone; TNF – tumor necrosis factor; IDO – indoleamine 2,3-dioxygenase; TDO – tryptophan 2,3-dioxygenase.

Serotonin

NAS

Melatonin

L-TRYPTOPHAN

Genes (IFNG, TNF-alpha)

IDO

TDO

Life stresses (cortisol, prolactin)

KYNURENINE

further insight in the etiological role of upregulated TRY – KYN metabolism in depression.

This review suggests that genetic and/or environmental (life stresses) factors trigger depression by upregulation of TRY-KYN metabolism. Effect of genetic factors (such as high producer alleles of pro-inflammatory genes) is mediated by cytokine-induced up-regulation of IDO. Combination of high producer alleles of IFNG (+874) and TNF-alpha (-308) genes might result in high production of these cytokines, and trigger the “super-induction” of IDO. The effect of life stressors might be mediated by hormonal activation of TDO. Cytokine–induced stimulation of cortisol production and augmentation of IFNG-induced activation of IDO by stress hormones suggest that the TRY-KYN pathway might be the converging point of gene-environmental interaction (e.g., like in aging) (Fig. 5).

Limitations Some major limitations should be mentioned with the hope to stimulate further evaluation of the proposed hypothesis.

The present hypothesis predicts higher frequency of carriers of high producer alleles of pro-inflammatory genes in subjects with mood disorders. While some studies of TNF-alpha genotypes support this suggestion (49-51), no publications regarding IFNG (or other pro-inflammatory cytokines) were found in the available sources.

The present hypothesis is based on the assumption that carriers of high producer alleles of cytokine genes have higher production of cytokines than carriers of low promoter alleles. These relationships were reported for healthy volunteers but were not studied in depressed patients. Similarly, the association between high producer alleles for the IFNG (+874) gene and high IDO 60

activity was observed in healthy volunteers but no studies done in depressed patients.

TRY–KYN metabolism as a new target for prevention and treatment of MDD and psychiatric complications of IFNalpha treatments Genotype assessment might help identify subjects-atrisk of developing depression in response to environmental stressors and/or to IFN-alpha therapy of hepatitis C, cancer, amyotrophic lateral sclerosis and multiple sclerosis.

Potential pharmacological interventions in identified subjects may include: a) inhibition of cytokine production by antibodies to TNF-alpha (e.g., etanercept, infiximab) and IFNG (64), and/or more careful selection of antidepressants.

While both selective 5-HT uptake inhibitor, fluoxetine (65) and the dopamine enhancer, wellbutrin (66) inhibit cytokine production, the latter might be of advantage considering the impaired 5-HT synthesis as a result of IDO activation. b) inhibition of IDO activity by MAO inhibitors (6769), minocycline (70) and 1-methyl-L-TRY (71). d) administration of methoxyindoles that might modulate the TRY-KYN pathway due to their inhibitory effect on cortisol (72) and proinflammatory cytokines (73-75) production. Methoxyindoles (melatonin, in particular) might attenuate excitatory, glutamate-mediated responses triggered by KYN pathway metabolites (76, 77).

Acknowledgement This paper is supported by NIH R21MH083225.

References 1. Lapin IP, Oxenkrug GF. Intensification of the central serotoninergic processes as a possible determinant of the thymoleptic effect. Lancet 1969; 1: 32-39.

2. Rubin RT, Mandell AJ, Crandall PH. Corticosteroid responses to limbic stimulation in man: Localization of stimulus sites. Science 1966;153:767-768.

3. Lapin IP. Convulsions and tremor in immature rats after intraperitoneal injection of kynurenine and its metabolites. Pharmacol Res Comm 1978;10: 81–84.

4. Lapin IP. Neurokynurenines (NEKY) as common neurochemical links of stress and anxiety. Adv Exp Med Biol 2003; 527:121-125.

Gregory F. Oxenkrug

5. Oxenkrug G F, Lapin IP. Kynurenine pathway of the metabolism of tryptophan and its possible neuropharmacologic role. In: Yakovelev V, editor. Chemistry and pharmacology of indole compounds. Kishinev: Stinza, 1975: pp. 5-18.

6. Lapin IP. Antagonism of kynurenic acid to anxiogens in mice. Life Sci 1998; 63:PL231-236.

7. Lapin IP. Kynurenines and anxiety. Adv Exp Med Biol 1996; 398:191-194.

8. Schwarcz R, Pellicciari R. Manipulation of brain kynurenines: glial targets, neuronal effects, and clinical opportunities. J Pharmacol Exp Ther 2002; 303:1-10.

9. Hayaishi O. Properties and function of indoleamine 2,3-dioxygenase. J Biochem (Tokyo) 1976;79:13P-21P. 10. GĂĄl EM, Sherman AD. L-kynurenine: its synthesis and possible regulatory function in brain. Neurochem Res 1980; 5:223-239.

11. Kopin IJ, Pare CB, Axelrod J, H Weissbach. The fate of melatonin in animals. J Biol Chem 1961; 236:3072-3075.

12. Walsh HA, Daya S. Inhibition of hepatic tryptophan-2,3dioxygenase: Superior potency of melatonin over serotonin. J Pineal Res 1997; 23:20-23.

13. Lewy AJ, Wehr TA, Goodwin FK, et al. Light suppresses melatonin secretion in humans. Science 1980; 210:1267-1269.

14. Brzezinski A, Vangel MG, Wurtman RJ, Norrie G, Zhdanova I, et al. Effects of exogenous melatonin on sleep: A meta-analysis. Sleep Med Rev 2005; 9:41-50.

15. Oxenkrug GF, Requintina PJ. Melatonin and jet lag syndrome: Experimental model and clinical implications.

CNS Spectrums 2003; 8:139-148.

16. Oxenkrug GF. Genetic and hormonal regulation of the kynurenine pathway of tryptophan metabolism: new target for clinical intervention in vascular dementia, depression and aging. Ann NY Acad Sci 2007; 1122: 35-49.

17. Guidetti P, Amori L, Sapko MT, et al. Mitochondrial aspartate aminotransferase: A third kynurenate-producing enzyme in the mammalian brain. J Neurochem 2007;102:103-111.

18. Zarate CA Jr, Quiroz JA, Singh JB, et al. An open-label trial of the glutamate-modulating agent riluzole in combination with lithium for the treatment of bipolar depression.

Biol Psychiatry 2005;57:430-432.

19. Krystal JH, Karper LP, Seibyl, JP, et al. Subanesthetic effects of the noncompetitive NMDA antagonist, ketamine, in humans. Psychotomimetic, perceptual, cognitive, and neuroendocrine responses. Arch Gen Psychiatry 1994;51:199-214.

20. Zawilska JB, Rosiak J, Senderecka M, Nowak JZ. Suppressive effect of NMDA receptor antagonist MK-801 on nocturnal serotonin N-acetyltransferase activity in the rat pineal gland. Pol J Pharmacol 1997; 49: 479-483.

21. MĂźller N, Schwarz M. A psychoneuroimmunological perspective to Emil Kraepelins dichotomy: Schizophrenia and major depression as inflammatory CNS disorders. Eur Arch Psychiatry Clin Neurosci 2008; 258:97-106.

22. Oxenkrug GF. Tryptophan metabolism as a new target for the treatment of schizophrenia. US Psychiatry Review 2007 (Touch Briefings): pp. 38-39.

23. Oxenkrug GF. Antioxidant effects of N-acetylserotonin: Possible mechanisms and clinical implications. Ann NY Acad Sci 2005;1053: 334-347.

24. Melillo G, Cox DW, Biragyn A, Sheffler LA. Regulation of nitric-oxide synthase mRNA expression by interferon-gamma and picolinic acid. J Biol Chem 1994; 269: 8128-8133.

25. Schwieler L, Erhardt S, Nilsson L, et al. Effects of COX-1 and COX-2 inhibitors on the firing of rat midbrain dopaminergic neurons - possible involvement of endogenous kynurenic acid. Synapse 2006 ; 59:290-298.

26. Manev G, Manev R. 5-lipoxygenase as a possible biological link between depressive symptoms and atherosclerosis.

Arch Gen Psychiatry 2007; 64:1333.

27. Oxenkrug GF. Metabolic syndrome, age-associated neuroendocrine disorders and dysregulation of tryptophan kynurenine pathway metabolism. Ann NY Acad Sci; 2009 (in press).

28. Oxenkrug GF. Antidepressant effect of N-acetylserotonin in relation to aging, hypertension and cancer. In: Berstein L, editor. Hormones, age and cancer. St. Petersburg: Nauka, 2005: pp. 140-158.

29. Forrest CM, Mackay GM, Stoy N, Egerton M, Christofides J, Stone TW, Darlington LG. Tryptophan loading induces oxidative stress. Free Radic Res 2004;38:1167-1171.

30. Rubin RT. Adrenal cortical activity changes in manic-depressive illness. Influence on intermediary metabolism of tryptophan. Arch Gen Psychiatry 1967;17:671-679.

31. Bender DA, Laing AE, Vale JA, et al. The effects of oestrogen administration on tryptophan metabolism in rats and in menopausal women receiving hormone replacement therapy. Biochem Pharmacol 1983; 32: 843-848.

32. Shibata K, Toda S. Effects of sex hormones on the metabolism of tryptophan to niacin and to serotonin in male rats.

Biosci Biotechnol Biochem 1997; 61:1200-1202.

33. Taylor MW, Feng GS. Relationship between interferongamma, indoleamine 2,3-dioxygenase, and tryptophan catabolism. FASEB J 1991; 5:2516-2522.

34. Widner B, Ledochowski M, Fuchs D. Interferon-gammainduced tryptophan degradation: Neuropsychiatric and immunological consequences. Curr Drug Metab 2000;1:193-204.

61

TRYPTOPHAN–KYNURENINE METABOLISM AS A COMMON MEDIATOR

35. Alberati-Giani D, Ricciardi-Castagnoli P, Köhler C, Cesura AM. Regulation of the kynurenine metabolic pathway by interferon-gamma in murine cloned macrophages and microglial cells. J Neurochem 1996; 66:996-1004.

48. Raitala A., Pertovaara M, Karjalainen J, et al. Association of interferon-gamma+874(T/A) single nucleotide polymorphism with the rate of tryptophan catabolism in healthy individuals. Scand J Immunol 2005; 61: 387-39046.

36. Sweeten TL, Ferris M, McDougle CJ, et al. Induction of indoleamine 2,3-dioxygenase in vivo by IFN-con1. J Interferon Cytokine Res 2001 ;21:631-633.

49. Jun TY, Pae CU, Hoon-Han, Chae JH, Bahk WM, Kim KS, Serretti A. Possible association between -G308A tumour necrosis factor-alpha gene polymorphism and major depressive disorder in the Korean population. Psychiatr Genet 2003;13:179-181.

37. Taylor JL, Grossberg SE. The effects of interferon-alpha on the production and action of other cytokines. Semin Oncol 1998;25:23-29.

38. Capuron L, Miller AH. Cytokines and psychopathology: Lessons from interferon-alpha. Biol Psychiatry 2004; 56: 819-824.

39. Wichers MC, Koek GH, Robaeys G, Verkerk R, Scharpé S, Maes M. IDO and interferon-alpha-induced depressive symptoms: A shift in hypothesis from tryptophan depletion to neurotoxicity. Mol Psychiatry 2005;10:538-544.

40. Bonaccorso S, Meltzer HY, Maes M. Psychological and behavioral effects of interferons. Curr Opin Psychiatry 2000; 13:673-677.

41. Robinson CM, Hale PT, Carlin JM. The role of IFN-gamma and TNF-alpha-responsive regulatory elements in the synergistic induction of indoleamine dioxygenase. J Interferon Cytokine Res 2005;25:20-30.

42. Wilson AG, de Vries N, Pociot F, et al. An allelic polymorphism within the human tumor necrosis factor-alpha promoter region is strongly associated with HLA A1, B8, and DR3 alleles. J Exp Med 1993; 177:557-560.

43. Liebau C, Baltzer A W, Schmidt S, et al. Interleukin-12 and interleukin-18 induce indoleamine 2,3-dioxygenase (IDO) activity in human osteosarcoma cell lines independently from interferon-gamma. Anticancer Res 2002; 22: 931-936.

44. Kwidzinski E, Bunse J, Aktas O, et al. Indolamine 2,3-dioxygenase is expressed in the CNS and down-regulates autoimmune inflammation. FASEB J 2005; 19:1347-1349.

45. O’Connor JC, André C, Wang Y, Lawson MA, Szegedi SS, Lestage J, Castanon N, Kelley KW, Dantzer R. Interferongamma and tumor necrosis factor-alpha mediate the upregulation of indoleamine 2,3-dioxygenase and the induction of depressive-like behavior in mice in response to bacillus Calmette-Guerin. J Neurosci 2009;29:4200-4209.

50. Pae CU, Lee KU, Han H, et al. Tumor necrosis factor alpha gene-G308A polymorphism associated with bipolar I disorder in the Korean population. Psychiatry Res 2004;125:65-68.

51. Czerski PM, Rybakowski F, Kapelski P, et al. Association of tumor necrosis factor -308G/A promoter polymorphism with schizophrenia and bipolar affective disorder in a Polish population. Neuropsychobiology 2008; 57:88-94.

52. Kawaguchi R, Ozawa-Kondo M, Ohta-Misaki I, et al.

Prolactin (PRL) up-regulates indoleamine 2,3-dioxygenase (IDO) expression in CD14+ cells. Nihon Rinsho Meneki Gakkai Kaishi 2005; 228:407-412.

53. Fox HS, Bond BL, Parslow TG, et al. Estrogen regulates the IFN-gamma promoter. J Immunol 1991; 146: 4362-4367.

54. Ozaki Y, Edelstein MP, Duch DS. The actions of interferon and antiinflammatory agents of induction of indoleamine 2,3-dioxygenase in human peripheral blood monocytes.

Biochem Biophys Res Commun 1987; 144: 1147-1153.

55. Leonard BE. The HPA and immune axes in stress: The involvement of the serotoninergic system. Eur Psychiatry 2005; 20: S302-S306.

56. Leonard BE, Myint A. The psychoneuroimmunology of depression. Hum Psychopharmacol 2009;24:165-175.

57. Dilman VM, Lapin IP, Oxenkrug GF. Serotonin and aging.

In: Essman W, editor. Serotonin in Health and Disease.

New York, London: Spectrum, 1979; vol. 5: pp. 111-123.

58. Oxenkrug GF, McIntyre I M, Gershon S, et al.

Dexamethasone suppression test: experimental model in rats and effect of age. Biol Psychiat 1984; 19: 413-416.

59. Oxenkrug GF, Pomara N, McIntyre I, et al. Aging and cortisol resistance to suppression by dexamethasone: A positive correlation. Psychiatry Res 1983; 10: 125-130.

46. Pravica V, Perrey C, Stevens A, et al. A single nucleotide polymorphism in the first intron of the human INFG gene: Absolute correlation with a polymorphic CA micro marker of high INFG gene. Hum Immunol 2000; 61: 8333-8366.

60. Rodríguez MI, Escames G, López LC, et al. Chronic melatonin treatment reduces the age-dependent inflammatory process in senescence-accelerated mice. J Pineal Res 2007; 42:272-279.

47. Anuradha B, Rakh SS, Ishaq M, et al. Interferon-gamma Low producer genotype +874 overrepresented in Bacillus Calmette-Guerin nonresponding children. Pediatr Infect Dis J 2008; 27:325-329.

61. Lio D, Scola L, Crivello A, et al. Allele frequencies of +874T-A single nucleotide polymorphism at the first intron of interferon-gamma gene in a group of Italian centenarians. Exp Gerontol 2002, 37: 315-319.

62

Gregory F. Oxenkrug

62. Pertovaara M, Raitala A, Lehtimaki T, et al. Indoleamine 2,3-dioxygenase activity in nonagenarians is markedly increased and predicts mortality. Mech Ageing Dev 2006; 127:497-499.

63. Oxenkrug GF. The extended life span of Drosophila melanogaster eye-color (white and vermilion) mutants with impaired formation of kynurenine. J Neural Transm 2010; 117: 23 -26.

64. Skurkovich B, Skurkovich S. Inhibition of IFN-gamma as a method of treatment of various autoimmune diseases, including skin diseases. Ernst Schering Res Found Workshop 2006;56:1-27.

65. Kenis G, Maes M. Effects of antidepressants on the production of cytokines. Int J Neuropsychopharmacol 2002; 5:401-412.

66. Brustolim D, Ribeiro-dos-Santos R, Kast RE, et al. A new chapter opens in anti-inflammatory treatments: The antidepressant bupropion lowers production of tumor necrosis factor-alpha and interferon-gamma in mice. Int Immunopharmacol 2006; 6:903-907.

67. Samsonova ML, Oxenkrug GF. Inhibition of substrate induction of TRY-pyrrolase in liver and increase in the content in 5-HT in brain under action of inhibitors of MAO. Vopr Med Khim 1972; 17: 198-201.

68. Oxenkrug GF. The acute effect of monoamine oxidase inhibitors on serotonin conversion to melatonin. In: Sandler M, Coppen A, Harnett S, editors. 5-hydroxytryptamine in psychiatry: A spectrum of ideas. Oxford, New York, Tokyo: Oxford University, 1991: pp: 98-109.

69. Sono M, Cady SG. Enzyme kinetic and spectroscopic studies of inhibitor and effector interactions with indoleamine 2,3-dioxygenase. 1. Norharman and 4-phenylimidazole binding to the enzyme as inhibitors and heme ligands.

Biochemistry 1989; 28:5392-5399.

70. Ryu J K, Choi H B, McLarnon J G. Combined minocycline plus pyruvate treatment enhances effects of each agent to inhibit inflammation, oxidative damage, and neuronal loss in an excitotoxic animal model of Huntingtonâ&#x20AC;&#x2122;s disease.

Neuroscience 2006;141:1835-1848.

71. Cady SG, Sono M. 1-Methyl-DL-tryptophan, beta-(3benzofuranyl)-DL-alanine (the oxygen analog of tryptophan), and beta-[3-benzo(b)thienyl]-DL-alanine (the sulfur analog of tryptophan) are competitive inhibitors for indoleamine 2,3-dioxygenase. Arch Biochem Biophys 1991;291:326-333.

