Neurobiology of Behaviour University of Toronto
Fall 2014
University of Toronto Fall 2014
Table of Contents 1. Expert Meditators are Better at Impulse Control than Non-Meditators - 1-4 Authors: Tineke Kruytbosch, Peter Nguyen, Monica Akula
2. Enhanced Learning Exhibited by Mice Expressing Humanized FOXP2 - 5-7 Authors: Ingrid Grozavu, Chloe Carducci, Nouran Sakr, Erik Friesen
3. A Potential New Drug for Depression - 8-12 Authors: Charmaine Chan, Jennifer Cheng, Jenny Chong, Sally Moy, Kenneth Nguyen
4. Stay Awake and Stressed for Better Memory - 13-17 Authors: Chuen, Victoria., Lee, Michelle., Zhang, Ifan (Audrey)
5. Functional Connectivity as a Predictor of rTMS Treatment Response in Major Depressive Disorder - 18-21 Authors: Fettes, Peter; Ceniti, Amanda; Burke, Charles
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Expert Meditators are Better at Impulse Control than Non-Meditators Authors: Tineke Kruytbosch, Peter Nguyen, Monica Akula University of Toronto - HMB420 October 28, 2014
Introduction and Background Benjamin Libet was one of the first to empirically study free will. His landmark 1983 paper showed that participants’ voluntary, spontaneous movement could be predicted by electroencephalography (EEG) recordings1. The basic method involved asking participants to click a button whenever they wanted. They had to note the time that they felt the initial will to move based on a clock in a corner of the experimental setup. While participants were carrying out these actions, EEG data was recorded1. Libet looked at slow cortical potentials (SCPs)2, which are a type of event-related potential3 resulting from slow, excitatory postsynaptic potentials in cortical dendrites that can either consist of negative or positive deflections. Libet particularly paid attention to readiness potentials (RPs), which are a type of SCP occurring right before a person carries out voluntary movements and are associated with negative deflections1. The results showed that button pressing could be predicted by RPs that occurred before participants were even aware of the intention to act1. This led to controversy, one of the reasons being that most people feel the subjective experience of having free will. Some scientists supported his findings, whereas many others criticized his experiment, particularly the experimental
design. Some criticism included the possible delay in subject reports of time4. This could have been a result of subjects possibly not focusing on the experiment, leading to inaccurate time reports5,6. Jo et al.’s general purpose was to study and integrate subjective experience of participants and objective data, adding to the overall understanding of free will7. The authors also tried to address the problem of subject attention by using one expert meditator named TL as part of the sample, because of past research suggests that meditation increases control of attention and more accurate reports of bodily sensations8. TL was paired with 5 age-, gender- and education-matched controls. Jo et al. asked participants to carry out 3 separate tasks, which were similar to Libet’s experiment. EEG recordings were taken, after which participants were given a post-task questionnaire to obtain subjective data8.
Results Session 1: M-task and W-task TL and the controls had to perform M-task (report button press) and W-task (report impulse arrival). In both M-task and W-task, TL and controls displayed a rising RP amplitude that precedes each button press (Figure 1 A/B)8. Likewise, a higher probability of pressing the button was observed during the negative deflections
2 for both TL and controls (Figure 1 C/D)8. Overall, TL displayed stronger RP amplitude and higher probability, suggesting his superior conscious control8. Figure 18. RPs (y-values inverted) for TL (black) and controls (greys) in M-task (A) and W-task (B). Button press indicated by vertical line (0 seconds). Light grey traces represent standard deviations (SD) of control mean.
Figure 28. Probabilities of pressing the button during each deflection (negative and positive) are visualized by superimposing a SCP wave in M-task (C) W-task (D). Black and white bars correspond to TL and controls (error bars represent SD) respectively.
Session 2: W-task and Holding task TL was the only participant and he had to perform W-task again and a Holding-task where he held onto an impulse for as long as possible and only then was he allowed to push the button. The same rising RP amplitude for W-task was observed, whereas a rising RP amplitude was also observed for Holding-task, but occurred at 3 seconds before the button press (how long TL held onto the impulse for)8. Both W-task and Holding-task displayed higher probabilities of button presses during the negative deflections rather than positive deflections of the SCP8. Figure 38. RPs (A) and probabilities (B) for (W-task (black) and Holding-task (grey) are displayed. The probability at 3 seconds before the button press in holding-task are represented by white bars.