72. Oxenkrug GF, McIntyre IM, Gershon S. Effects of pinealectomy and aging on the serum corticosterone circadian rhythm in rats. J Pineal Res 1984; 1:181-185.

73. Perianayagam MC, Oxenkrug GF, Jaber B. Immune modulating effects of melatonin, N-acetyl Serotonin and N-acetyl Dopamine. Ann NY Acad Sci 2005; 1053:386-393.

74. Requintina PJ, Oxenkrug GF. Differential effects of lipopolysaccharide on lipid peroxidation in F344N, SHR rats and BALB/C mice and protection of melatonin and NAS against its toxicity. Ann NY Acad Sci 2003; 993:325-333.

75. Bachurin S, Oxenkrug G, Lermontova N, et al. Nacetylserotonin, melatonin and their derivatives improve cognition and protect against beta-amyloid-induced neurotoxicity. Ann NY Acad Sci 1999; 890:155-166.

76. Lapin IP, Mirzaev SM, Oxenkrug GF, et al. Anticonvulsant activity of melatonin against seizures induced by quinolinate, kainate, glutamate, NMDA and pentylenetetrazole in mice. J Pineal Res 1998; 24: 215-218.

77. Prakhie IV, Oxenkrug GF. The effect of nifedipine, Ca++ antagonist, on activity of MAO inhibitors, N-acetylserotonin and melatonin in the mouse tail suspension test. Intern J Neuropsychopharmacol 1998; 1:35-40.

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Isr J Psychiatry Relat Sci - Vol 47 - No. 1 (2010)

Biochemical and Anatomical Substrates of Depression and Sickness Behavior Thomas C. Hanff, MD, Stephanie J. Furst, and Thomas R. Minor, PhD Departments of Psychology and Neuroscience, University of California, Los Angeles, California, U.S.A.

ABSTRACT This paper reviews recent research on the contribution of the proinflammatory cytokine interleukin-1β (IL1β) and the purine nucleoside adenosine in mediating behavioral depression and related symptoms of conservation-withdrawal in animal models of both major depression and illness. Activation of brain IL1β receptors appears to contribute to conservationwithdrawal symptoms in animals treated with reserpine or lipopolysaccharide, suggesting a common underlying mechanism. Moreover, brain cytokine signaling is capable of recruiting adenosine signaling at adenosine A2A receptors, which directly mediate symptoms of behavioral depression. The adenosine receptors densely populate spiny GABAergic neurons in the striopallidal tract in the striatum and form part of an A2A/D2/mGLU receptor complex. Activation of these A2A receptors functionally uncouples dopamine’s excitatory motivation influence from ongoing behavior, leading to a state of conservation-withdrawal, and antagonism of the ventral medial striatum A2A receptors in reserpinated rats relieves symptoms of behavioral depression.

Introduction Sickness behavior – the lethargy, hypoactivity, decreased libido, anorexia, anhedonia, and increased sleep that accompanies infectious disease (1, 2) was once thought to be a maladaptive consequence of an animal’s immune response, damaging the animal’s ability to successfully interact with its environment. However, two decades of research into the biological mechanism of sickness behavior have modified this view, demonstrating that

sickness behavior is an adaptive central motivational state necessitated by the metabolic constraints for mounting a fever (3).

The characterization of sickness behavior as a motivational state underscores similarities among mood disorders, the reaction to traumatic stress and recuperation from injury (4). Each of these conditions is associated with intense catabolic output (5) and is automatically and unconditionally followed by a compensatory shift to a behavioral state termed conservation-withdrawal (6). The sensory unresponsiveness, cognitive dullness and behavioral depression that characterize this state are adaptive mechanisms for husbanding limited resources and facilitating the recovery of energy homeostasis.

Conservation-withdrawal is an integral component of major depression and related mood disorders (7).

This reaction most closely corresponds to the affect-less, fatigue components of depression, rather than subsuming the entirety of the behavioral, cognitive, emotional and motivational symptoms that comprise the disorder – it also represents the aspects of affective disorders that are most accurately modeled in animals (8). Conservationwithdrawal also is a key component of the after-reaction to physical and psychological stress. Thus symptoms of conservation-withdrawal are seen after a patient leaves the intensive care unit following a serious injury and are often confused with major depression (9). These same symptoms also are the hallmark of the after-reaction to traumatic psychological stress that has been variously termed learned helplessness (10), behavioral depression (11) and the distress syndrome (12), and they comprise a critical component of sickness behavior.

This paper reviews research from our laboratory on conservation-withdrawal reactions in two animal paradigms that have been used to model symptoms of major depression: induction of sickness behavior with endotoxin or interleukin-1β (IL-1β), and systemic injection

Address for Correspondence: Thomas R. Minor, Departments of Psychology and Neuroscience, University of California, Los Angeles, CA 900951563, U.S.A. minor@psych.ucla.edu

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of reserpine. We have examined the potential contribution of two signaling pathways – one involving the purine nucleoside adenosine, and the other involving the proinflammatory cytokine IL-1β – to the induction of conservation-withdrawal in each of these paradigms.

Both of these molecules modulate excitable tissue in the periphery (13, 14). They also produce global organized responses by binding to receptors in the central nervous system to alter neural signaling (15, 16). More important, both of these pathways are capable of producing the symptoms of conservation-withdrawal. A brief review of cytokine and adenosine signaling are presented below.

I. Modulators of Conservation-withdrawal

A. Brain Adenosine Signaling. Adenosine participates in a purine feedback pathway that regulates excitable tissue with respect to available energy.

The purine nucleotide adenosine triphosphate (ATP) and the nucleoside adenosine (ADO) serve as endpoints in intracellular metabolism, with expendable energy represented in the number of high-energy phosphate bonds (17). These molecules also are liberated into extracellular space to convey information concerning intracellular energy state. Adenosine signaling is actively engaged by challenges to metabolic homeostasis (18). The nucleoside exerts very potent inhibition on excitatory transmission in brain as a compensatory reaction to neural energy failure (19).

Adenosine is extruded into extracellular space or hydrolyzed from extracellular nucleotides whenever the rate of adenosine triphosphate (ATP) hydrolysis exceeds the synthesis rate (20). Such an imbalance of the energy supply/demand ratio can result from excessive neural activation or from a shortage in brain glucose or oxygen.

The extracellular nucleoside binds to specific G-protein linked adenosine receptors (A1, A2A, A2B, & A3), which are widely distributed on pre- and post-synaptic membranes and in the microvascular bed (21). Adenosine interacts with a number of cellular effector systems via these receptors to decrease membrane excitability and inhibit transmitter release, thereby decreasing metabolic demand in the target neuron (22). Adenosine also acts at the system level to produce a number of changes that protect neural tissue from the potentially excitotoxic effects of activation in the absence of sufficient energy (23).

Extracellular adenosine concentrations normally are controlled by high- and low-affinity nucleoside uptake transporters (24). Adenosine is rapidly converted to

5'AMP by adenosine kinase once inside the cell, which decreases subsequent extrusion via gradient transport.

The receptor-mediated effects of the ligand also are regulated by a degradation pathway located on glia, which converts extracellular adenosine to inactive inosine and eventually to uric acid (25).

The features of adenosine signaling that are most relevant to conservation-withdrawal, and particularly the aspects of the syndrome related to behavioral depression, involve activation of A2A receptors in the striatum. The striatum (caudate-putamen, nucleus accumbens and olfactory tubercle) is the major component of the basal ganglia in which excitatory glutamatergic inputs from the cortex, thalamus, and limbic areas are integrated with dopaminergic inputs from the mesencephalon.

These processes converge primarily on medium-sized spiny (GABAergic) neurons that project either to the substantia nigra (via the direct or strionigral/entopeduncular paths) or to the globus pallidus (via the indirect or striopallidal path) (26). The striatum plays a critical role in integrating sensory, emotional, motivational and motor components of ongoing action (27). As such, the striatum is a plausible center for the uncoupling of motivation and action during behavioral depression.

Moreover, activation of striopallidal A2A receptors is precisely the type of molecular mechanisms that is anticipated by the concept of conservation-withdrawal.

Adenosine modulates dopaminergic functions in the dorsal and ventral striatum where the nigrostriatal, mesostriatal, and mesolimbic neuronal pathways terminate. Strong evidence now implies the existence of an adenosine A2A/D2 heteromeric complex coupled in an antagonistic relationship. Hillion et al. (28) showed that when cells stably transfected with D2 receptors were transiently cotransfected with a tagged A2A receptor, they formed receptor complexes in the absence of exogenous agonists for either receptor. Not only does binding of the A2A receptor in the heteromeric complex result in conformational changes in the D2 receptor, but it also decreases D2 activated coupling to its Gi-protein (29). Recent evidence suggests that A 2A receptors modulate glutamatergic afferents to the region via an antagonistic coupling to dopamine D2 receptors (30), and a synergistic coupling to metabotropic glutamate (mGLU5) receptors (31). The functional consequences of this arrangement are that activation of D2 receptors augments ongoing action. By contrast, activation of A2A or mGLU5 receptors antagonizes dopamine’s effect on ongoing action to produce behavioral depression.

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B. Bidirectional Immune-to-Brain Signaling. Systemic administration of lipopolysaccharide (LPS), the active fragment of endotoxin from gram-negative bacteria, induces the synthesis of proinflammatory cytokines in peripheral macrophages – interleukin-1β (IL-1β), IL-6, and tumor necrosis factor (TNFα) (32). Kupffer cells in the liver also express IL-1β as a consequence of LPS administration and may serve as the primary immuneto-brain communication pathway. This signal is transferred via the vagal nerve complex to the brain nucleus tractus solitarius (NTS) where IL-1β is then expressed.

The cytokine also is expressed relatively quickly thereafter in a variety of other brain nuclei, particularly in the hypothalamus (33).

IL-1β binds to specific receptors distributed throughout the brain to induce sickness behavior − lethargy, hypoactivity, decreased libido, anorexia, anhedonia and increased sleep (32). This dramatic shift in ongoing activity, along with the induction of fever, is assumed to be a highly adaptive strategy to fight infection.

Symptoms of sickness behavior and major depression overlap considerably (33).

Plasma cytokine concentrations are elevated in depressed patients and normalize with electroconvulvsive therapy and a return of normal affect (34). In animal models, systemic administration of endotoxin not only increases brain concentrations of IL-1β, but also produces swim deficits (35), as well as other experimental indexes of depression (36). These LPS-induced ailments are reversed by chronic (but not acute) treatment with tricyclic antidepressants (36).

Important for the present purpose is the recent finding that LPS-induced swim deficits is reversed by systemic administration of an A2A receptor antagonist (37).

These data support an important interaction between purine (adenosine) and cytokine (IL-1β) signaling in one model of behavioral depression and suggest that the interaction may occur in the central nervous system as well as in the periphery. We provide additional support for this hypothesis in the following review.

II. Reserpine-Induced Depression.

We have conducted a considerable number of studies on the ability of reserpine to induce behavioral depression or conservation-withdrawal in rats (38, 39). Unlike the data from the learned helplessness paradigm, the reserpine data provide very clear support for a role of both adenosine signaling and cytokine signaling in conservation-withdrawal.

66

Reserpine was introduced in the United States in the early 1950s as a treatment for hypertension (40).

The extract reduces both cardiac output and peripheral vascular resistance by depleting stores of biogenic amines in the central and autonomic nervous systems.

Reserpine binds irreversibly to storage vesicles in monoaminergic neurons (41). The vesicles become “leaky,” resulting in seepage of transmitter into the cytoplasm, where it is either destroyed by intraneuronal monoamine oxidase or diffuses into the synaptic cleft. The end result is that little or no active transmitter is released at the synapse following depolarization. Recovery from the effects of reserpine requires synthesis of new storage vesicles, which can take several days to accomplish after discontinuing drug treatment (42).

The historic significance of reserpine is more related to unwanted side-effects than its efficacy as an antihypertensive or tranquilizing agent. Unfortunately, a significant portion of the population undergoing reserpine treatment for hypertension developed severe symptoms of major depression. These inconsistencies led us to revisit the animal literature on reserpine to ask whether behavioral depression is due to depletion of brain monoamines or some secondary consequence of drug treatment. Below we review the major findings from a lengthy series of experiments that implicate brain adenosine A2A and IL-1β signaling in reserpineinduced depression in rats.

A. IL-1β Signaling. One of the main problems in making a convincing case for a direct role of the biogenic amines in reserpine-induced depression is that the time course for their depletion does not fit the time course for behavioral impairment. For instance, Bean et al. (43) reported that dopamine (DA) depletion in striatum, n. accumbens, and frontal cortex occur rapidly and reach a maximum about 6 hours after an ip injection of 6 mg/kg of reserpine. DA levels remained at floor levels until the 18-hour point and then increases thereafter, such that DA baselines recover within 48 hours.

As shown below, a behavioral deficit is evident before maximum DA depletion occurs and persists after DA levels recover.

We determined the time course for reserpine’s effect by injecting groups of rats with a single, 6 mg/kg, ip dose of reserpine or DMSO vehicle. Rats were tested in a forced swim task 1, 24, 48, 72 or 168 hours after drug treatment. Reserpine, in vehicle controls, resulted in large deficit in swim performance with close to a twofold increase in floating times.

Thomas C. Hanff ET AL.

Peroxide radicals are produced as a byproduct of the degradation of biogenic amines by monamine oxidase (44). The concentration of these radicals may be sufficient to damage tissue when monoamines are rapidly depleted with a large reserpine dose. Activation of brain IL-1β receptors in the hypothalamus and elsewhere would produce the symptoms of sickness behavior and conservation-withdrawal. Because behavioral depression is a major component of these reactions, IL-1β is a plausible mediator of swim deficits following reserpine treatment.

We tested this possibility by stereotaxically implanting groups of rats with guide cannulae in the right lateral ventricle. Groups either received an ip injection of 6 mg/kg of reserpine or injected with DMSO vehicle.

DMSO-treated groups received a 6 µg infusion of the IL-1 receptor antagonist (IL-1ra). All groups were tested for swim performance 15 minutes later.

Reserpine-treated rats showed a large swim deficit relative to the DMSO control 1 hour and also 24 hours after the injection. Icv injection of the IL-1ra had no untoward effect on swim performance and also failed to improve swim performance in reserpine-treated rats. We then assessed IL-1β potential contribution at 48 hours post-injection.

It seemed possible that IL-1β is induced at longer post-injection times to mediate swim deficits following reserpine treatment.

To test this possibility rats were implanted with a guide cannula in the right lateral ventricle. Groups received an ip injection of DMSO vehicle or an ip injection of 6 mg/kg reserpine. All rats were tested for swim performance 48 hours later. One reserpine group received icv infusion of saline vehicle and one reserpine group received icv infusion of the IL-1ra. Results revealed reserpine treatment produced a large increase in floating time 48 hours later. These data provide clear evidence for a long-term deficit that is mediated by IL-1β. Thus, reserpine-induced depression consists of multiple deficits, depending on whether or not a proinflammatory cytokine is induced.

If the above is correct, temporal variations in brain IL-1β should parallel the behavioral results for the IL-1ra. Rats were injected with 6 mg/kg of reserpine or DMSO vehicle. Sacrifice occurred at 0, 1, 24, 48, 72, or 168 hours post drug treatment. Brains were dissected into hypothalamus and hippocampus because these regions are implicated in sickness behaviors.

Reserpine produced a large rise in the concentration of IL-1β in the hypothalamus, and to a lesser degree in

the hippocampus, 48 and 72 hours after the injection, but not at earlier times. IL-1β concentrations returned to baseline within 168 hours of reserpine treatment.

These data suggest that there are two temporally distinct components to reserpine-induced depression.

An early component is evident 1 hour after reserpine treatment and persists for 24 hours. This deficit is not occasioned by a rise in brain IL-1β concentrations and is not reversed by the IL-1ra.

Some other mechanism must mediate this initial component of reserpine-induced depression. A second, late component is evident 48 hours after reserpine treatment, persists for at least 72 hours, and recovers within 168 hours. Brain IL-1β concentrations rise substantially at these times, particularly in the hypothalamus (45).

Moreover, the IL-1ra completely reversed swim deficits at the 48 and 72-hours points. Overall, these data indicate that a rise in brain IL-1β is a sufficient, but not a necessary condition for behavioral depression or conservation-withdrawal in this paradigm.

B. Adenosine A2A Signaling. One condition under which adenosine exerts potent compensatory inhibition is during excessive neural activation (46). Large amounts of monoamine transmitter are likely to diffuse into the synaptic cleft with the destruction of storage vesicles upon initial reserpine treatment. The resulting neuronal excitation might be sufficient to compromise metabolic homeostasis and provoke adenosine-mediated inhibition as a compensatory mechanism. If so, then the early component of the reserpine swim deficit, in particular, is likely to involve adenosine signaling. A potential contribution of this mechanism to the latecomponent deficit is less clear, but certainly possible.

Thus, the present experiments assessed the ability of nonselective (A1/A2), or highly selective A1, A2, A2A, or A2B, adenosine receptor antagonists to reverse deficits in forced swim performance occurring 1 or 48 hours after an injection of 6 mg/kg of reserpine.

Swim deficits 1 and 48 hours after reserpine treatment were reversed by the nonselective adenosine receptor antagonist caffeine and by the moderately selective A2 antagonist 3,7-Dimethyl-1-propargylxanthine (DMPX) in a dose-dependent manner. However, no benefit was afforded by any dose of an A1 antagonist 8-Phenyltheophylline (8-PT), or the A 2B antagonist alloxazine (AX) at either time point.

Data displayed evidence that swim deficits 1 hour after reserpine treatment are mediated at adenosine A2A receptors. Groups of rats received an ip injection 67

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of 6 mg/kg reserpine or DMSO vehicle. Forty-five minutes later, one DMSO-treated group was injected with 1.0 mg/kg of the selective A2A antagonist CSC. Reserpine-treated groups received an ip injection of 0, 0.01, 0.1, or 1.0 mg/kg of CSC. All rats were tested for swim performance 15 minutes later. As evident in the figure, CSC alone had no effect on swim performance relative to the DMSO control. Pretreatment with reserpine produced a large swim deficit 1 hour later and this effect was completely reversed by CSC in a dosedependent manner.