Session 3: Less-Effort task and Strong-Effort task TL performed a less-effort-task and a strong-effort-task where he waited for an upcoming impulse and then act or alternatively “actively� act in the absence of an impulse, respectively. In less-efforttask, the ongoing negativity (blue) began much earlier, was much stronger, and had more localization (supplementary motor area) than in strong-effort-task8.
3 Also, stronger RP amplitude and higher probability of button presses during negative deflections were observed in less-effort-task (figures not shown)8. Figure 48. Topographic maps based on EEG recordings of TL’s cortical activity during these tasks. Blue areas represent rising RP amplitudes.
Conclusions and Discussion Jo et al.’s (2014) paper reached two main conclusions regarding slow cortical potentials: negative deflections in SCPs precede one’s intention to act; and these negative deflections indicate that the action took less effort than if there had been no negative deflection in SCP8. Regarding ‘free will’ and the purpose of Libet’s original study, Jo et al. (2014) concluded that this experiment did not test free will. Rather, it tested impulse control8. In spite of the attempt by Jo et al. (2014) to improve upon Libet’s original research, the paper had some major faults. Using a very small sample size of N=1, for example, was a weakness that calls the results into question. Expanding the sample size and
using a more diverse group of participants, both meditators and non-meditators alike, could have easily improved this. In addition, the reliance on meditation and participant interviews brought an element of subjectivity into the study, which can be seen as a weakness. Possible future directions for this area of research could be to expand more on the ideas of free will and meditation. The results of Jo et al.’s (2014) paper show that the expert meditator was more accurate and in-tune with his inner impulse to act than the controls. This calls into question whether expert meditators, such as Buddhist monks, experience ‘free will’ differently to non-experts. Future studies could also include more diverse actions to more accurately test free will. For example, participants could have the choice to click a button, wave their hand, or shrug their shoulder, and the choice the make one of these may be a more accurate test of free will.
References 1. Libet, B., Gleason, C. A., Wright, E. W., & Pearl, D. K. Time of conscious intention to act in relation to onset of cerebral activity (readiness-potential). The unconscious initiation of a freely voluntary act. Brain 106, 623–642 (1983). 2. Strehl, U. Slow cortical potentials neurofeedback. Journal of Neurotherapy 13, 117-126 (2009). 3. Ibanez, A., Melloni, M., Huepe, D., Helgiu, E., Rivera-Rei, A., Canales-Johnson, A., Baker, P., & Moya, A. What event related potentials (ERPs) bring to social n euroscience? Social Neuroscience 7, 632-649 (2012).
4 1. Ahmadian, P., Sanei, S., Ascari, L., Gonzalez-Villanueva, L., & Umilta, M. A. (2013). Constrained blind source extraction of readiness potentials from EEG. IEEE Trans. Neural. Syst. Rehabil. Eng. 21, 567-575 (2013). 2. Stanley, K. Libet’s Research on the Timing of Conscious Intention to Act: A Commentary. Conscious. Cogn. 11, 273 279 (2002). 3. Danquah, A. N., Farrell, M. J., & O’Boyle, D. J. Biases in the subjective timing of perceptual events: Libet et al. (1983) revisited. Conscious. Cogn. 17, 616–627 (2008). 4. Guggisberg, A. G., & Mottaz, A. Timing and awareness of movement decisions: does consciousness really come too late? Front. Hum. Neurosci. 7, 1–11 (2013). 5. Jo, H-G., Wittmann, M., Borghardt, T.L., Hinterberger, T., & Schmidt, S. First person approaches in neuroscience of consciousness: Brain dynamics correlate with the intention to act. Conscious Cogn. 26, 105-116 (2014).
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Enhanced Learning Exhibited by Mice Expressing Humanized FOXP2 Authors: Ingrid Grozavu, Chloe Carducci, Nouran Sakr, Erik Friesen University of Toronto - HMB420 October 27, 2014
FoxP2 is a gene of interest as it was the first to be linked to language and speech impairments1. Research done in the early 1990s on the British, multi-generational KE family was a crucial first step in forming this association. Over half the members in this family exhibited a form of language deficit, proposed to be developmental verbal dyspraxia (DVD). Since then, extensive research has further implicated FoxP2’s role in language disorders2. Evolutionary analysis has indicated that two amino acid substitutions in FoxP2 have been positively selected for during evolution3. Previous research has shown that introducing these substitutions to the endogenous FoxP2 gene in mice resulted in increased synaptic plasticity in the striatum, altered dopamine levels4, and differences in ultrasonic vocalizations. In addition, it has been proposed that different regions of the striatum mediate different forms of learning5. For instance, the dorsomedial striatum (DMS) has been shown to mediate declarative learning, while the dorsolateral striatum (DLS) has been implicated in procedural learning. Homologous regions have also been discovered in human brains, which respond to each mode of learning preferentially6.
In this paper7, the researchers investigated how these striatumdependent learning systems are affected in mice with the ‘humanized’ form of the FoxP2 gene, a topic that has not yet been addressed in the literature. It was hypothesized that FoxP2 has evolved in humans in order to adapt and prepare the brain circuits for language acquisition, through the differential tuning of coritcostriatal systems that are involved in declarative and procedural learning. The first set of experiments conducted were maze navigation by Foxp2hum/hum versus WT mice, where spatial cues were either present or absent. It was found that the humanized mice learned faster in the cueenriched setup, however no difference was uncovered between the strains in the cue-deprived maze. Reduced competition between the DMS and DLS was proposed as the underlying mechanism. To test this hypothesis, the mice were forced to shift from declarative to procedural learning and vice-versa using a previously described cross-maze paradigm. Trials surfaced that humanized mice transitioned faster than WT from declarative to procedural learning, while no difference was observed when shifting from procedural to declarative learning.
To assess why this occurred, neurophysiological testing was performed where mRNA expression levels, dopamine levels, and ease of LTD induction were measured. Firstly, mRNA expression profiles found up-regulation of “groups” of genes in the DMS Foxp2hum/hum mice only, including signaling, neurotransmitter transporter, and synaptic regulatory genes. Secondly, dopamine levels were 70% lower in the DMS of Foxp2 mice hum/hum compared to WT. Finally, the ease of LTD induction by tetanic high frequency stimulation was tested to a depolarization of -70mV (weak) and -15mV (strong). Results showed stronger LTD in the DLS of humanized mice relative to WT mice at -70mV, a difference that was eliminated at a stronger (-15mV) depolarization. This concluded that LTD is most easily inducible in the DLS of Foxp2hum/hum mice. Molecular mechanisms of the above results were then investigated as follows. Sulpiride (a D2R antagonist) treatment completely eliminated LTD induction in the striatum of humanized mice, revealing that the mechanism is D2R-linked. It is important to note that LTP may act as a confounding variable, making it essential to check against with APV treatment. This trial demonstrated no change in LTD response of the DLS of WT mice, meaning that LTP did not skew previous results. Conversely, APV addition prevented LTD induction in the DLS of Foxp2hum/hum mice, thus demonstrating that the mechanism involves NMDA receptors. Finally, LTD induction was observed to require postsynaptic activation of NMDA receptors, because LTD induction was blocked in MK801 treatment. On a behavioural level, the key result of this study was clear: in mice, humanized FoxP2 allows for a more efficient interaction between declarative and procedural learning.
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Though clear, this result fails to explain (1) its neurophysiological underpinnings, (2) how it relates to the function of FoxP2 in human memory systems, and (3) how this relates to human language development. The researchers only addressed the first of these problems. However, though these tests yielded some interesting results, they none the less failed to elucidate whether the observed neurophysiological changes were actually causing the different learning patterns in humanized mice, and if so, the mechanism underlying these changes. Therefore, from a neurophysiological standpoint, there remains much work to be done in uncovering the cellular mechanisms of FoxP2 in modulating learning behaviours in mice. Another important future direction is to reconcile the gap between the role of ‘humanized’ FoxP2 in mice and its role in humans. This reconciliation is important for a number of reasons: first, understanding the neurophysiology of FoxP2 in humans could allow for the development of effective treatments for DVD, as this is a condition associated with the mutation of one FoxP2 allele10. Second, gaining a better understanding of how (or if) foxP2 mediates the interaction between the human procedural and declarative memory systems could be important in other regards. For example, researchers in the field of Neuroeconomics are currently trying to understand the interaction between multiple memory systems during human decision-making8. Understanding a genetic factor, such as FoxP2, that appears to influence the relationship between declarative and procedural learning may therefore allow for large strides to be made in our understanding of human decision-making.