The A2A antagonist is equally effective at reversing the late component of the reserpine-induced deficit, whereas other types of adenosine receptor antagonists are not. The moderately selective A2 antagonist DMPX reversed swim deficits 48 hours after reserpine treatment, however, no benefit was afforded by any dose of the A1 antagonist 8-PT, or the A2B antagonist AX. The beneficial effects of DMPX were due to its action at A2A receptors, at a dose of 1.0 mg/kg of CSC, which completely reversed swim deficits 48 hours after reserpine treatment.

The evidence linking behavioral depression in several paradigms to adenosine signaling at A2A receptors provides significant insight as to where this response is organized in the brain. Given the limited distribution of A2A receptors in brain, the critical role of A2A receptor activation in behavioral depression as reviewed above, and the role of the striatum in linking motivation and action, this brain region would seem to be an excellent candidate to mediate conservation-withdrawal reactions following reserpine treatment.

Figure 1. Effects of bilateral microinfusion of CSC on reserpine-induced swim deficits. Rats received an ip injection of reserpine or DMSO vehicle 24 hours before forced swim testing. CSC or vehicle was directly infused into the ventral medial striatum 10 minutes before swim testing.

24 Hours post reserpine treatment 15

Float Time (minutes)

12 9 6 3 0

68

Res/DMSO

Res/CSC DMSO/DMSO DMSO/CSC Treatment

Figure 1 provides preliminary evidence activation of A2A receptors in the ventral medial striatum are critical mediators of reserpine-induced depression. Rats were implanted with bilateral guide cannulae in the ventral medial striatum. After recovery, groups either received an ip injection of 6 mg/kg reserpine or DMSO vehicle.

To activate A2A receptors, 30 nM of CSC or vehicle was infused in to the ventral medial striatum twenty-four hours later. Reserpine again produced a large deficit in swim performance relative to the vehicle control. These data provide preliminary evidence that A2A receptors in striatum mediate swim deficits following reserpine treatment, and more generally, the behavioral depression component of conservation-withdrawal.

III. Sickness Behavior

Enhanced synthesis of IL-1β in the brain parenchyma is evident as early as 2 hours after systemic treatment with LPS, with substantially higher concentrations of the proinflammatory cytokine being expressed 6 hours later in the NTS and the PVN and arcuate nuclei of the hypothalamus (47). Activation of hypothalamic IL-1β receptors is thought to mediate sickness behavior and related symptoms of conservation-withdrawal (48).

The foregoing experiments on reserpine-induced depression suggest that brain adenosine signaling might contribute importantly to sickness behavior as well.

Treatment with reserpine in the previous studies presumably produced some condition – e.g., cell damage in the periphery or brain – that resulted in the induction of brain IL-1ß. The symptoms of behavioral depression occurring with the induction of the brain cytokine appear to depend on concomitant activation of striatum A2A receptors, which ultimately impairs swim performance in reserpine-treated rats. As noted earlier, adenosine A2A signaling provides inhibitory feedback on proinflammatory cytokine signaling in peripheral immune cells (49). Evolution may have capitalized on this primitive cytokine-purine interaction to produce a more complicated brain interaction that regulates overt behavior at times of illness. The suppression of ongoing behavior is certainly an important aspect of sickness as a means of conserving resources and regulating fever. Although the precise nature of the interaction between brain IL-1β and adenosine is not clear, activation of A2A receptors in the striatum is precisely the type of mechanism to accomplish this outcome.

A preliminary test of this potential interaction was assessed by inducing sickness behavior with a system injection of LPS. All rats initially had a guide cannula

Thomas C. Hanff ET AL.

inserted in the right lateral ventricle during stereotaxic surgery. Following recovery, groups were injected with saline or LPS. Twenty-four hours later, one LPS-treated group received icv infusion of the IL-1ra and an ip injection of DMSO. Another LPS-treated group received icv infusion of saline and an ip injection of CSC. Groups Vehicle and LPS received icv infusion of saline and an ip injection of DMSO. All rats were tested in the forced swim task 15 minutes later.

A comparison of groups Vehicle and LPS indicate that treatment with endotoxin produced a large deficit in swim performance 24 hours later. This increase in floating time was completely reversed by either icv infusion of the IL-1ra or peripheral injection of the A2A receptor antagonist CSC. Additional evidence for a purine-cytokine interaction in sickness behavior was obtained by infusing IL-1β directly into the right lateral ventricle 1 hour before swim testing. The ensuing deficit in performance was completely reversed by icv infusion of the IL-1ra or ip injection of CSC. These data are in good agreement with recent findings in other animal models of depression. Yirmiya (38) initially reported that peripheral injection of endotoxin produced symptoms of major depression, including evidence of conservation-withdrawal within 24 hours of the injection. El Yacoubi et al. (50) demonstrated that endotoxin-induced swim deficits are reversed by systemic administration of A2A receptor antagonists.

The most plausible explanation for these data, in conjunction with those already discussed, is that behavioral depression or conservation-withdrawal is a “downstream” consequence of brain cytokine signaling.

In this context, activation of brain IL-1β receptors is a sufficient, but not a necessary condition for impaired test performance. Because the A2A receptor antagonist CSC universally reverses evidence of impaired test performance (51), it seemed likely that activation of brain IL-1β receptors recruits adenosine signaling at A2A to produce symptoms of behavioral depression.

Although data are still lacking, the likely site of adenosine’s action is the spiny GABAergic neurons of the striatum.

Overview This article has reviewed the evidence concerning the contribution of the proinflammatory cytokine IL-1β

and the purine nucleoside adenosine to the symptoms of conservation-withdrawal in two different paradigms (injection with reserpine and injection of endotoxin).

Symptoms of conservation-withdrawal are observed in both of these paradigms and are assumed to be an after reaction to intense catabolic output. The sensory unresponsiveness, cognitive dullness, and behavioral depression that characterize this state are assumed to be adaptive mechanisms for husbanding limited resources and facilitating the recovery of metabolic homeostasis.

IL-1β contributes directly to conservation-withdrawal following systemic treatment with reserpine or LPS. The cytokine’s contribution is delayed for 48 hours after reserpine treatment. At this point, IL-1β concentration rise substantially in the hypothalamus and hippocampus. The IL-1ra also is effective in reversing swim deficits at this time, but not earlier. We suspect that this rise in brain IL-1β may be related to tissue damage and the consequent activation of cytokine-producing microglia in the brain following reserpine treatment. Systemic LPS has a more immediately effect on brain IL-1β concentrations (52) and behavioral depression. The mechanism of the increase in this instance is likely to be immune-to-brain communication via the vagus (53).

The most direct mechanism of behavioral depression in these paradigms is activation of brain adenosine A2A receptors. Although more research is needed for stronger conclusions, the likely locus of these receptors is the spiny GABAergic neurons of the striopallidal (or indirect) path in the striatum.

Activation of these receptors appears to be necessary and sufficient for evidence of conservation-withdrawal in the learned helplessness, reserpine-induced depression, and LPS-induced sickness behavior paradigms. Moreover, behavioral deficits resulting from IL-1β receptor activation are reversed by antagonists of the A2A receptor, although the reverse is not true. This pattern of data may suggest that Il-1β receptor activation may recruit adenosine signaling in striatum A2A receptors to uncouple motivational influences from ongoing behavior.

There are a number of potentially important implications to these data. First, cytokine signaling appears to exacerbate and prolong the conservation-withdrawal reactions in major depression and illness. Downregulating this form of signaling should hasten recovery.

More important, the affect-less, fatigue components of stress, depression, and illness should be directly alleviated by manipulating the A2A/D2/mGLU heteromeric receptor complex. The present data clearly suggest that 69

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blockade of the A2A receptor produces direct benefit. If the conceptualization of this receptor complex is correct, additional benefit should be derived from a combination of A2A antagonists, D2 agonists, and mGLU, antagonists.

Such a combination should minimize fatigue and recouple motivation influences on ongoing behavior.

14. Sitovsky MV, Lukashev D, Apasov S. Physiological control of immune response and inflammatory tissue damage by hypoxia-inducible factors and adenosine A2A receptors.

Annu Rev Immunol 2004; 22:657-682.

References

16. Rosin DL, Hettinger BD, Lee A, Linden J. Anatomy of adenosine A2A receptors in brain: Morphological substrates for integration of striatal function. Neurology 2003;61:10-11.

1. Dantzer R. Cytokine-induced sickness behavior: Where do we stand? Brain Behav Immun 2001;15:7-24.

15. Popoli P, Minghetti L, Tebano MT. Adenosine A2A receptor antagonism and neuroprotection: Mechanisms, lights, and shadows. Crit Rev Neurobiol 2004;16: 99-106.

2. Larson SJ, Dunn AJ. Behavioral effects of cytokines. Brain Behav Immun 2001; 15:371-387.

17. Meghji P, Pearson JD, Slakey LL. Regulation of extracellular adenosine production by ectonucleotidases of adult rat ventricular myocytes. Am J Physiol 1992; 263:40-47.

3. Maes M. Evidence for an immune response in major depression: A review and hypothesis. Prog Neuropsychopharmacol Biol Psychiatry 1995;19:11-38.

18. Phillis JW. Adenosine and adenine nucleotides as regulators of cerebral blood flow: Roles of acidosis, cell swelling, and KATP channels. Crit Rev Neurobiol 2004;16:237-270.

4. Dantzer R. Cytokine-induced sickness behavior: Mechanisms and implications. Immunol Allergy Clin North Am 2009; 29:247-264.

19. Meghji P. Adenosine production and metabolism in the nervous system. In: Stone TW, editor. Adenosine in the nervous system. San Diego, Cal.: Academic, 1991.

5. Koo JW, Duman RS. Evidence for IL-1 receptor blockade as a therapeutic strategy for the treatment of depression.

Curr Opin Investig Drugs 2009;10:664-671.

20. White TD, Hoehn K. Release of adenosine and ATP from nervous tissue. In: Stone TW, editor. Adenosine in the nervous system. San Diego, Cal.: Academic, 1991: pp.173-196.

6. Engel GL, Schmale AH. Conservation-withdrawal: A primary regulatory process for organismic homeostasis. Ciba Found Symp 1972; 8:57-75.

21. Pickel VM, Chan J, Linden J, Rosin DL. Subcellular distributions of adenosine A1 and A2A receptors in the rat dorsomedial nucleus of the solitary tract at the level of the area postrema. Synapse 2006;60:496-509.

7. Woodson JC, Minor TR, Job RF. Inhibition of adenosine deaminase by erythro-9-(2-hydroxy-3-nonyl) adenine (EHNA) mimics the effect of inescapable shock on escape learning in rats. Behav Neurosci 1998; 112: 399-409.

8. Minor TR, Huang Q, Foley EA. Cytokine-purine interactions in behavioral depression in rats. Integr Physiol Behav Sci 2003; 38:189-202.

9. Mohta M, Sethi AK, Tyagi A. Psychological care in trauma patients. Injury 2003; 34:17-25.

10. Overmier JB, Seligman ME. Effects of inescapable shock upon subsequent escape and avoidance responding. J Comp Physiol Psychol 1967; 63:28-33.

11. West CH, Weiss JM. A selective test for antidepressant treatments using rats bred for stress-induced reduction of motor activity in the swim test. Psychopharmacology 2005;182:9-23.

22. Burns RF. Role of adenosine in supply/demand balance.

Nucleos Nucleotides 1991;10:931-933.

23. Marangos P. Potential therapeutic roles for adenosine in neurologic disease. In: Stone TW, editor. Adenosine in the nervous system. San Diego, Cal.: Academic, 1991: pp. 173-196.

24. Geiger JD, Padua RA, Nagy JI. Adenosine deaminase regulation of purine actions. In: Phillis JW, editor. Adenosine and adenine nucleotides as regulators of cellular function.

Boca Raton, Florida: CRC, 1991: pp. 71-84.

25. Rall TW. Drugs used in the treatment of asthma: The methylxanthines, cromolyn sodium, and other agents. I. In: Goodman-Gilman A, Rall TW, Nies AS, Taylor P, editors. Goodman and Gilman's the pharmacological basis of theraputics. New York: Pergamon, 1990: pp. 618-637.

26. Herrero MT, Barcia C, Navarro JM. Functional anatomy of thalamus and basal ganglia. Childs Nerv Syst 2002;18:386-404.

12. Minor TR, Dess NK, Overmier JB. Inverting the traditional view of â&#x20AC;&#x153;learned helplessness,â&#x20AC;? in fear, avoidance, and phobias: A fundamental analysis. In: Denny MR, editor.

Hillsdale, N.J.: Lawrence-Erlbaum, 1991: pp. 87-133.

27. Ikemoto K. [Human striatal D-neurons and their significance]. Nihon Shinkei Seishin Yakurigaku Zasshi 2002; 22:131-135.

13. Lukashev D, Ohta A, Apasov S. Cutting edge: Physiological attenuation of proinflammatory inscription of the Gs proytein-coupled A2A adenosine receptor in vivo. J Immunol 2004;173:21-24.

28. Hillion J, Canals M, Torvinen M. Coaggregation, cointernalization, and codesensitization of adenosine A2A receptors and dopamine D2 receptors. J Biol Chem 2002; 277:18091-18097.

70

Thomas C. Hanff ET AL.

29. Fuxe K, Agnati LF, Jacobsen K. Receptor heteromerization in adenosine A2A receptor signaling: Relevance for striatal function and Parkinson's disease. Neurology 2003; 61:19-23.

Vesicles store and release. In: Kandel ER, Schwartz JH, editors. Principles of neural science. London: E. Arnold, 1981: pp. 113-115.

30. Svenningsson P, Tzavara ET, Witkin JM. Involvement of striatal and extrastriatal DARPP-32 in biochemical and behavioral effects of fluoxetine (Prozac). Proc Natl Acad Sci USA 2002; 99:3182-3187.

43. Bean AJ, During MJ, Deutch AY. Effects of dopamine depletion on striatal neurotensin: Biochemical and immunohistochemical studies. J Neurosci 1989;9:4430-4438.

31. Diaz-Cabiale Z, Vivo M, Del Arco A. Metabotropic glutamate mGlu5 receptor-mediated modulation of the ventral striopallidal GABA pathway in rats. Interactions with adenosine A(2A) and dopamine D(2) receptors. Neurosci Lett 2002; 324:154-158.

32. Roitt I. Essential immunology. Malden, Mass.: Blackwell Science, Inc., 1997.

33. Maier SF, Watkins L. Cytokines for psychologists: Implications of bidirectional immune-to-brain communication for understanding behavior, mood, and cognition.

Psychological Review 1998;105:83-107.

34. Dantzer R. Cytokine, sickness behavior, and depression.

Brain Behav Immun 2009;15:7-24.

35. Pollack Y, Yirmiya R. Cytokine-induced changes in mood and behavior: Implications for “depression due to a general medical condition,” immunotherapy and antidepressive treatment. Int J Neuropsychopharmacology 2002;5:389-399.

36. Hestad KA, Tonseth S, Stoen CD Ueland T, Aukrust P. Raised plasma levels of tumor necrosis factor alpha in patients with depression: Normalization during electroconvulsive therapy. J ECT 2004;19:183-188.

37. del Cerro S, Borrell J. Interleukin-1 affects the behavioral despair response in rats by an indirect mechanism which requires endogenous CRF. Brain Res 1990; 528:162-164.

38. Yirmiya R. Endotoxin produces a depressive-like episode in rats. Brain Res 1996; 711:163-174.

39. Huang QJ, Jiang H, Hao XL, Minor TR. Brain IL-1 beta was involved in reserpine-induced behavioral depression in rats. Acta Pharmacol Sin 2004;25:293-296.

40. Huang Q, Jiang H, Minor TR. Reserpine increases brain IL-1beta expression in rat brain. Chinese J Immunol 2003; 19:765-767.

41. Minor TR. Conditioned fear and neophobia following inescapable shock. Anim Learn Behav 1990;18: 212-226.

42. Schwartz JZ. Chemical basis of synaptic transmission:

44. Soto-Otero R, Mendez-Alvarez E, Sanchez-Sellero I. Reduction of rat brain levels of the endogenous dopaminergic proneurotoxins 1,2,3,4-tetrahydroisoquinoline and 1,2,3,4-tetrahydro-beta-carboline by cigarette smoke.

Neurosci Lett 2001; 298:187-190.

45. Kelley KW, Bluthé RM, Dantzer R, Zhou JH, Shen WH, Johnson RW, Broussard SR. Cytokine-induced sickness behavior. Brain Behav Immun 2003;15:112-118.

46. Koga K, Takaesu G, Yoshida R, Nakaya M, Kobayashi T, Kinjyo I, Yoshimura A. Cyclic adenosine monophosphate suppresses the transcription of proinflammatory cytokines via the phosphorylated c-Fos protein. Immunity 2009; 30:372-383.

47. Chennaoui M, Drogou C, Gomez-Merino D. Effects of physical training on IL-1beta, IL-6 and IL-1ra concentrations in various brain areas of the rat. Eur Cytokine Netw 2008;19:8-14.

48. Ferre S, Fredholm BB, Morelli M. Adenosine-dopamine receptor-receptor interactions as an integrative mechanism in the basal ganglia. Trends Neurosci 1997;20:482-487.

49. Trincavelli ML. Costa B, Tucano D, Lucacchini A, Martin C. Up-regulation of A2A adenosine receptors by proinflammatory cytokines. Biochem Pharamocol 2001;4:625-631.

50. El Yacoubi M, Ledent C, Parmentier M. Adenosine A2A receptor antagonists are potential antidepressants: Evidence based on pharmacology and A2A receptor knockout mice.

Br J Pharmacol 2001;134:68-77.

51. Minor TR, Rowe M, Cullen PK, Furst SJ. Enhancing brain adenosine signaling with the nucleoside transport blocker NBTI (S-(4-nitrobenzyl)-6-theoinosine) mimics the effects of inescapable shock on later shuttle-escape performance in rats. Behav Neurosci 2008;122:1236-1247.

52. Licinio J, Wong ML. Interleukin 1 receptor antagonist gene expression in rat pituitary in the systemic inflammatory response syndrome: Pathophysiological implications. Mol Psychiatry 1997; 2:99-103.