In conclusion, the transition from declarative to procedural learning is enhanced by the integration of the humanized FOXP2 gene into the murine genome. This conclusion implies that the premise for the evolution of the human FOXP2 gene may have occurred in order to refine the systems that are explicitly involved in the automizationproceduralization of learning in humans. However, direct evolutionary conclusions of FOXP2 have yet to still be analyzed. Furthermore, experiments investigating the role of FOXP2 in multiple memory systems and how FOXP2 is intricately involved in the decision making process is of great interest for future research. In short, further investigation would be to demonstrate how FOXP2 brings about a behavioural change in murine models.
References 1. Nudel, R. & Newbury, D (2013). Foxp2. WIREs Cogn Sci , 4, 547-560. 2. Fisher, S. & Scharff, C. (2009). FOXP2 as a molecular window into speech and language. Trends in Genetics, 25 (4), 166-177. 3. Enard, W. (2011), FOXP2 and the role of cortico-basal ganglia circuits in speech and language evolution. Current Opinion in Neurobiology, 21, 415-424. 4. Enard, W. et al. (2009). A humanized version of Foxp2 affects cortico-basal ganglia circuits in mice. Cell, 137, 961-971. 5. Yin, HH. Et al. (2009). Dynamic reorganization of the striatal circuits during acquisition and consolidation of a skill. Nat Neurosci, 12 (3), 333-341.
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6. Balleine, B. & O’Doherty, J.. (2009). Human and rodent homologies in action control Corticostriatal determinants of goal-directed and habitual action. Neuropsychopharmacology, 25, 48-69. 7. Schreiweis C, Bornschein U, Burguière E, Kerimoglu C, Schreiter S, Dannemann M, Goyal S, Rea E, French CA, Puliyadi R, Groszer M, Fisher SE, Mundry R, Winter C, Hevers W, Pääbo S, Enard W, Graybiel AM. Humanized Foxp2 accelerates learning by enhancing transitions from declarative to procedural performance. Proc Natl Acad Sci U S A. 2014 Sep 30;111(39):14253-8. 8. Delgado MR, Dickerson KC. Reward-related learning via multiple memory systems. Biol Psychiatry. 2012 Jul 15;72(2):134-41. 9. French CA, Fisher SE. What can mice tell us about Foxp2 function? Curr Opin Neurobiol. 2014 Oct;28C:72-79. 10. MacDermot KD, Bonora E, Sykes N, Coupe AM, Lai CS, Vernes SC, Vargha-Khadem F, McKenzie F, Smith RL, Monaco AP, Fisher SE. Identification of FOXP2 truncation as a novel cause of developmental speech and language deficits. Am J Hum Genet. 2005 Jun;76(6):1074-80.
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A Potential New Drug for Depression Authors: Charmaine Chan, Jennifer Cheng, Jenny Chong, Sally Moy, Kenneth Nguyen University of Toronto - HMB420 October 28, 2014
Introduction In North America, depression has become an increasingly more prevalent mental disorder and in response to this, there is a growing need for effective antidepressant medication. However, classical antidepressants are only most effective on patients suffering from severe depression2. As a result, alternative forms of treatment are needed. Drawing from traditional Chinese medicine, β-asarone derived from wild ginger has long been known to treat depression yet the scientific community has only recently examined its mechanisms. Although clinical advantages to β-asarone have already been demonstrated7, this study is the first to suggest that β-asarone promotes adult neurogenesis through the ERK1/2-CREB cascade, providing a cellular molecular explanation. From
Background the monoamine hypothesis, which was based on looking at the mechanisms of action of known antidepressants, depression is thought to be induced by a chemical imbalance that resulted in a deficiency of three neurotransmitters, serotonin, norepinephrine, and dopamine. This deficiency causes an up-regulation of specific postsynaptic monoamine receptors. However, support for this hypothesis was unclear as symptom ameliorated only after prolonged usage of
antidepressants ( neurotransmitters levels increased immediately) and decreasing these neurotransmitters did not ensure depression. The neurotrophic theory of depression ushered in a new framework, placing emphasis on downstream cascades and gene expression. Beta-asarone targets BDNF, a downstream target that determines neuronal survival, through the ERK ½-CREB cascade. In depression, repressed BDNF can lead to neuronal atrophy in the hippocampus. Clinical imaging studies provide support for this as reduced brain volumes were shown to result from knockout studies. Adding to the existing literature supporting the neurotrophin theory of depression and focusing on downstream targets to treat depression, studying β-asarone seems like the right way to go in order to find alternative treatments for depression. Using the Chronic Unpredictable Mild Stress (CUMS) mouse model, a widely used model for stress-induced depression, rats were placed in an aversive environment through food deprivation and random strobe light stimuli8. Extracted from Acornus tatarinowii Schott (wild ginger), β-asarone has already previously demonstrated various advantages to classical antidepressants such as minimal side effects due to the downstream neurotrophin targeting, fast acting effects as it passes through the blood brain
barrier, and the ease of oral ingestion as a method of intake7. Moreover, evidence for neuroprotective effects have been provided through research into brain ischemia11 and Alzheimer’s6, this has also shown that beta-asarone is safe to use in the brain. Most importantly, the researchers sought to answer whether experimental science could reconcile the reasoning behind why wild ginger is used to treat depression in Traditional Chinese Medicine with empirical findings from experimental science. Furthermore, this paper also addresses whether β-asarone treatment could serve as an alternative treatment for depression as this method is the first to demonstrate adult hippocampal neurogenesis.