53. Anisman H, Merali Z. Anhedonic and anxiogenic effects of cytokine exposure. Adv Exp Med Biol 1999;461:199-233.

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Isr J Psychiatry Relat Sci - Vol 47 - No. 1 (2010)

Genetics of Unipolar Major Depressive Disorder Tanya Goltser-Dubner, MSc,1 Esti Galili-Weisstub, MD,2 and Ronnen H. Segman, MD1 1

Department of Psychiatry, Hadassah Hebrew University Hospital, Jerusalem, Israel Herman Dana Division of Child and Adolescent Psychiatry, Department of Psychiatry, Hadassah Hebrew University Hospital, Jerusalem, Israel

2

ABSTRACT Major depressive disorder (MDD) is a heterogeneous, highly prevalent, and moderately heritable disorder.

A complex and diverse genetic-environmental interplay converges to set apart a significant minority that is susceptible to MDD, from among those who experience shorter lived and less recurrent intensive and incapacitating forms of sadness. The major technological advances of deciphering the human genome reference sequence and its common gene variations are beginning to allow cost effective genetic studies of unprecedented scale, applying increasingly denser genome wide mapping to increasingly larger case control samples. This effort is now at the initial stages of unraveling the genetic architecture of several complex phenotypes. Despite a tardy beginning, MDD genetic research is maturing from modest scale candidate gene association studies to include family-based linkage studies, and will soon allow genome wide case control association studies. Replicated risk conferring gene variants discovered so far exert a modest effect size that appears to contribute to overt phenotype expression in the context of a highly intricate concert of interrelated epigenetic and epistatic modifiers.

The unraveling of additional previously unimplicated MDD risk conferring genes, that will throw light on molecular mechanisms mediating such susceptibilities, is necessary for progressing beyond current generation monoamine modulating antidepressant drugs. The review outlines basic concepts and current progress briefly overviews major replicated gene findings that to date mostly stem from hypotheses driven candidates, and ends with a discussion of current directives,

including sample size and phenotype considerations and advancement of systematic studies of the functional significance of implicated gene variants, beyond their current exploratory stage.

Introduction Basic understanding of MDD pathophysiology is limited, and has been driven in large part by extrapolating from putative mechanisms of drugs found to possess antidepressant efficacy. Genetics circumvents the need to access the relevant molecular pathophysiology, by associating sequence variation with phenotype, thus offering an important window for discovering previously unimplicated mechanisms. The completion of the genome reference sequence and increasing availability of genome scale molecular tools is beginning to transform all areas of medicine. Our understanding of complex psychiatric phenotypes such as MDD has much to gain from this genomic revolution, as the majority of common psychiatric disorders as we currently define them, based on clinical presentation and subjective symptom resemblance, show considerable heritability.

Heritability Family twin and adoption studies are used to estimate the extent of heritable contribution for a phenotype of interest. MDD aggregates in families as demonstrated by a 2.84 increased risk for a broad MDD phenotype among first degree relatives of affected probands (1).

Whereas mere familial clustering of cases does not distinguish between a shared environmental vs. a shared

Address for Corrsepondence: Ronnen H. Segman, MD, Department of Psychiatry, POB 24035, Hadassah Mt. Scopus Hospital, Jerusalem 91240, Israel. â&#x20AC;&#x2020; ronense@ekmd.huji.ac.il

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genetic cause, a heritable effect is demonstrated when concordance for sharing the phenotype among relatives increases as a function of increased sharing of genetic sequence (e.g., as occurs among relatives with decreasing familial distance, or when comparing monozygotic twins that share the same DNA sequence, with dizygotic twins that share half of their sequence on average, as would any sibling pair). Heritability estimates for MDD based on monozygotic vs. dizygotic twin concordance differences exhibit a modest heritable contribution of 37% (1) with a greater estimated contribution of 42% for women vs. 29% for men (2). Limited adoption studies point to an important genetic impact of parental depression (3), but also a significant environmental impact of maternal depression in mediating depression among adopted adolescents (4). Although the heritable contribution for broadly defined MDD is about half of that found for bipolar disorder or schizophrenia, its prevalence is several fold higher, and its permissive phenotype definition may be more prone to the inclusion of multiple etiologies, if these share a sufficient number of similar diagnostic criteria. Such heterogeneity may be one reason behind the low rates of response to generic antidepressants that target monoaminergic neurons (5).

Increased severity, high recurrence rate and early onset phenotype characteristics exhibit increased heritability (as gauged by the observation that such characteristics among index probands predict higher rates of depression among their relatives) (6). When applying such model fitting and accounting for measurement error, heritability estimates for a restrictive MDD phenotype can be shown to increase to over 70% (reviewed in 7). Chronic illness course has also been shown to be associated with increased familial aggregation among recurrent, early-onset MDD pedigrees (8). Restrictive phenotype aspects have been applied to guide ascertainment considerations in several linkage based genome scan studies described below. Another important facet of diagnostic specificity relates to disorders that share some phenotypic aspects with MDD, such as bipolar disorder, anxiety disorders or internalizing personality traits. It is clear that some cases observed to satisfy diagnostic criteria for unipolar depression may have differing etiologies (e.g., bipolar disorder is one example).

Misdiagnoses potentially carry important treatment implications (e.g., bipolar depression is better treated with mood stabilizers), and there is an ongoing debate as to their prevalence (9). There is evidence for familial aggregation and some shared heritability for MDD with

both BPD (10) and anxiety disorders (11). Neuroticism, to name a different diagnostic construct, reflects in large part genetic liability for MDD (12). As these partially overlapping phenotype distinctions are likely polygenic in etiology (13, 14), we might expect that a single small effect gene could contribute in a modular way to more than one categorical phenotype, depending on individual constellations of interactive epistatic and epigenetic modifiers. Such a possibility is supported both by evidence for some shared inheritance between these phenotypes, as well as some overlap in clinical presentation (e.g., some shared symptoms, medication response, etc.). Indeed, as detailed below, some gene polymorphisms, such as the serotonin transporter promoter polymorphism and neurotrophic tyrosine kinase receptor 3 (NTRK3) gene variations, have been shown to predispose to more than one categorical phenotype.

Currently employed categorical phenotype definitions are formulated using threshold-based descriptive symptom clustering that mostly relies on subjective self report and lack valid biomarkers, resulting in permissive boundaries that constrain our ability to ascertain cases sharing homogeneous biological underpinnings.

From a geneticistâ&#x20AC;&#x2122;s point of view, this compromises our odds for locating common causative gene variations.

Issues of phenotype definition for genetic studies are further discussed in more detail below.

Genes Strategies for locating involved genes

Once a heritable contribution has been established, we may turn to locate the actual genes or, more precisely, genomic template nucleotide sequence variations that transmit such heritable risk. Historically, efforts for locating disease risk conferring genes have either focused on hypothesis driven candidates or employed an unbiased exploration using linked polymorphic markers that span the whole genome in search of previously unimplicated loci. Allele and genotype frequencies of nucleotide sequence variations within a candidate gene, that is thought to possess a priori relevance to MDD pathogenesis, may be compared between cases vs. ethnically matched healthy unrelated controls in search of association (or by using parental DNA in search of deviation from expected even transmission rates to affected offspring). Genome wide linkage is based on genome scale systematic mapping of the inheritance patterns of evenly spaced polymorphic markers in 73

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search of linked markers that are co-inherited with a risk conferring gene variant among affected family relatives, thus implicating susceptibility linked chromosomal regions harboring a causative variant. Allelic variants in both adjacent marker and risk gene loci may be co-transmitted if their proximity on the same chromosome impedes crossing over between them during meiosis, resulting in deviation from independent assortment. The completion of the human genome reference sequence and increasingly detailed knowledge of single nucleotide polymorphisms (SNPs) now allow genome wide association studies through dense mapping at a range of 106 SNPs. Here adjacent SNPs that are in linkage disequilibrium with a risk conferring polymorphism form haplotype blocks that also show association with the phenotype and may be used to fine-map the co-transmitted risk variant. Alternatively, a functional SNP that has a causative role may be directly detected by showing deviant allelic frequencies among affected individuals as compared with controls. SNP microarrays thus allow cost effective genome wide association studies (GWAS) using increasingly larger case control samples. Furthermore, genomic microduplications and microdeletions, also known as structural variants or copy-number variants (CNVs), have been demonstrated to have a high prevalence as well as a causative role in mediating complex psychiatric disorders (15), and genome scale copy-number variation can be studied alongside single nucleotide variation using a single microarray platform (16).

Modification of gene variation effects

An implicated gene sequence variation may or may not acquire functional significance for conferring risk to a phenotype of interest, in the context of a complex interchange that includes several players. Genes are transcribed into mRNA and translated into protein in a cell X time X function specific manner. Epigenetic modifications of the template DNA nucleotide sequence are a set of environmentally modulated complex molecular interactions (e.g., DNA methylation, chromatin remodeling) regulating the where, when, and to what extent a certain gene may be expressed. A Gene X Environment (GXE) interaction occurs when nucleotide sequence variations become important in mediating risk for a phenotype in the context of relevant exposure, as has been arguably demonstrated for the serotonin transporter gene promoter polymorphism and discussed in detail below. The impact of a risk conferring gene varia74

tion may be further modified by interactive effects of other variations in the same gene or in other relevant genes. Epistasis or Gene X Gene (GXG) interaction takes place when the action of one gene is modified by that of another or several other (e.g., modifier) genes, and the phenotypic consequences of any one allele may generally depend on multiple other alleles in a highly complex interactive manner.

Implicated MDD risk gene variants Genome scale studies

Despite its high prevalence, the study of MDD has lagged behind and has been less extensively addressed by genome wide studies, as compared with the less frequent but more heritable psychiatric disorders (e.g., schizophrenia and bipolar disorder). A number of MDD linkage studies have been performed to date (1720, 21-24) with several chromosomal regional implications reported by more than one study (e.g., reviewed in 22, 25, 26). Most of these studies ascertained familial cases with homogeneous restrictive and highly heritable phenotypic characteristics (e.g., featuring early age at onset, high recurrence rate, a more severe and chronic clinical course, high familial loading, gender specific sub analyses etc.), that are more likely to share common genetic underpinnings. In addition to the lower statistical power of linkage based analysis (see, for example, 27), a major limitation of family based linkage studies is an inherent difficulty to ascertain sizable numbers of families with multiple affected relatives sharing restrictive phenotype characteristics that will allow a large enough sample powered to detect variants with a small effect size. Further, large chromosomal regions shared among family members constrain the narrowing down of a linkage signal sufficiently to identify a causative gene. Both within a cohort and between cohorts, the percentage of families sharing a similar founder effect may be limited, despite efforts to ascertain cohorts with a homogeneous ethnic background (e.g., in both North American and Northern European populations there is little evidence for founder effects, 28). Detection of a genomic signal may thus be muted in a large cohort in which only some of the families carry a certain risk conferring variant, and replication of reported signals may be limited by genetic heterogeneity in different cohorts (29). Genome wide MDD case control association samples (GWAS) are currently being ascertained (30, 31), with an emphasis on large scale samples (e.g., incorporating a growing understanding in the field of

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genetics of complex traits that even several thousand cases and controls may not be enough to capture modest risk conferring variants). The long-term effort required for ascertaining such exceedingly large samples may be very much worth while, as they are likely to eventually shed new light on the largely uncharted biology of MDD. NTRK3 gene localization through positional cloning. The Genetics of Recurrent Early-Onset Depression (GenRED) linkage project reported preliminary genome scan evidence from 297 families implicating the 15q25.326.2 chromosomal region with a restrictive phenotype of recurrent early-onset MDD (17). This region was independently implicated by two other independent linkage studies that focused on a similar restrictive phenotype, the European-U.S. Depression Network (DeNt) study (21) with 497 affected Sibling pairs, and a linkage study of 87 extended pedigrees from Utah (24), as well as in a final extended genome scan analysis (23, 32). Linkage fine mapping of this region with SNPs in 631 families produced genome-wide significant evidence for linkage (32). The authors further genotyped 1,195 individuals from 300 informative European ancestry pedigrees with multiple relatives with recurrent early onset MDD, for 795 SNPs, applying linkage disequilibrium mapping, and located several nominally significant candidate genes in the region (33). One of these, the NTRK3 gene, encodes a receptor that binds neurotrophin 3 (NT3), and may be involved in MDD pathogenesis.

The NTRK3 finding was recently replicated in a sample consisting of 603 families with 723 affected children and adolescents diagnosed with a mood disorder with onset of the first episode by age 15 (34). The NTRK3 gene thus represents a potentially important candidate gene successfully arrived at through linkage derived positional cloning. The gene encodes trkC which is preferentially expressed in relevant brain regions and constitutes a particularly attractive candidate that fits elegantly with the neural plasticity theory of antidepressant treatment mechanism and depression (35, 36), and warrants further replications attempts, and functional characterization.

Candidate genes A specific gene may either be implicated as a candidate for conferring risk to a phenotype through a previous hypothesis that purports its function may possess pathogenetic implication, or is identified through

being linked with an informative marker in the context of a genome screen (positional cloning, such as with NTRK3). Hypotheses driven candidate gene search is difficult to conduct when the disorder at hand has little in the way of known pathobiology. Important leads that have attracted extensive interest in terms of generating hypotheses for MDD candidate gene search include antidepressant drug targets and the neuroendocrine and neuroimmune pathways. Currently available antidepressants act to increase monoaminergic neurotransmission, and may exert a therapeutic effect in part through augmenting neural plasticity processes (37). Sequence variations in genes encoding antidepressant drug targets constitute immediate candidates for influencing therapeutic and adverse response patterns, but may also impact on risk for MDD. Candidate gene associations with MDD have been recently reviewed (38, 39). Lopez-Leon et al. reviewed MDD association studies reported in 183 papers that studied 393 polymorphisms in 102 genes (38). Only 22 of these polymorphisms were investigated by three or more studies allowing a meta-analysis, of these they found significant evidence for association for APOE, DRD4, GNB3, MTHFR, SLC6A3 and SLC6A4 (38). Below we review some of the candidate gene findings replicated in several studies or meta-analyses, and important initial findings from larger scale studies.

The serotonin transporter. The serotonin transporter gene solute carrier family 6 (neurotransmitter transporter, serotonin), member 4 (SLC6A4) gene located on chromosome 17q11.1â&#x20AC;&#x201C;q12, is the exclusive therapeutic target for the derived second generation selective serotonin reuptake inhibitor class of drugs. Several sequence variations have been described in the gene, the most extensively studied is 5-hydroxytriptamine transporterlinked polymorphic region (5-HTTLPR) a 43 base pair Insertion/Deletion short/long (S/L) polymorphism in the promoter (40), which together with rs25531 an A/G substitution within the Longer allele creates a functional AP2 transcription-factor binding site reported to interact to reduce gene expression to a level comparable to that of the short allele (41). Additional variations have been described in the same polymorphic region that may result in a further effect on transcription (42, 43). The ancillary pharmacogenetic arm of the Sequenced Treatment Alternatives to Relieve Depression (STAR*D) study, the largest single study to date with 1,953 participants, could not confirm association with therapeutic response, but reported a positive association of the grouped short allele 75

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and long-G allele combination (e.g., alleles reported to reduce mRNA expression) with adverse effect burden induced by the SSRI citalopram (44). A recent metaanalysis summarizing nine studies with 2,462 participants found significant evidence for association of the 5-HTTPLR with antidepressant response, as well as for additional polymorphisms in smaller overall samples, including the serotonin transporter intron 2 (STin.2) and sequence variations in the serotonin receptor genes HTR1A, HTR2A, the tryptophan hydroxylase gene (TPH1) and the brain derived neurotrophic factor (BDNF) gene (45). The serotonin transporter promoter variation has also been associated with suicidality (46), and with bipolar disorder, albeit with a modest OR of 1.12 (46). Evidence of association with anxiety related personality traits of harm avoidance (47) and neuroticism (48) (e.g., shown to possess partly shared heritability with depression) does not appear to hold for harm avoidance, and requires further study for neuroticism (e.g., 49, 50).

Stressful events have been shown to increase risk for MDD (51), and different adverse events are causally associated with distinct depressive symptoms (52).

Caspi et al. (53), employing a follow up study of 1,000 children through young adulthood, demonstrated 5-HTTLPR short allele carriers were more prone to serious depression if experiencing stressful life events either during childhood or during the years preceding the depressive episode, suggesting a GXE interactive effect whereby the capacity of early stress exposure to alter the brain’s susceptibility for depression, depends in part on 5-HTTPLR genotype. This report was followed by numerous reports investigating a role for stress exposure as a modifier of 5-HTTPLR impact on depression, with inconclusive over all replication results to date (e.g., reviewed in 54), and more recently refuted in a large meta-analysis (55). Proposed methodological strategies for gene environmental studies have been previously reviewed (56), and further investigation of the molecular mechanisms mediating such GXE interactive effect remains an important field for further study requiring much larger sample sizes (57).

Other MDD risk genes. Lopez-Leon et al. found positive evidence for association for the MTHFR C677T with MDD employing a meta-analysis of six association studies with a total of 875 cases and 3,859 controls with a combined OR of 1.20 (CI, 1.07–1.34) for the T allele (38). They also found the APOE epsilon2 allele to show a significant protective effect for MDD in a meta-analysis of seven studies including a total of 827 cases and 1,616 76

controls, with a pooled OR of the epsilon2 allele compared to the epsilon3 of 0.51 (CI, 0.27–0.97) (38). LopezLeon et al. (38) further calculated positive associations for the dopamine D4 receptor gene (DRD4) 2 allele with unipolar disorder and mixed affective (bipolar and unipolar) disorder in a meta-analysis of 12 samples with a total of 917 affective patients and 1,164 controls, the GNB3 C825T using three studies with a total of 375 cases and 492 controls (T allele OR, 1.38; CI, 1.13–1.69); and the dopamine transporter gene (SLC6A3) 40 base pair VNTR showed positive association calculated from three studies including a total of 151 cases and 272 controls, with a pooled OR for the 9/10 genotype compared to the 10/10 genotype of 2.06 (CI, 1.25–3.40) (38). The G72 gene, previously associated with schizophrenia and BPD, was reported to associate with MDD and neuroticism (58).

Several immune related gene associations with MDD have been reported. The P2RX7 gene is located within a region on chromosome 12q24.31 that has been identified as a susceptibility locus for affective disorders by linkage and association studies. P2RX7 is a purinergic ATP-binding calcium channel that modulates monocyte/macrophageinduced inflammatory response and is also expressed in neurons and glia. A non-synonymous coding SNP in the P2RX7 gene (rs2230912) resulting in amino acid substitution (Gln460Arg), that was previously found to be associated with bipolar disorder (59), was significantly associated with MDD, among 1,000 German Caucasian patients compared with controls (60). Proteasome beta4 subunit (PSMB4), and TBX21 (T bet) genes important in T lymphocyte function (61), and cyclic nucleotide phosphodiesterases PDE9A and PDE11A genes (62) were also reported to associate with MDD risk. All these association reports await further replications.

Endophenotype gene associations. Endophenotypes represent more elementary phenotypic measures that may be more likely to correlate with small effect sequence variants. Depression may be associated with hippocampal volume reduction, in part through a reduced capacity for neurogenesis, and neurotrophic factors including brain-derived neurotrophic factor (BDNF) and vascular endothelial growth factor (VEGF) have both been implicated in modulating hippocampal plasticity (35, 63). There is no consistent evidence associating common BDNF gene polymorphisms with MDD, although significantly smaller hippocampal volumes were observed for patients and for controls carrying the BDNF Val66Met Met allele (64-66). Similarly VEGF gene SNP variations have been shown to corre-

Tanya Goltser-Dubner et al.

late with hippocampal volume among healthy subjects (67). The 5-HTTPLR short allele has been reported to correlate with augmented amygdala activation to fearful stimuli which may be related to trait negative affectivity and depression (68). A summary of several replication attempts provides some support to this finding (69).

To conclude, antidepressant drug targets and also neuroimmune and neuroendocrine modulators have been extensively invoked to guide candidate gene selection for MDD association studies. Methodological aspects of candidate gene association studies have progressed to employ increasingly larger case control samples or meta-analytic summaries to increase sample size (38, 70). The majority of reported studies to date have failed to incorporate comprehensive scans encompassing all known sequence variations in any gene of interest. As an example, although the SLCA4 gene contains multiple known variations (42, 43), the majority of studies focused on promoter variations (e.g., mostly 5-HTTLPR). Studies that examined additional SLC6A4 SNPs and haplotypes reported their relevance for both gene expression (42, 43) (71) as well as for mediating stress X SLC64A genotype effects on the depressive phenotype (72). Further, few studies have looked for epistatic interactions between different candidate susceptibility loci (72-76). Such studies generally would require larger samples unless a large effect size is produced by interactive loci.

The following step of unraveling the biological significance of implicated risk variations is currently at its beginning stages. It follows both that functional knowledge of gene variations is currently too scant to justify premature focus of phenotype associations on any single sequence variant, and that reciprocal systematic input from such studies would result in a more informed focus on risk conferring gene variations for functional studies. Recent initiatives for ascertainment of large scale samples for genome wide association studies may yield new findings of previously unknown MDD risk conferring genes in the near future.

Current directives for study

With few exceptions, replicated MDD gene association findings to date derive from modestly sized metaanalyses that combine small case control samples with non-uniform ascertainment characteristics. Further, a majority of studies to date address hypotheses driven candidates that mostly revolve around known mechanisms of antidepressant drugs. As noted, such studies have prematurely devoted disproportionate focus to

few highly reported polymorphisms (e.g., such as the 5-HTTPLR promoter variation), neglecting comprehensive coverage of the coding and promoter regions of the gene of interest. Despite the extensive efforts described and some progress made, the hypotheses driven candidate gene strategy is unlikely to provide ground-breaking perspectives that will furnish a new understanding of the largely unknown biology of depression.

Elucidation of previously unimplicated genes may come from unbiased genome scale family based linkage studies, and eventually from genome scale case control studies once large enough samples (e.g., the lower limit of which is several thousands cases) have been ascertained, as has recently been the case for schizophrenia (77). A conspicuous finding from numerous recent replicated gene findings across several complex phenotypes such as schizophrenia (77-79), type II diabetes (80, 81), and rheumatoid arthritis (82) is the small effect size conferred by implicated gene variants. Such findings could only be reliably detected with employing exceedingly large case control samples, highlighting the importance of multicenter collaborative efforts and meta-analyses for meaningful GWAS results. This does not discount smaller family based linkage studies, as these may discover mutations that bear a higher relative risk for familial restrictive MDD phenotypes among a cohort that shares a founder effect, despite a lower overall sample size. A common genetic variant in the same gene may confer a much lower relative risk for sporadic MDD, in which case a population based GWAS may require a much larger unrelated case control sample to detect a minor effect size that exhibits genome wide significance level. The frustratingly low and variable relative risks attributable to implicated gene variations in the context of recent population based case control GWA studies of multiple complex disorders among samples sized at 104 subjects pose the immediate question of how relevant these true replicated findings may be for predicting actual risk for the disorder. The unfolding genetic landscape of schizophrenia thus far appears to be populated with both a prevalent polygenic type resulting from the convergence of several common polygene variants each contributing a minute relative risk to the sporadic disorder, as well as rare cases of copy number variations that hold a very high relative risk for carriers (15, 83, 84). This combination of cases may account for a stable rate of schizophrenia over generations despite reduced fecundity of those affected, as the heritable pool for common gene variations that confer risk to the sporadic phenotype lies with the unaffected 77

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general population, whereas rare CNVs may mostly arise among affected individuals de novo, and are therefore independent of fecundity. Such patterns raise questions about future practicality of predictive tests, as inherited minute effect poly genes cannot be ascribed deterministic stigmatizing predictions for an individual carrier, but rather portray probabilistic propensities that may become conductive of a categorical phenotype only within a given epistatic and epigenetic context. If common gene variants each contribute so little to the biology of the complex phenotype, and rare structural variants apply to very few of the patients, what implications could such findings possibly have for guiding the development of novel targeted preventive and palliative molecular interventions that could be useful to the majority of patients?

Several of the structural variants as well as minor risk SNP variations discovered appear to converge in disrupting common neurodevelopmental pathways such as neuregulin signaling (e.g., 15), or the interactive DISC I and PDE4B genes, the disruption of either of which may result in a schizophrenia phenotype (85), leading to the suggestion that genes in such relevant pathways may either be severely disrupted leading to rare cases, or contain common minor variations contributing to the common polygenic phenotype (15). Discoveries of either type of variation may converge to implicate the same pathogenetically relevant genes and pathways that possess common relevance as targets for significant interventions.

Conclusion Non-hypotheses driven GWA studies are transforming our understanding of the genetic architecture and pathophysiology of common complex medical disorders. Since 2005, nearly 100 replicated risk conferring gene variants have been reported for as many as 40 common diseases (86). This progress contrasts with the slow progress in complex trait research during the previous two decades, and largely stems from rapid increments in the ability to apply denser SNP mapping to larger case control samples. MDD research is currently on the verge of applying such large scale GWA studies. The published literature surveyed above is to date mostly compiled of reports stemming from modest case control samples exploring candidate gene association with categorical MDD, few of which withstand meta-analytic validation of independent replication attempts. In a similar vein, reviews of hundreds of association reports across multiple medical disorders demonstrate the importance of 78

large scale independent replication and meta-analyses for teasing out replicated findings from frequent nonreplications that may result from population stratification, phenotype differences, selection biases, genotyping errors, etc. (e.g., 87, 88). Detailed guidelines have recently been proposed for replicating genetic association (89) and for assessing the validity of cumulative evidence for genetic association findings (90, 91). Empirically derived statistical power estimates suggest that GWAS for complex phenotypes will typically require several thousands of cases to study main effects and several tens of thousands of cases to properly support the investigation of gene X gene or gene X environment interactions (92).

The modest relative risks conferred by hitherto discovered common gene variations in psychiatric genetics signify probabilistic propensities, with little if any practical diagnostic predictive value, and are not likely to bear deterministic (e.g., neither stigmatizing nor of practical clinical value) attributes for the individual. Future comprehensive knowledge of sets of multiple interacting small effect risk loci might have practical predictive implications, in the context of clinical risk factors and ethnic derivation. The small effect size of risk conferring gene variations makes a much harder case for efforts to decipher their incremental biological contribution to the pathogenesis of a polygenic multifactorial disorder. This is in contrast to karyotype abnormalities and CNVs that have been identified so far to result in very high relative risks for overlapping major psychiatric disorders and cognitive deficits, but apply to rare cases and may mostly arise de novo. Although unlikely to reform the next 2012 addition of DSM-V, genetics will definitely have an increasing input into diagnostic reclassification. More importantly, the discovery of risk loci will furnish the basis for a fresh understanding of the pathophysiology of these idiopathic conditions, offering hope for the design of selective molecular interventions for the prevention and treatment of psychiatric disorders.

Acknowledgements This work was supported by a NARSAD independent investigator award, an Israel Science Foundation Grand 1563-08 and the HWZOA women's health foundation award to RHS.

References 1. Sullivan PF, Neale MC, Kendler KS. Genetic epidemiology of major depression: review and meta-analysis. Am J Psychiatry 2000; 157: 1552-1562.

2. Kendler KS, Gatz M, Gardner CO, Pedersen NL. A Swedish

Tanya Goltser-Dubner et al.

national twin study of lifetime major depression. Am J Psychiatry 2006; 163: 109-114.

3. Wender PH, Kety SS, Rosenthal D, Schulsinger F, Ortmann J, Lunde I. Psychiatric disorders in the biological and adoptive families of adopted individuals with affective disorders. Arch Gen Psychiatry 1986; 43: 923-929.

4. Tully EC, Iacono WG, McGue M. An adoption study of parental depression as an environmental liability for adolescent depression and childhood disruptive disorders. Am J Psychiatry 2008; 165: 1148-1154.

5. Trivedi MH, Rush AJ, Wisniewski SR, Nierenberg AA, Warden D, Ritz L, et al. Evaluation of outcomes with citalopram for depression using measurement-based care in STAR*D: implications for clinical practice. Am J Psychiatry 2006; 163: 28-40.

6. Kendler KS, Gardner CO, Prescott CA. Clinical characteristics of major depression that predict risk of depression in relatives. Arch Gen Psychiatry 1999; 56: 322-327.

7. McGuffin P, Cohen S, Knight J. Homing in on depression genes. Am J Psychiatry 2007; 164: 195-197.

8. Mondimore FM, Zandi PP, Mackinnon DF, McInnis MG, Miller EB, Crowe RP, et al. Familial aggregation of illness chronicity in recurrent, early-onset major depression pedigrees. Am J Psychiatry 2006; 163: 1554-1560.

9. Stensland MD, Schultz JF, Frytak JR. Diagnosis of unipolar depression following initial identification of bipolar disorder: A common and costly misdiagnosis. J Clin Psychiatry 2008; 69: 749-758.

10. Karkowski LM, Kendler KS. An examination of the genetic relationship between bipolar and unipolar illness in an epidemiological sample. Psychiatr Genet 1997; 7: 159-163.

11. Kendler KS, Gardner CO, Gatz M, Pedersen NL. The sources of co-morbidity between major depression and generalized anxiety disorder in a Swedish national twin sample. Psychol Med 2007; 37: 453-462.

12. Kendler KS, Gatz M, Gardner CO, Pedersen NL. Personality and major depression: A Swedish longitudinal, population-based twin study. Arch Gen Psychiatry 2006; 63: 1113-1120.

13. Nurnberger JI, Jr. A simulated genetic structure for bipolar illness. Am J Med Genet B Neuropsychiatr Genet 2008; 147B: 952-956.

16. McCarroll SA, Kuruvilla FG, Korn JM, Cawley S, Nemesh J, Wysoker A, et al. Integrated detection and populationgenetic analysis of SNPs and copy number variation. Nat Genet 2008; 40: 1166-1174.

17. Holmans P, Zubenko GS, Crowe RR, DePaulo JR, Jr., Scheftner WA, Weissman MM, et al. Genomewide significant linkage to recurrent, early-onset major depressive disorder on chromosome 15q. Am J Hum Genet 2004; 74: 1154-1167.

18. Nurnberger JI, Jr., Foroud T, Flury L, Su J, Meyer ET, Hu K, et al. Evidence for a locus on chromosome 1 that influences vulnerability to alcoholism and affective disorder.

Am J Psychiatry 2001; 158: 718-724.

19. Zubenko GS, Maher B, Hughes HB, 3rd, Zubenko WN, Stiffler JS, Kaplan BB, et al. Genome-wide linkage survey for genetic loci that influence the development of depressive disorders in families with recurrent, early-onset, major depression. Am J Med Genet B Neuropsychiatr Genet 2003; 123B: 1-18.

20. Abkevich V, Camp NJ, Hensel CH, Neff CD, Russell DL, Hughes DC, et al. Predisposition locus for major depression at chromosome 12q22-12q23.2. Am J Hum Genet 2003; 73: 1271-1281.

21. McGuffin P, Knight J, Breen G, Brewster S, Boyd PR, Craddock N, et al. Whole genome linkage scan of recurrent depressive disorder from the depression network study. Hum Mol Genet 2005; 14: 3337-3345.

22. Middeldorp CM, Sullivan PF, Wray NR, Hottenga JJ, de Geus EJ, van den Berg M, et al. Suggestive linkage on chromosome 2, 8, and 17 for lifetime major depression. Am J Med Genet B Neuropsychiatr Genet 2009; 150B:352-358.

23. Holmans P, Weissman MM, Zubenko GS, Scheftner WA, Crowe RR, Depaulo JR, Jr., et al. Genetics of recurrent early-onset major depression (GenRED): Final genome scan report. Am J Psychiatry 2007; 164: 248-258.

24. Camp NJ, Lowry MR, Richards RL, Plenk AM, Carter C, Hensel CH, et al. Genome-wide linkage analyses of extended Utah pedigrees identifies loci that influence recurrent, early-onset major depression and anxiety disorders. Am J Med Genet B Neuropsychiatr Genet 2005; 135B: 85-93.

25. Hamet P, Tremblay J. Genetics and genomics of depression. Metabolism 2005; 54: 10-15.

26. Craddock N, Forty L. Genetics of affective (mood) disorders. Eur J Hum Genet 2006; 14: 660-668.

14. Hovatta I, Barlow C. Molecular genetics of anxiety in mice and men. Ann Med 2008; 40: 92-109.

27. Risch N, Merikangas K. The future of genetic studies of complex human diseases. Science 1996; 273: 1516-1517.

15. Walsh T, McClellan JM, McCarthy SE, Addington AM, Pierce SB, Cooper GM, et al. Rare structural variants disrupt multiple genes in neurodevelopmental pathways in schizophrenia. Science 2008; 320: 539-543.

28. Hardy J, Low N, Singleton A. Whole genome association studies: Deciding when persistence becomes perseveration. Am J Med Genet B Neuropsychiatr Genet 2008; 147B: 131-133.

79

GENETICS OF UNIPOLAR MAJOR DEPRESSIVE DISORDER

29. Brzustowicz LM. Size matters: the unexpected challenge of detecting linkage in large cohorts. Am J Psychiatry 2007; 164: 192-194.

30. Manolio TA, Rodriguez LL, Brooks L, Abecasis G, Ballinger D, Daly M, et al. New models of collaboration in genome-wide association studies: The Genetic Association Information Network. Nat Genet 2007; 39: 1045-1051.

31. Boomsma DI, Willemsen G, Sullivan PF, Heutink P, Meijer P, Sondervan D, et al. Genome-wide association of major depression: description of samples for the GAIN Major Depressive Disorder Study: NTR and NESDA biobank projects. Eur J Hum Genet 2008; 16: 335-342.

32. Levinson DF, Evgrafov OV, Knowles JA, Potash JB, Weissman MM, Scheftner WA, et al. Genetics of recurrent early-onset major depression (GenRED): significant linkage on chromosome 15q25-q26 after fine mapping with single nucleotide polymorphism markers. Am J Psychiatry 2007; 164: 259-264.

33. Verma R, Holmans P, Knowles JA, Grover D, Evgrafov OV, Crowe RR, et al. Linkage disequilibrium mapping of a chromosome 15q25-26 major depression linkage region and sequencing of NTRK3. Biol Psychiatry 2008; 63: 1185-1189.

34. Feng Y, Vetro A, Kiss E, Kapornai K, Daroczi G, Mayer L, et al. Association of the neurotrophic tyrosine kinase receptor 3 (NTRK3) gene and childhood-onset mood disorders. Am J Psychiatry 2008; 165: 610-616.

35. Duman RS, Heninger GR, Nestler EJ. A molecular and cellular theory of depression. Arch Gen Psychiatry 1997; 54: 597-606.

36. Duman RS. Depression: A case of neuronal life and death?

Biol Psychiatry 2004; 56: 140-145.

37. Malberg JE, Eisch AJ, Nestler EJ, Duman RS. Chronic antidepressant treatment increases neurogenesis in adult rat hippocampus. J Neurosci 2000; 20: 9104-9110.

38. Lopez-Leon S, Janssens AC, Gonzalez-Zuloeta Ladd AM, Del-Favero J, Claes SJ, Oostra BA, et al. Meta-analyses of genetic studies on major depressive disorder. Mol Psychiatry 2008; 13: 772-785.

39. Levinson DF. The genetics of depression: A review. Biol Psychiatry 2006; 60: 84-92.

40. Heils A, Teufel A, Petri S, Stober G, Riederer P, Bengel D, et al. Allelic variation of human serotonin transporter gene expression. J Neurochem 1996; 66: 2621-2624.

41. Hu XZ, Lipsky RH, Zhu G, Akhtar LA, Taubman J, Greenberg BD, et al. Serotonin transporter promoter gainof-function genotypes are linked to obsessive-compulsive disorder. Am J Hum Genet 2006; 78: 815-826.

42. Kraft JB, Slager SL, McGrath PJ, Hamilton SP. Sequence analysis of the serotonin transporter and associations with 80

antidepressant response. Biol Psychiatry 2005; 58: 374-381.

43. Wendland JR, Moya PR, Kruse MR, Ren-Patterson RF, Jensen CL, Timpano KR, et al. A novel, putative gain-offunction haplotype at SLC6A4 associates with obsessivecompulsive disorder. Hum Mol Genet 2008; 17: 717-723.