Major Results The authors used a series of methods to measure the effects of β-asarone treatment, administered orally for 28 days concurrent with CUMS exposure, on male Spraque-Dawley rats. Significant decrease in sucrose preference and increased immobility time from the sucrose preference test (SPT) and forced swim test (FST) respectively1 ensured anti-depressive behavior in rats from CUMS exposure. In comparison to the saline treatment, β-asarone treated rats demonstrated a significant increase in sucrose preference and decrease in immobile time1, almost fully (SP) and partially (immobility) reversing the stressed-induced effects. The authors observed the distribution of newly born cells with the use of BrdU labeling, in the subgranular zone (SGZ) of the rat hippocampi, one of the two areas of adult neurogenesis. With β-asarone treatment after CUMS exposure, not only
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was there a significant increase of BrdU+ cells compared to saline-treated rats (Fig. 1A), but the distribution of SGZ newborn cells were comparatively more condensed into layers, suggesting increased sitespecific neurogenesis (Fig. 1B)1.
Fig. 1A The number of BrdU positive cells in the subgranular zone of the rat hippocampi presented in a histogram. A comparison between rats receiving the saline treatment and β-asarone treatment across stress/nonstress conditions. The mean number BrdU+ cells were not significantly different between the two stress conditions in β-asarone-treated rats, whereas the mean number of BrdU+ cells were in saline-treated rats.
Fig. 1B Photomicrographs of the distribution of newborn cells in the subgranular layer of rat hippocampi, labeled with BrdU and stained blue. A comparison between of distribution of cells of rats receiving the saline treatment and β-asarone in stress/non-stress conditions are shown.
Because the ERK1/2 and CREB signaling pathway had been proposed to be involved, the authors performed Western blots to determine levels of p-CREB and p-ERK1/2. β-asarone treated CUMS rats were found to have a significant increase in p-CREB and p-ERK1/2 expression (Fig. 2A and 2B) which indicate activation of the ERK1/2 pathway.
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Fig. 2A Graph of phosphorylated CREB levels normalized to total CREB levels. Note that total CREB levels and GAPDH (loading control used in the immunoblot analysis) remained constant, and the graph only documents changes in p-CREB levels. p-CREB levels of β-asarone-treated rats were not significant across the two stress conditions, whereas the p-CREB levels of saline-treated rats were.
Fig. 2B Graph of phosphorylated ERK1/2 levels normalized to total ERK1/2 levels. Note that total ERK1/2 levels and GAPDH (loading control used in the immunoblot analysis) remained constant, and the graph only documents changes in p-ERK1/2 levels. p-ERK1/2 levels of β-asarone-treated rats were not significant across the two stress conditions, whereas the p-ERK1/2 levels of saline-treated rats were.
They also looked at protein levels and mRNA levels of BDNF (using RTPCR). β-asarone treated CUMS rats had a significant increase of 63% in protein levels compared to the saline treated CUMS rats (Fig. 3A). An increase of BDNF mRNA levels was also found, indicating an increase in BDNF gene expression (Fig. 3B).1
Fig. 3A Graph of BDNF protein levels normalized to GAPDH, loading control. BDNF protein levels were acquired through immunoblotting techniques. BDNF protein levels in the no stress conditions were similar for both the saline and β-asarone treated rats. Significant increase in BDNF protein levels was observed in the β-asarone CUMS treated rats compared to the saline-treated CUMS rats
Fig. 3B Graph of phosphorylated ERK1/2 levels normalized to total ERK1/2 levels. Note that total ERK1/2 levels and GAPDH (loading control used in the immunoblot analysis) remained constant, and the graph only documents changes in p-ERK1/2 levels. p-ERK1/2 levels of β-asarone-treated rats were not significant across the two stress conditions, whereas the p-ERK1/2 levels of saline-treated rats were.