44. Hu XZ, Rush AJ, Charney D, Wilson AF, Sorant AJ, Papanicolaou GJ, et al. Association between a functional serotonin transporter promoter polymorphism and citalopram treatment in adult outpatients with major depression. Arch Gen Psychiatry 2007; 64: 783-792.

45. Kato M, Serretti A. Review and meta-analysis of antidepressant pharmacogenetic findings in major depressive disorder.

Mol Psychiatry 2008; Nov. 4. [Epub ahead of print] 46. Li D, He L. Meta-analysis supports association between serotonin transporter (5-HTT) and suicidal behavior. Mol Psychiatry 2007; 12: 47-54.

47. Munafo MR, Clark T, Flint J. Does measurement instrument moderate the association between the serotonin transporter gene and anxiety-related personality traits? A meta-analysis. Mol Psychiatry 2005; 10: 415-419.

48. Sen S, Burmeister M, Ghosh D. Meta-analysis of the association between a serotonin transporter promoter polymorphism (5-HTTLPR) and anxiety-related personality traits. Am J Med Genet B Neuropsychiatr Genet 2004; 127B: 85-89.

49. Willis-Owen SA, Turri MG, Munafo MR, Surtees PG, Wainwright NW, Brixey RD, et al. The serotonin transporter length polymorphism, neuroticism, and depression: A comprehensive assessment of association. Biol Psychiatry 2005; 58: 451-456.

50. Munafo MR, Freimer NB, Ng W, Ophoff R, Veijola J, Miettunen J, et al. 5-HTTLPR genotype and anxiety-related personality traits: A meta-analysis and new data. Am J Med Genet B Neuropsychiatr Genet 2008; 150B:271-281.

51. Kendler KS, Karkowski LM, Prescott CA. Causal relationship between stressful life events and the onset of major depression. Am J Psychiatry 1999; 156: 837-841.

52. Keller MC, Neale MC, Kendler KS. Association of different adverse life events with distinct patterns of depressive symptoms. Am J Psychiatry 2007; 164: 1521-1529; quiz 1622.

53. Caspi A, Sugden K, Moffitt TE, Taylor A, Craig IW, Harrington H, et al. Influence of life stress on depression: moderation by a polymorphism in the 5-HTT gene.

Science 2003; 301: 386-389.

54. Munafo MR, Durrant C, Lewis G, Flint J. Gene x environment interactions at the serotonin transporter locus. Biol Psychiatry 2009; 65:211-219.

55. Risch N, Herrell R, Lehner T, Liang KY, Eaves L, Hoh J, et al. Interaction between the serotonin transporter gene

Tanya Goltser-Dubner et al.

(5-HTTLPR), stressful life events, and risk of depression: A meta-analysis. JAMA 2009; 301: 2462-2471.

56. Moffitt TE, Caspi A, Rutter M. Strategy for investigating interactions between measured genes and measured environments. Arch Gen Psychiatry 2005; 62: 473-481.

57. Carola V, Frazzetto G, Pascucci T, Audero E, PuglisiAllegra S, Cabib S, et al. Identifying molecular substrates in a mouse model of the serotonin transporter x environment risk factor for anxiety and depression. Biol Psychiatry 2008; 63: 840-846.

58. Rietschel M, Beckmann L, Strohmaier J, Georgi A, Karpushova A, Schirmbeck F, et al. G72 and its association with major depression and neuroticism in large population-based groups from Germany. Am J Psychiatry 2008; 165: 753-762.

59. Barden N, Harvey M, Gagne B, Shink E, Tremblay M, Raymond C, et al. Analysis of single nucleotide polymorphisms in genes in the chromosome 12Q24.31 region points to P2RX7 as a susceptibility gene to bipolar affective disorder. Am J Med Genet B Neuropsychiatr Genet 2006; 141B: 374-382.

60. Lucae S, Salyakina D, Barden N, Harvey M, Gagne B, Labbe M, et al. P2RX7, a gene coding for a purinergic ligand-gated ion channel, is associated with major depressive disorder. Hum Mol Genet 2006; 15: 2438-2445.

61. Wong ML, Dong C, Maestre-Mesa J, Licinio J. Polymorphisms in inflammation-related genes are associated with susceptibility to major depression and antidepressant response. Mol Psychiatry 2008; 13: 800-812.

62. Wong M-L, Whelan F, Deloukas P, Whittaker P, Delgado M, Cantor RM, et al. Phosphodiesterase genes are associated with susceptibility to major depression and antidepressant treatment response. Proc Nat Acad Sci 2006; 103: 15124-15129.

63. Warner-Schmidt JL, Duman RS. Hippocampal neurogenesis: Opposing effects of stress and antidepressant treatment. Hippocampus 2006; 16: 239-249.

64. Szeszko PR, Lipsky R, Mentschel C, Robinson D, GunduzBruce H, Sevy S, et al. Brain-derived neurotrophic factor val66met polymorphism and volume of the hippocampal formation. Mol Psychiatry 2005; 10: 631-636.

65. Bueller JA, Aftab M, Sen S, Gomez-Hassan D, Burmeister M, Zubieta JK. BDNF Val66Met allele is associated with reduced hippocampal volume in healthy subjects. Biol Psychiatry 2006; 59: 812-815.

66. Frodl T, Schule C, Schmitt G, Born C, Baghai T, Zill P, et al. Association of the brain-derived neurotrophic factor Val66Met polymorphism with reduced hippocampal volumes in major depression. Arch Gen Psychiatry 2007; 64: 410-416.

67. Blumberg HP, Wang F, Chepenik LG, Kalmar JH, Edmiston E, Duman RS, Gelernter J. Influence of vascular endothelial growth factor variation on human hippocampus morphology. Biol Psychiatry 2008; 64: 901-903.

68. Hariri AR, Mattay VS, Tessitore A, Kolachana B, Fera F, Goldman D, et al. Serotonin transporter genetic variation and the response of the human amygdala. Science 2002; 297: 400-403.

69. Munafo MR, Brown SM, Hariri AR. Serotonin transporter (5-HTTLPR) genotype and amygdala activation: A metaanalysis. Biol Psychiatry 2008; 63: 852-857.

70. Levinson DF. Meta-analysis in psychiatric genetics. Curr Psychiatry Rep 2005; 7: 143-151.

71. Martin J, Cleak J, Willis-Owen SA, Flint J, Shifman S. Mapping regulatory variants for the serotonin transporter gene based on allelic expression imbalance. Mol Psychiatry 2007; 12: 421-422.

72. Lazary J, Lazary A, Gonda X, Benko A, Molnar E, Juhasz G, et al. New evidence for the association of the serotonin transporter gene (SLC6A4) haplotypes, threatening life events, and depressive phenotype. Biol Psychiatry 2008; 64: 498-504.

73. Wichers M, Kenis G, Jacobs N, Mengelers R, Derom C, Vlietinck R, et al. The BDNF Val(66)Met x 5-HTTLPR x child adversity interaction and depressive symptoms: An attempt at replication. Am J Med Genet B Neuropsychiatr Genet 2008; 147B: 120-123.

74. Pezawas L, Meyer-Lindenberg A, Goldman AL, Verchinski BA, Chen G, Kolachana BS, et al. Evidence of biologic epistasis between BDNF and SLC6A4 and implications for depression. Mol Psychiatry 2008; 13: 709-716.

75. Neff CD, Abkevich V, Packer JC, Chen Y, Potter J, Riley R, et al. Evidence for HTR1A and LHPP as interacting genetic risk factors in major depression. Mol Psychiatry 2009; 14: 621-630.

76. Kaufman J, Yang BZ, Douglas-Palumberi H, Grasso D, Lipschitz D, Houshyar S, et al. Brain-derived neurotrophic factor-5-HTTLPR gene interactions and environmental modifiers of depression in children. Biol Psychiatry 2006; 59: 673-680.

77. OážżDonovan MC, Craddock N, Norton N, Williams H, Peirce T, Moskvina V, et al. Identification of loci associated with schizophrenia by genome-wide association and follow-up. Nat Genet 2008; 40:1053-1055.

78. Allen NC, Bagade S, McQueen MB, Ioannidis JP, Kavvoura FK, Khoury MJ, et al. Systematic meta-analyses and field synopsis of genetic association studies in schizophrenia: The SzGene database. Nat Genet 2008; 40: 827-834.

79. OážżDonovan MC, Norton N, Williams H, Peirce T, Moskvina V, Nikolov I, et al. Analysis of 10 independent samples provides evidence for association between schizophrenia and 81

GENETICS OF UNIPOLAR MAJOR DEPRESSIVE DISORDER

a SNP flanking fibroblast growth factor receptor 2. Mol Psychiatry 2009; 14: 30-36.

86. Pearson TA, Manolio TA. How to interpret a genome-wide association study. JAMA 2008; 299: 1335-1344.

80. Zeggini E, Scott LJ, Saxena R, Voight BF, Marchini JL, Hu T, et al. Meta-analysis of genome-wide association data and large-scale replication identifies additional susceptibility loci for type 2 diabetes. Nat Genet 2008; 40: 638-645.

87. Ioannidis JP, Ntzani EE, Trikalinos TA, ContopoulosIoannidis DG. Replication validity of genetic association studies. Nat Genet 2001; 29: 306-309.

81. Frayling TM, McCarthy MI. Genetic studies of diabetes following the advent of the genome-wide association study: where do we go from here? Diabetologia 2007; 50: 2229-2233.

82. Barton A, Thomson W, Ke X, Eyre S, Hinks A, Bowes J, et al. Re-evaluation of putative rheumatoid arthritis susceptibility genes in the post-genome wide association study era and hypothesis of a key pathway underlying susceptibility.

Hum Mol Genet 2008; 17: 2274-2279.

83. The International Schizophrenia Consortium. Rare chromosomal deletions and duplications increase risk of schizophrenia. Nature 2008; 455: 237-241 (letter).

88. Lohmueller KE, Pearce CL, Pike M, Lander ES, Hirschhorn JN. Meta-analysis of genetic association studies supports a contribution of common variants to susceptibility to common disease. Nat Genet 2003; 33: 177-182.

89. Chanock SJ, Manolio T, Boehnke M, Boerwinkle E, Hunter DJ, Thomas G, et al. Replicating genotype-phenotype associations. Nature 2007; 447: 655-660.

90. Ioannidis JP, Boffetta P, Little J, OážżBrien TR, Uitterlinden AG, Vineis P, et al. Assessment of cumulative evidence on genetic associations: Interim guidelines. Int J Epidemiol 2008; 37: 120-132.

84. Stefansson H, Rujescu D, Cichon S, Pietilainen OP, Ingason A, Steinberg S, et al. Large recurrent microdeletions associated with schizophrenia. Nature 2008; 455: 232-236.

91. Moonesinghe R, Khoury MJ, Liu T, Ioannidis JP. Required sample size and nonreplicability thresholds for heterogeneous genetic associations. Proc Natl Acad Sci U S A 2008; 105: 617-622.

85. Millar JK, Pickard BS, Mackie S, James R, Christie S, Buchanan SR, et al. DISC1 and PDE4B are interacting genetic factors in schizophrenia that regulate cAMP signaling. Science 2005; 310: 1187-1191.

92. Burton PR, Hansell AL, Fortier I, Manolio TA, Khoury MJ, Little J, et al. Size matters: Just how big is BIG?: Quantifying realistic sample size requirements for human genome epidemiology. Int J Epidemiol 2008; 38:263-273.

82

Jonathan Benjamin

commentary: Etiological hypotheses of mental disorders at the molecular level may not help psychiatry Jonathan Benjamin, MD, MHA “Clalit” Sick Fund, Israel, Professor Emeritus, Ben-Gurion University, Beersheba, Israel

This issue of the “Israel Journal of Psychiatry” is devoted to etiological hypotheses of mental disorders at the molecular level. I appreciate the invitation to write a brief commentary in support of my view that as of 2008 such hypotheses seem less likely to be helpful than they were expected to be 50 years ago.

The short explanation for this is that they have not been helpful yet, so why should we expect a change? We in fact know no more about the etiology of schizophrenia, depression or borderline personality disorder today than we did in 1958. We knew then, as now, that all the disorders we study and treat have a genetic component, and that psychoses and neuroses are helped but not cured by drugs affecting dopamine in the first instance, and noradrenaline and/or serotonin in the second. The success of drugs initially discovered by chance, and the demonstration of some of their actions (but this is not the same as the demonstration of the mechanism of action that treats the disorder), led to a series of molecular hypotheses which began at the level of messenger (neurotransmitter) and receptor, and then progressed to the level of second-messenger and onward into the interior of the cell (growth factors, response elements, nuclear receptors). This is a logical path to follow for a biochemist or cell biologist attempting to elucidate the working of the signal that begins at the synapse with a single quantum of messenger, and then devolves inward as an increasingly complex cascade of molecular events and adaptations (e.g., 1). But the quest for scientific knowledge has not yet helped the doctor understand the disease, let alone treat the patient. Virtually every piece of knowledge painstakingly learned in this way has been

E-mail:

offered as a tentative explanation for an aspect of mental illness; none has survived. The oldest and most famous hypothesis, that every medical student and practitioner still instinctively feels is “right,” the mono-amine hypothesis of depression (2, 3), suggests that depression is a disease of too little serotonin and/or noradrenaline.

But tianeptine, an effective antidepressant available in Europe, is a selective serotonin reuptake enhancer, i.e., lowers the levels of synaptic serotonin (4).

The last five decades have seen a partial retreat from dynamic psychotherapy, and growing enthusiasm for cognitive-behavioral therapy, as cost-effective treatments for some mild but common mental disorders (and for some other problems not really considered “disorders”). There has been no parallel development in biological psychiatry, only the discovery of “me-too” drugs with arguably fewer side effects than those that were known in 1958. None of these new drugs were developed using new etiological hypotheses, nor indeed any etiological hypotheses; all were extensions of drugs previously discovered by chance. Deliberate attempts to treat schizophrenia by glycine agonists (5), and depression with corticotropin-releasing factor (CRF) receptor antagonists (6), both based on etiological hypotheses, have not led to breakthroughs.

The success of the genetic study of a few single-gene diseases in other areas of medicine prompted the search for linkages and then genes for mental illnesses. This is a logical path to follow for a geneticist; given the demonstration that a phenotype is heritable, scan every chromosome for linkage (impossible in 1958 but possible now), and then focus on the areas of linkage until you

zola@netvision.net.il

83

Isr J Psychiatry Relat Sci - Vol 47 - No. 1 (2010)

find (a) mutation(s) or polymorphism(s) responsible for the phenotype. Virtually every chromosome has been implicated in the etiology of serious mental disorders like schizophrenia, no linkage has been consistently replicated (7), and no genes that can reliably be said to cause (or account for a major part of) a mental disorder have been found. A report in “Science” described a linkage to schizophrenia on chromosome 1 with chances of less than one in a million of a false positive (8); this too did not replicate (e.g., 9).

The situation is similar in bipolar disorder. Here too promising genes continue to be reported (10), but we do not have a genetic understanding of the illness.

Researchers tell us that this is partly because many genes are responsible for major mental disorders, each gene of very small effect. But this claim does not accord with mathematical models of the genetics of schizophrenia (11), for example, nor with our acquaintance with the risk of schizophrenia in identical twins, other sibs, and second-degree relatives (around 45%, 10% and 3% respectively). Three or four genes for schizophrenia are more compatible with the evidence. Depression, which seems to have a far bigger environmental component in its etiology (12), looks like an even less promising candidate for a molecular-genetic explanation.

The “British Journal of Psychiatry” recently convened a debate with the chilling title “Research into putative biological mechanisms of mental disorders has been of no value to psychiatry.” In a paragraph addressing etiology, even the scholar chosen to refute the thesis made no attempt to offer examples of insights from molecular science, and wrote instead, “Biology of course does not just mean drugs or genes: Freud … considered himself a biologist” (13). One of the guest editors of the current issue of the “Israel Journal of Psychiatry,” reviewing major depression for the “New England Journal of Medicine,” recently wrote, “Depression is a … disorder … with … no established mechanism” (12). This is as good a summary of the current situation as we are likely to get.

I said above that the failure of molecular hypotheses in psychiatry thus far is the “short” explanation for my concern that they may also fail in the future. There is also an explanation of another kind. Mental disorders are extremely complex sets of phenomena, and the path from molecules, which undoubtedly play an important role, to mental disorders must be long and tortuous. A single example can suffice: alcohol is a simple molecule, with familiar effects on subjective states and behavior.

Yet these vary tremendously across time and individu84

als. Nevertheless, inhibition of social restraint and of fine motor control and of wakefulness may all be plausibly explained by alcohol’s effects on the benzodiazepineGABA receptor complex. When we come to the mental disorders associated with alcohol use, which range from withdrawal delirium to persistent dementia to paranoid jealousy, a plausible molecular hypothesis to account for these seems more remote, even though the “responsible” molecule is already known.

Of course, nothing in this brief commentary can convince us that a true breakthrough is not just around the corner. Too much pessimism can paralyze science, and it then fulfills its own prophecy. Society owes it to itself to continue to fund the study of promising hypotheses, and to reward fruitful research. Nevertheless, the experience of the last 50 years in psychiatry suggests that hopeful prognostications on grant applications, in scientific journals, and in the popular media, should be swallowed in small doses, and washed down with a large measure of thoughtful reflection.

References 1. Duman RS, Heninger GR, Nestler EJ. A molecular and cellular theory of depression. Arch Gen Psychiatry 1997; 54:597-606.

2. Schildkraut J. The catecholamine hypothesis of affective disorders. A review of supporting evidence. Int J Psychiatry 1967; 4:203-217.

3. Murphy DL, Andrews AM, Wichems CH, Li Q, Tohda M, Greenberg B. Brain serotonin neurotransmission: An overview and update with an emphasis on serotonin subsystem heterogeneity, multiple receptors, interactions with other neurotransmitter systems, and consequent implications for understanding the actions of serotonergic drugs.

J Clin Psychiatry 1998; 59:4-12.

4. Kasper S, McEwen BS. Neurobiological and clinical effects of the antidepressant tianeptine. CNS Drugs 2008; 22:15-26.

5. Buchanan RW, Javitt DC, Marder SR, Schooler NR, Gold JM, McMahon RP, Heresco-Levy U, Carpenter WT. The Cognitive and Negative Symptoms in Schizophrenia Trial (CONSIST): The efficacy of glutamatergic agents for negative symptoms and cognitive impairments. Am J Psychiatry 2007; 164:1593-1602.

6. Holsboer F, Ising M. Central CRH system in depression and anxiety - Evidence from clinical studies with CRH(1) receptor antagonists. Eur J Pharmacol 2008; 583:350-357.

7. DeLisi LE, Fleischhaker W. Schizophrenia research in the era of the genome, 2007. Curr Opin Psychiatry 2007; 20:109-110.

Jonathan Benjamin

8. Brzustowicz LM, Hodgkinson KA, Chow EW, Honer WG, Bassett AS. Location of a major susceptibility locus for familial schizophrenia on chromosome 1q21-q22. Science 2000; 288:678-682.

9. Levinson DF, Holmans PA, Laurent C, Riley B, Pulver AE, Gejman PV, Schwab SG, Williams NM, Owen MJ, Wildenauer DB, Sanders AR, Nestadt G, Mowry BJ, Wormley B, Bauché S, Soubigou S, Ribble R, Nertney DA, Liang KY, Martinolich L, Maier W, Norton N, Williams H, Albus M, Carpenter EB, DeMarchi N, Ewen-White KR, Walsh D, Jay M, Deleuze JF, O'Neill FA, Papadimitriou G, Weilbaecher A, Lerer B, O'Donovan MC, Dikeos D, Silverman JM, Kendler KS, Mallet J, Crowe RR, Walters M. No major schizophrenia locus detected on chromo-

Book reviews History of the Introduction of Lithium into Medicine and Psychiatry - Birth of Modern Psychopharmacology 1949 J. Schioldann, MD; Preface by G.E. Berrios, MD Adelaide Academic Press, 363 pages ISBN: 978-0-9805477-0-2

A

s medical professionals we have come to accept and rely on lithium therapy as one of the essential tools of modern psychiatry; however not many among us have tried to delve deeper into the history behind the emergence and acceptance of this therapy. It is widely accepted that the rediscovery of lithium's antimanic properties by John F. Cade has helped to establish the field of modern psychopharmacology. Therefore, a deeper understanding of the events which have led to this discovery will help us to better understand one of the forming events on which the modern psychiatric profession is based.

Professor Johan Schioldann, of the University of Adelaide, reveals a fascinating chapter of the early history of modern psychopharmacology. His meticulous study, often through the use of original sources which have not been researched before, tells the history of lithium therapy from the mid-19th century to John Cade's discovery, during the 1940s, of lithium’s effects on patients with mood disorders. The tale behind the origins of John Cade's ideas provides us with further insight into the field

some 1q in a large multicenter sample. Science 2002; 296:739-741.

10. Serretti A, Mandelli L. The genetics of bipolar disorder: Genome “hot regions,” genes, new potential candidates and future directions. Mol Psychiatry 2008; 13:742-771.

11. Risch N. Linkage strategies for genetically complex traits.

I. Multilocus models. Am J Hum Genet 1990; 46:222-228.

12. Belmaker RH, Agam G. Major depressive disorder. N Eng J Med 2008; 55-68.

13. Kingdon D, Young AH. Research into putative biological mechanisms of mental disorders has been of no value to psychiatry. Br J Psychiatry 2007; 191:285–290.

of modern psychopharmacology, which is one of the cornerstones of contemporary psychiatry.

The book is written in a style which makes it equally appealing to medical professionals and to laypeople interested in psychiatry, psychopharmacology, or the history of medicine. It is extremely well referenced and presents us with several original documents which allow us a unique perspective of the events which are being described. Although the work touches on several issues which are quite controversial from a nationalistic point of view, this is done with utmost regard for national sensitivities and respect for the accepted historical narrative.

The first part of the book gives an account of the early history of lithium therapy from its earliest applications as a gout remedy, within the context of the then accepted correlation between gout and mental illness, to the first documented uses of lithium in the treatment of mania by Carl and Fritz Lange in the 1850s.

The second part of the book gives a most detailed and meticulously researched account of John Cade's renewed discovery of the therapeutic properties of lithium and subsequent medical trials, and the controversies concerning the early uses of lithium in the treatment of manic disorders, as well as painting a portrait of the man himself both through his research and through accounts of his contemporaries.

This work is highly recommended for all who wish to obtain a deeper and more encompassing comprehension of the field of modern psychiatry and more particularly of modern psychopharmacology.

Igor Plopski, Bat Yam

85

Isr J Psychiatry Relat Sci - Vol 47 - No. 1 (2010)

Reelin Glycoprotein: Structure, Biology and Roles in Health and Disease

Generalized Anxiety Disorder Across the Lifespan - An Integrative Approach

S. Hossein Fatemi, editor Springer, 2008, 443 pages

Michael E. Portman Springer, 2009, Hardcover, pp. 182.

ISBN 978-0-387-89242-9

"

Reelin glycoprotein is a major secretory protein with an important role in embryogenesis and during adult life," starts S. Hossein Fatemi in the preface to his book on reelin, a truly masterful rollercoaster for the basic scientist, mainly on this molecule that has a deep impact on brain health and disease.

The book takes the reader through the chemistry, anatomy and molecular functions of reelin to its influence upon cognition, liver function and odontogenesis, proving it is a still much neglected but nevertheless essential molecule. It describes the homo and heterozygous mouse mutant to reelin, giving us, thus, the basic understanding for its major role. It relates to its function and lack in brain disease like lissencephaly, Alzheimer's disease, autistic disorder, stroke, schizophrenia and bipolar disorder. For the clinician the book starts at Chapter 21 and ends at Chapter 27, about 100 pages on reelin causing the smooth (agyric) brain of lissencephaly or the synaptogenesis deficit postulated to be the basis of schizophrenia and its role in synaptic plasticity crucial for cognitive abilities.

More so, it introduces the concept of impaired reelin/ApoE receptor dependent neuromodulation that might contribute to synaptic loss and cognitive deficit in Alzheimer's disease.

Graphically the book is exceptionally attractive, having a large number of colored and black and white figures and graphs. Didactically organized and chaptered it deals for its better part, two-thirds on basic science and a last third of preclinical studies.

I would warmly recommend it to the basic scientist on his way to understanding the "molecular brain" â&#x20AC;&#x201C; a vivacious addition to tau and Apo E, in an era where we look at human brain diseases through mutant mice and mutated molecules. I would less warmly recommend it to the brain clinician as it relates only vaguely to therapeutic options for brain disease in Chapters 23-24.

Overall a mighty effort for what will prove to be a new significant molecule, or not.

eneralized Anxiety Disorder (GAD) is a mental disorder characterized by excessive worry and anxiety that is difficult for the person to control.

GAD causes significant disruption in the patient's life.

Approximately 3% of people may develop the disorder during any given year, and up to 5% of people will suffer from GAD at some point in their lives.

Michael Portman's book on GAD is written for practicing clinicians, psychologists, social workers, researchers and even interested laymen. In a concise, but comprehensive form, the author reviews the major theories regarding GAD, as well as providing a comprehensive and balanced overview of the most recent research on the subject. In addition we are presented with a description of the main strategies and methodologies for treating this disabling and under-recognized condition.

The author, Michael Portman, DPhil, LISW-S, avails us with his extensive clinical experience, gained from his work as a clinical social worker and his private practice specializing in anxiety disorders, and his widely ranging knowledge of both the professional and empirical literature pertaining to the understanding and treatment of GAD and its definition as a separate disorder.

The book has eight chapters covering a wide range of subjects from the definition, history and the nature of GAD, the most current diagnostic tools, the better founded conceptual models regarding GAD, the various therapeutic approaches (both psychosocial and pharmacological), the specifics of treating various special needs groups such as minors and older adults, some less common issues, such as prevention and culture specific symptom clusters and finally the direction of future developments in the field of GAD. The book is both user-friendly and informative. The writing is clear and the review is up to date. Both professionals and laypeople interested in the subject may benefit from reading this book.

Diana Paleacu, Bat Yam

Shelly Tadger, Bat Yam

86

G

‫ישיבת ועד האיגוד הפסיכיאטרי ‪-‬‬ ‫כנס ים המלח‪ ,‬ינואר ‪2010‬‬ ‫‪ .1‬האיגוד אימץ את עקרונות ההתערבות לטיפול בהפרעת‬ ‫דחק בתר־חבלתית כפי שגובשו על ידי הקונסורציום הלאומי‬ ‫והוצגו על ידו‪ .‬האיגוד רואה את ההצעה כדינמית‪ ,‬מתפתחת‬ ‫ומשתנה לאור הספרות המקצועית והטכנולוגיות החדשות‪.‬‬ ‫תועבר תוספת להשלמת הפרק העוסק בהתערבות בטראומה‬ ‫של ניצולי שואה‪.‬‬ ‫‪ .2‬לקראת הכנס הקרוב בנושא‪" ,‬תרופות טיפיות או א־טיפיות?"‬ ‫מונתה ועדה לגיבוש מתכונת הכנס ורשימת המשתתפים‪ .‬בסיום‬ ‫הדיונים יגובש מסמך מקצועי אשר יסכם את התהליך והעמדות‬ ‫שיוצגו בו‪.‬‬ ‫‪ .3‬מונתה ועדה מקצועית שתרכז את נושא "הפסיכותרפיה‬ ‫באיגוד"‪ .‬תגובש הצעה לאישור המליאה לגבי תחומי העיסוק‬ ‫והיעדים לפעילותה‪.‬‬ ‫‪ .4‬האיגוד רואה חשיבות בהרחבת ההתנסות וגוף הידע בתחום‬ ‫הפסיכוגריאטריה אצל המתמחים‪ ,‬וכן יגדיל את היקף הנושא‬ ‫בהרכב הבחינה‪ .‬ועדת ההתמחות תבדוק את משמעות חיוב‬ ‫הרוטציה במחלקות הפסיכו־גריאטריות במסלול ההתמחות‪.‬‬ ‫‪ .5‬האיגוד רואה חשיבות להרחבת העיסוק בתחום "הפסיכיאטריה‬ ‫החיובית"‪ .‬הוקמה ועדה שתגבש את הפעילות הנדרשת‪ ,‬תפיץ‬ ‫חומר רקע לחברים ותציע סדר יום לקידום הנושא‪.‬‬

‫‏‪ 10‬בינואר ‪2010‬‬

‫‪ .6‬מונה צוות מוביל אשר יגבש קווים אדומים בהתקיים‬ ‫הרפורמה הביטוחית בבריאות הנפש וההתנהלות באין רפורמה‪,‬‬ ‫דרכים לפיתוח ולהשלמת השירותים הקהילתיים ולהתאמת‬ ‫מספר מיטות האשפוז הפסיכיאטריות‪ .‬נייר העקרונות שיגובש‬ ‫יוצג בפני גורמי משרד הבריאות העוסקים בנושא ויועציו‪.‬‬ ‫‪ .7‬הוועד המנהל באיגוד יבדוק דרכים להפעלת הגב' צופית‬ ‫גרנט כדוברת ויועצת התקשורת של האיגוד‪ ,‬ואת המשמעויות‬ ‫להעסקת חברת ייעוץ ויחסי ציבור חיצונית‪.‬‬ ‫‪ .8‬האיגוד ייזום יום עיון ארצי לסטודנטים לרפואה בשנה ו'‬ ‫במסגרת העלאת המודעות למקצוע‪ ,‬שיפור מעמד הפסיכיאטר‬ ‫ומשיכת מתמחים מצטיינים לתחום‪.‬‬ ‫‪ .9‬יום העיון השנתי של האיגוד יאורגן על ידי ועד סניף‬ ‫ירושלים‪ ,‬לזכרו של ד"ר קרלוס בראל ז"ל‪.‬‬ ‫‪ .10‬ברכות לחברנו ד"ר הירשמן‪ ,‬על כניסתו לתפקיד ראש‬ ‫שירותי בריאות הנפש במשרד הבריאות עד לקיום המכרז‪ .‬עם‬ ‫סיום תפקידו במשרד הבריאות יחזור לפעילות מלאה באיגוד‪.‬‬ ‫ ‬ ‫ד"ר נמרוד גריסרו‬ ‫ ‬ ‫מזכיר‬

‫פרופ' זאב קפלן‬ ‫יו"ר‬

‫איגוד הפסיכיאטריה בישראל‬ ‫ההסתדרות הרפואית ‪ -‬המועצה המדעית‬

‫‪Israeli Psychiatric Association‬‬

‫יו"ר‪ :‬פרופ' זאב קפלן ‪President: Prof. Z. Kaplan /‬‬ ‫‪Zeev.kaplan@pbsh.health.gov.il‬‬ ‫מזכיר‪ :‬ד"ר נמרוד גריסרו ‪President: Dr. N. Grisaru /‬‬ ‫‪grisarun@gmail.com‬‬ ‫יו"ר נבחר‪ :‬פרופ' משה קוטלר ‪Elected President: Prof. M. Kotler /‬‬ ‫‪Moshe.kotler@beerness.health.gov.il‬‬

‫יו"ר יוצא ואחראי קשרי חו"ל‪ :‬פרופ' אבי בלייך ‪/‬‬ ‫‪President Emeritus and Foreign Affairs: Prof. A. Bleich‬‬ ‫‪lean@bgu.ac.il‬‬

‫ ‬

‫‪ableich@lev-hasharon.co.il‬‬

‫‪87‬‬

‫המרכז לבריאות הנפש באר שבע‬ ‫‪Beer-Sheva Mental Health Center‬‬

‫ טל'‪ ;08-6401606 :‬פקס‪08-6401621 :‬‬ ‫ רח' הצדיק מירושלים ‪ ,2‬באר שבע‪ ,‬ת"ד ‪ 4600‬‬ ‫ ‪Hazadik from Jerusalem St. P.O. Box 4600‬‬ ‫ ‪www.psychiatry.org.il‬‬

‫באדנוזין דרך רצפטורי אדנוזין מסוג ‪ ,A2A‬המבקרים ישירות‬ ‫סימנים התנהגותיים של דיכאון‪ .‬צפיפות גבוהה של רצפטורים‬ ‫לאדנוזין נמצאת בעצבי ה–‪ GABA‬במסלול ה–‪Striopallidal‬‬ ‫בסטריאטום ומרכיבים חלק מקומפלקס הרצפטורים‬ ‫‪ .A2A/D2/mGLU‬הפעלה של רצפטורי ‪ A2A‬הללו מפרידה‬ ‫פונקציונלית בין ההשפעה של עירור המוטיבציה תלוית‬ ‫דופאמין לבין ההתנהגות המתמשכת מסוג ‪conservation-‬‬ ‫‪ ,withdrawal‬המובילה להתנהגות נסיגה‪ .‬אנטגוניזם של‬ ‫רצפטורי ‪ A2A‬באזור הוונטראלי–מדיאלי של הסטריאטום‬ ‫בחולדות שטופלו ברזרפין מקל סימפטומים של התנהגות‬ ‫דיכאונית‪.‬‬

‫הגנטיקה של מחלת הדיכאון המג'ורי החד־קוטבי‬ ‫כדוגמה לגנטיקה בהפרעות מורכבות‬ ‫ט' גולצר־דובנר‪ ,‬א' גלילי־וייסטוב‪ ,‬ר' ה' סגמן‪ ,‬ירושלים‬

‫הפרעת הדיכאון המג'ורי היא הטרוגנית‪ ,‬בעלת שכיחות גבוהה‬ ‫ובמידה מסוימת הינה הפרעה תורשתית‪ .‬שילוב של פעולה‬ ‫ותגובה הדדית בין גנים לסביבה מפריד מתוך כלל אלו שחווים‬ ‫סוגי עצב שונים‪ ,‬קצרים יותר והדירים ומשתקים פחות‪ ,‬קבוצה‬ ‫קטנה אך משמעותית שעלולה לפתח את מחלת הדיכאון‪.‬‬ ‫ההתקדמות הטכנולוגית האדירה בפענוח רצפי ייחוס של הגנום‬ ‫האנושי ווריאציות שכיחות בגנום מאפשרת מחקרים גנטיים‬

‫בקנה מידה חסר תקדים מבחינת צפיפות הגנום הממופה וגודל‬ ‫אוכלוסיות המדגם בעלויות סבירות יחסית‪ .‬המאמץ המחקרי‬ ‫הזה נמצא בשלבים הראשונים לקראת חשיפת הארכיטקטורה‬ ‫הגנטית של כמה פנוטיפים מורכבים‪.‬‬ ‫למרות ההתחלה המקרטעת‪ ,‬המחקר הגנטי של מחלת הדיכאון‬ ‫מבשיל ממחקר אסוציאציה צנוע–ממדים של גנים נבחרים‬ ‫למחקרי לינקנג' במשפחות‪ ,‬ויאפשר בקרוב מחקרי אסוציאציה‬ ‫מבוקרים למלוא אורכו של הגנום‪ .‬חוזק ההשפעה (‪effect‬‬ ‫‪ )size‬של וריאנטים גנטיים שהתגלו עד כה‪ ,‬הקשורים לסיכון‬ ‫לפתח דיכאון ואוששו במחקרים שונים‪ ,‬הינו צנוע‪ ,‬אך עם זאת‬ ‫נדמה שהוא תורם לביטויו של הפנוטיפ כחלק ממערך מורכב‬ ‫של גורמים אפיגנטיים ואפיסטטיים התלויים זה בזה‪ .‬גילוי של‬ ‫גנים נוספים שתורמים לסיכון לפתח דיכאון‪ ,‬שיחשפו מנגנונים‬ ‫מולקולריים הקשורים ברגישות למחלה‪ ,‬הוא הכרחי לפיתוח‬ ‫תרופות אנטי–דיכאוניות בעלות מנגנון פעולה שונה מאלה‬ ‫הקיימות שמווסתות העברה מונואמינרגית‪.‬‬ ‫מאמר הסקירה הזה משרטט תפישות בסיסיות ומעדכן את‬ ‫ההתקדמות שנעשתה במחקר‪ .‬כמו כן הוא סוקר בקצרה גנים‬ ‫מרכזיים בעלי הדירות גבוהה‪ ,‬שנבחרו מפני היותם תואמים‬ ‫להיפותזות הביולוגיות הקיימות לדיכאון‪ ,‬ומסתיים בדיון‬ ‫בכיווני המחקר העכשוויים שכוללים שיקולי פנוטיפים‪ ,‬גודל‬ ‫מדגם וקידומם של מחקרים שיטתיים‪ ,‬שמלבד גילוי הווריאנטים‬ ‫הגנטיים‪ ,‬יחקרו גם את משמעותם הפונקציונלית‪.‬‬

‫‪88‬‬

‫ובסכיזופרניה‪ .‬גישות טיפוליות המכוונות לשינוי תזמון ומשך‬ ‫השינה‪ ,‬לכוונון מחדש של המחזוריות היומית ולהארכת זמן‬ ‫העירות‪ ,‬כמו טיפול באור או בתרופות המשפיעות על גורמים‬ ‫אלו‪ ,‬נמצאו יעילות בטיפול במחלות מצב הרוח‪.‬‬ ‫מחקרים שבהם ניסו לשפר את השינה והפרעות במחזוריות‬ ‫היומית קיימים אך נדירים‪ ,‬על אף שנמצא כי מתן מלטונין‬ ‫משפר את מהלך השינה ומפחית סימנים פסיכוטיים‪ .‬לאחרונה‬ ‫צבר תאוצה מחקר המנגנונים המולקולריים במחלות פסיכוטיות‬ ‫הקשורים בשעון הביולוגי (‪.)clock‬‬ ‫מחקרים גנטיים מצאו קשר בין מחלת הסכיזופרניה וגנים מקבוצת‬ ‫ה–‪ Clock‬כגון‪ CLOCK ,PER1 ,PER3 :‬ו–‪.TIMELESS‬‬ ‫רוב המחקר במחלה דו–קוטבית התרכז בפולימורפיזם של הגן‬ ‫‪ ,CLOCK‬אבל ייתכן שהגן ‪ GSK–3‬שהוא גן מטרה של ליתיום‪,‬‬ ‫משחק גם הוא תפקיד חשוב בתהליך‪ .‬מחקר מעמיק‪ ,‬שבוחן את‬ ‫התפקיד של תהליכי המחזוריות היומית וגנים מקבוצת ‪Clock‬‬ ‫במחלות נפש‪ ,‬סביר שיביא להתפתחות מואצת של טיפול‬ ‫המבוסס על התערבות בתהליכים אלו‪.‬‬

‫השפעתם של גורמים תוך־רחמיים‬ ‫על התפתחות דיכאון קליני‬ ‫מ' ויינשטוק‪ ,‬ירושלים‬

‫על אף שהגורמים המובילים להתפתחות דיכאון קליני אינם‬ ‫ידועים‪ ,‬התהליך מתרחש בעיקר בפרטים בעלי רגישות יתר‪,‬‬ ‫בעקבות חשיפה למאורעות חיים בעלי אופי שלילי‪ .‬סקירה זו‬ ‫בוחנת את המעורבות של גורמים תוך־רחמיים‪ ,‬אשר מתבטאים‬ ‫כתוצאה מחשיפה להורמוני דחק‪ ,‬ברגישות המוגברת של הפרט‬ ‫להתפתחותו של דיכאון קליני‪ .‬דחק אמהי חמור או צריכת‬ ‫אלכוהול במהלך השליש השני והשלישי להיריון גורמים לעליה‬ ‫מוגברת בשחרור של ‪CRH – Corticotropin Releasing‬‬ ‫‪ Hormone‬ושל ההורמון קורטיזול‪.‬‬ ‫ההורמונים הללו קשורים לירידה במשקל הוולד‪ ,‬פוגעים‬ ‫במנגנוני ההחזר המבקרים את ציר ההיפותלמוס־היפופיזה־‬ ‫אדרנל (‪ )HPA axis‬ובמסלול העצבי המועבר באמצעות‬ ‫הקולטנים לסרוטונין‪ 5-HT1A ,‬ו־‪ ,5-HT2A‬באזורים עיקריים‬ ‫של המוח‪ .‬דחק טרום־לידתי וצריכת אלכוהול בחיות מעבדה‬ ‫גורמים להופעת תסמינים התנהגותיים דמויי דיכאון בחולדות‪,‬‬ ‫וכן לשינויים במחזור השינה ובמחזור הצירקדי‪ ,‬בדומה‬ ‫לתופעות הנצפות בבני אדם הלוקים בדיכאון‪ .‬הטיפול בתרופות‬ ‫נוגדות דיכאון‪ ,‬אשר מוביל לשיפור בתסמיני הדיכאון הקליני‪,‬‬ ‫תורם להחזרת הבקרה התקינה על ציר ה־‪ HPA‬ושל ההעברה‬ ‫העצבית המתווכת על ידי סרוטונין‪.‬‬

‫הביולוגיה של דלדול טריפטופן והקשר למצב הרוח‬ ‫ל' טוקר‪ ,‬ש' עמר‪ ,‬י' ברסודסקי‪ ,‬ג' בנימין‪ ,‬א' קליין‪ ,‬ג' אגם‪ ,‬באר שבע‬

‫קיימות עדויות רבות למעורבות המערכת הסרוטונרגית‬ ‫‪89‬‬

‫במחלות נפש‪ .‬הורדת רמות טריפטופן היא שיטה מוכרת‬ ‫המשמשת לחקר ההשפעה של רמות סרוטונין נמוכות במוח‪,‬‬ ‫אך התוצאות הנצפות אינן חד–משמעיות ולעתים אף מנוגדות‪.‬‬ ‫העבודה הנוכחית סוקרת את הסתירות הללו ודנה בהן תוך‬ ‫שימת דגש על חשיבות הפרטים המתודולוגיים כגון הורדה‬ ‫כרונית מול קצרת טווח של רמות טריפטופן‪ ,‬הדיאגנוזה של‬ ‫החולה ומצב המחלה (מאוזן מול פאזה חריפה) וטיפול תרופתי‬ ‫קודם‪ .‬הורדה קצרת טווח של רמות טריפטופן מוצעת כמבחן‬ ‫חיזוי לטיפול אנטי–דיכאוני מותאם אישית‪.‬‬

‫מטבוליזם של טריפטופן־קינורנין כמתווך משותף‬ ‫של פגיעות גנטיות וסביבתיות בדיכאון מג'ורי‪:‬‬ ‫בחינה מחדש של "השערת הסרוטונין" לאחר ‪ 40‬שנה‬ ‫ג' פ' אוקסנקרוג‪ ,‬בוסטון‪ ,‬ארה"ב‬

‫מאמרנו ב–‪ Lancet 1969‬העלה את ההשערה הבאה‪" :‬בעת‬ ‫דיכאון‪ ,‬מטבוליזם של טריפטופן מוסט ממסלול יצירת סרוטונין‬ ‫לכיוון יצירת קינורנין"‪ .‬הממצא שלפיו לקינורנינים יש פעילות‬ ‫נוירו־טרופית תומך באפשרות שהגברת מסלול הטריפטופן–‬ ‫קינורנין לא רק מגבירה את החסר בסרוטונין אלא גם נוגעת‬ ‫לחרדה הקשורה לדיכאון‪ ,‬לפסיכוזה ולירידה קוגניטיבית‪ .‬מאמר‬ ‫הסקירה הזה מציע כי מסלול הטריפטופן–קינורנין מעורב‬ ‫במנגנונים גנטיים וסביבתיים הקשורים בדיכאון‪.‬‬ ‫אנזימים קובעי קצב בסינתזת קינורנין מופעלים על ידי‬ ‫הורמוני עקה (‪)TDO – Tryptophan 2,3-dioxygenase‬‬ ‫או על ידי ציטוקינים פרו–דלקתיים (‪IDO – Indolamine‬‬ ‫‪ .)2,3-dioxygenase‬הימצאות בו–זמנית של אללים המבטאים‬ ‫ביתר גנים של ציטוקינים פרו–דלקתיים (כגון אינטרפרון‬ ‫גאמא ו–‪ )tumor necrosis factor alpha‬משפיעה על‬ ‫נטייה גנטית לדיכאון על ידי הגברת ה–‪ ,IDO‬בעוד ההשפעה‬ ‫של הסביבה מתווכת על ידי הפעלה הורמונלית של ‪.TDO‬‬ ‫מסלול הטריפטופן–קינורנין מייצג נקודת מפגש מרכזית‬ ‫של אינטראקציה בין גנים וסביבה בדיכאון‪ ,‬ומטרה חדשה‬ ‫להתערבות פרמקולוגית במחלה‪.‬‬ ‫חפיפה בין מרכיבים ביוכימיים ואנטומיים בדיכאון‬ ‫ובהתנהגות במצב של חולי‬ ‫ט' ס' הנף‪ ,‬ס' ג' פורסט‪ ,‬ט' ר' מינור‪ ,‬לוס אנג'לס‪ ,‬ארה"ב‬

‫מאמר זה סוקר מחקרים עדכניים על התרומה של הציטוקין‬ ‫הפרו–דלקתי אינטרפרון ‪1β -‬ח)‪ )IL-1β‬והנוקלאוזיד הפוריני‪,‬‬ ‫אדנוזין‪ ,‬בבקרת התנהגות דמוית דיכאון ותגובת נסיגה‬ ‫המכונה ‪ conservation-withdrawal‬בחיות מודל של דיכאון‬ ‫ושל חולי‪ .‬הפעלה של רצפטורי ‪ IL-1β‬במוח תורמת כנראה‬ ‫לתגובת ‪ conservation-withdrawal‬בחיות שטופלו ברזרפין‬ ‫או בליפופוליסכריד‪ ,‬מה שמרמז על מנגנון משותף בבסיס‬ ‫השפעת חומרים אלו‪ .‬בנוסף‪ ,‬מעבר האותות בעקבות שחרור‬ ‫ציטוקין במוח מסוגל להפעיל את מעבר האותות הקשור‬

‫כתב עת ישראלי‬ ‫לפסיכיאטריה‬ ‫תקצירים‬ ‫תיאוריות גלוטמטרגיות של סכיזופרניה‬ ‫ד' ס' ג'ביט‪ ,‬ניו יורק‪ ,‬ארה"ב‬

‫סכיזופרניה הינה הפרעה נפשית חמורה הפוגעת בכ־‪1%‬‬ ‫מהאוכלוסייה העולמית‪ .‬במודלים מסורתיים של המחלה‬ ‫הודגש תפקוד דופמינרגי לקוי‪ .‬עם זאת‪ ,‬ב־‪ 20‬השנים‬ ‫האחרונות מגבלות מודל הדופאמין הפכו ברורות יותר‬ ‫ויותר‪ ,‬ונוצר הצורך בפיתוח מודלים חלופיים‪ .‬המודלים‬ ‫הגלוטמטרגיים מבוססים על האבחנה כי גורמים פסיכו־‬ ‫מוטוריים כגון )‪ phencyclidine (PCP‬ו־‪ketamine‬‬ ‫משרים תסמינים פסיכוטיים והפרעות נוירו־קוגניטיביות‬ ‫הדומים לאלו של סכיזופרניה‪ ,‬זאת על ידי חסימת קולטני‬ ‫‪type glutamate‬־)‪aspartate (NMDA‬־‪D‬־‪methyl‬־‪.N‬‬ ‫משום שקולטני ‪ glutamate/NMDA‬מפוזרים במוח‪,‬‬ ‫המודלים הגלוטמטרגיים מנבאים הפרעה קורטיקלית נרחבת‬ ‫בשילוב מעורבות ייחודית של קולטני ה־‪ NMDA‬במוח‪.‬‬ ‫כמו כן‪ ,‬הימצאות קולטני ‪ NMDA‬הממוקמים במעגלים‬ ‫מוחיים ומווסתים את שחרור הדופאמין מציעה כי גירעונות‬ ‫דופאמינרגיים בסכיזופרניה עשויים להיות משניים להימצאות‬ ‫הפרעות גלוטמטרגיות‪ .‬גירוי העברה המתווכת על ידי קולטני‬ ‫‪ ,NMDA‬כגון ‪ glycine-site agonists‬ומעכבי העברת ‪,glycine‬‬ ‫הציג תוצאות מעודדות במחקרים קדם־קליניים‪ ,‬וכרגע מתקיימים‬ ‫פיתוחים קליניים בתחום‪ .‬כמו כן נצפו תוצאות קליניות בגורמים‬ ‫כגון החומר הכימי המתחרה ‪ metabotropic 2/3‬המפחית‬ ‫את רמת ה־‪ ,resting glutamate‬היפוך פוטנציאל ההפרעה‬ ‫בתבנית הירי בקורטקס הפרה־פרונטלי‪ ,‬וייתכן שגם באזורים‬ ‫אחרים במוח‪ .‬באופן כללי‪ ,‬על סמך תוצאות אלו ניתן להציע‬ ‫כי תיאוריות גלוטמטרגיות עשויות להוביל להמשגות חדשות‬ ‫ולגישות טיפוליות אשר לא היו אפשריות בהתבסס על תיאוריות‬ ‫הדופאמין לבדן‪.‬‬

‫פגיעה בבקרה האינהיביטורית של תנודות‬ ‫מחזוריות של מקבצי עצבים פירמידאליים נמצאת‬ ‫בבסיס ההפרעה בזיכרון העבודה ובפונקציות‬ ‫ביצועיות גבוהות בסכיזופרניה‪ :‬הרציונל‬ ‫להתערבויות תרופתיות גאבארגיות (‪(GABAergic‬‬ ‫ס' א' דויטש ‪ ,‬ב' ל' רוס‪ ,‬ג' שוורץ‪ ,‬ג' א' מאסטרופאולו‪,‬‬ ‫ג' א' בורקט‪ ,‬א' וייצמן‪ ,‬נורפולק‪ ,‬וירג'יניה‪ ,‬ארה"ב‬

‫‪israel journal of‬‬

‫‪psychiatry‬‬ ‫כרך ‪ ,47‬מס' ‪2010 ,1‬‬

‫חומצה גאמא–אמינו בוטירית (‪ )GABA‬היא הנוירוטרנסמיטר‬ ‫האינהיביטורי המרכזי במוח‪ .‬היא מסונתזת מגלוטמט‪ ,‬שאף‬ ‫היא מתפקדת כנוירוטרנסמיטר מקבוצת החומצות האמיניות‪,‬‬ ‫ומאוכסנת בעצבי קישור (‪ )interneurons‬דיפרנציאטיביים‬ ‫המאופיינים בפעילות מעכבת‪ .‬אינטרנוירונים מעכבים גאבארגיים‬ ‫נבדלים זה מזה במיקומם האנטומי‪ ,‬בצורתם ובאתרי המטרה של‬ ‫קצותיהם הסינפטיים‪ ,‬וכן בתכולת החלבונים והנוירופפטידים‬ ‫שלהם (לדוגמה פארוואלבומין ‪ ;parvalbumin -‬כולציסטוקינין‬ ‫ ‪ ;cholecystokinin‬וראלין ‪.)reelin -‬‬‫תאי עצב גאבארגיים אינהיביטוריים בקורטקס הקדמי‬ ‫יכולים ליצור קשרים סינפטיים עם יותר מ–‪ 200‬תאי עצב‬ ‫פירמידאליים שונים‪ ,‬שהינם תאי הפלט העיקריים‪ .‬יתר על‬ ‫כן‪ ,‬הם יוצרים כמה סינפסות על התאים הפירמידאליים‪ ,‬כולל‬ ‫על אלומות הדנדריטים (‪ )shafts of dendritic spines‬שעל‬ ‫הסתעפויות הדנדריטים‪ ,‬גופי התאים והקטעים התחיליים של‬ ‫התפצלות האקסונים‪ ,‬שיכולים לקבוע את אופי הפלט וקצב‬ ‫הירי‪ .‬לכן הפעילות הפאזית של עצבים גאבארגיים יכולה לבקר‬ ‫את התנודות הפריודיות של מקבצי עצבים פירמידאליים‪,‬‬ ‫שתזמונן המדויק הוא חיוני לתפקודים קורטיקליים גבוהים‬ ‫כמו זיכרון עבודה‪ .‬האות הגאבארגי מועבר באמצעות קולטנים‬ ‫ממשפחת הקולטנים שהתקשרותם עם הליגנד שלהם פותחת‬ ‫תעלת יונים‪ .‬הקולטן מורכב מחמש תת–יחידות חלבוניות‪.‬‬ ‫חמש התת–יחידות שמרכיבות את הקולטן ל–‪GABA‬‬ ‫ממוקמות בממברנה‪ ,‬ויוצרות תעלת כלור שפתיחתה‬ ‫מתאפשרת על ידי קישור ה–‪ .GABA‬ישנן כמה דרכי פעולה‬ ‫תרופתיות אפשריות להגברת ההעברה הגאבארגית‪ ,‬חלקן‬ ‫מאוד סלקטיביות ומכוונות לקולטני ‪ GABAA‬המורכבים‬ ‫מתת–יחידות מסוימות‪ .‬שאלה מרכזית במחקר התרופתי היא‬ ‫האם התערבות תרופתית טונית‪ ,‬אפילו היא סלקטיבית‪ ,‬יכולה‬ ‫לדמות בצורה נכונה את הפגיעה בתנודות הפריודיות של‬ ‫מקבצי עצבים פירמידאליים‪.‬‬ ‫המחזורים הצירקדיים וגנים הקשורים‬ ‫לשעון הביולוגי בהפרעות פסיכוטיות‬ ‫א' וודינגטון־למונט‪ ,‬ד' ל' קוטו‪ ,‬נ' סרמאקיין‪ ,‬ד' ב' בויווין‪,‬‬ ‫מונטריאול‪ ,‬קנדה‬

‫עדויות רבות מצביעות על כך שהפרעות במערכת המחזוריות‬ ‫היומית (המערכת הצירקדית) תורמות להתפתחות הפרעות‬ ‫פסיכיאטריות ולקשת התסמינים הקליניים שלהם‪ .‬הפרעות‬ ‫שינה‪ ,‬בעיקר בשנת תע"מ (‪ )REM‬המאופיינת בתנועות‬ ‫עיניים מהירות‪ ,‬נצפו במחלה דו–קוטבית (‪)Bipolar disorder‬‬ ‫‪90‬‬