Conclusion These results implicate β-asarone as a potentially effective antidepressant compound, possibly through induction of hippocampal neurogenesis. Furthermore, activation of the ERK1/2 and CREB signaling pathway and increased BDNF expression were observed, indicating a neuroprotective effect. However, the results of this study are preliminary and have not addressed some key issues.
Although the authors successfully demonstrated a significant increase of newborn cells from the β-asarone treatment using BrdU labeling, which is an appropriate measure of neurogenesis, it would further benefit the study if the authors also measured cell apoptosis of SGZ newborn cells through a TUNEL assay3. BrdU is also implicated in DNA repair and gene duplication, and these cellular events should be differentiated from neurogenesis10. Figure 2B has poor resolution and has possibly been manipulated as some parts of the western blot appear to be superimposed. The full, unaltered western blot should be included with the journal article in the supplementary materials to dispel notions of tampering. In addition, the article suggests that β-asarone reverses CUMS-induced depression, but treatment was administered concurrently with CUMS stressors, and the possibility that it only slows down or stunts the effects of stressinduced depression remains. To address this issue, a long-term study should be conducted with varying β-asarone administration time points throughout the experimental timeline. The inclusion of other tests for depression in addition to the sucrose preference test and forced swim test are recommended to understand what types of stress β-asarone may be effective for. Although the ERK1/2 and CREB signaling pathway has been shown to be a target of antidepressant drugs in other research9, causation was not established between this pathway and β-asarone’s antidepressant property specifically. The possibility that β-asarone’s antidepressant property is derived from a different pathway cannot be completely dismissed.
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Thus further research on the mechanism of β-asarone is necessary to understand how its antidepressant property is derived. Due to its effect on neurogenesis and neuroprotection, β-asarone may be a potential treatment not just for stress-induced depression, but also for neurodegenerative diseases. It has since been implicated as a potential alternative treatment for Parkinson’s Disease when co-administered with dopamine through promotion of L-dopa conversion in the striatum5.
References 1. Dong, H.Y., Gao, Z.Y., Rong, H., Jin, M., Zhang, X.J. β-asarone Reversed Chronic Unpredictable Mild Stress-Induced Depression-Like Behavior and Promotes Hippocampal Neurogenesis in Rats. Molecules. 19, 5634-5649. (2014) 2. Fournier, J.C., DeRubeis, R.J., Hollon, S.D., Dimidjian, S., Amsterdam, J.D., Shelton, R.C., & Fawcett, J. Antidepressant drug effects and depression severity: a patient-level meta-analysis. JAMA. 303(1), 47-53 (2010). 3. Gavrieli, Y., Sherman, Y., & Ben-Sasson, S.A. Identification of Programmed Cell Death In Situ via Specific Labeling of Nuclear DNA Fragmentation. J Cell Biol. 119(3), 493-501 (1992). 4. Geng, Y., Li, C., Liu, J., Xing, G., Zhou, L., Dong, M., Li, X., & Niu, Y. Beta-Asarone Improves Cognitive Function by Suppressing Neuronal Apoptosis in the Beta-Amyloid Hippocampus Injection Rats. Biological & Pharmaceutical Bulletin. 33(5), 836-843 May (2010). 5. Huang, L., Deng, M., Zhang, S., Fang, Y., & Li, L. Co-administration of β-asarone and Levodopa increase dopamine in rat’s brain via accelerating transformation of Levodopa: a different mechanism from madopar. Clin Exp Pharmacol Physiol. 19, 5634-5649 (2014).
6. Li, C., Xing, G., Dong, M., Zhou, L., Li, J., Wang, G., Zou, D., Wang, R., Liu, J., & Niu, Y. Beta-asarone protection against betaamyloid-induced neurotoxicity in PC12 cells via JNK signaling and modulation of bcl-2 family proteins. Eur J Pharmacol. 635(1-3), 96-102 (2010). 7. Li, Z., Zhao, G., Qian, S., Yang, Z., Chen, X., Chen, J., Cai, C., Liang, X., & Guo, J. Cerebrovascular protection of β-asarone in alzheimer’s disease rats: A behavioral, cerebral blood flow, biochemical and genic study.J Ethnopharmacol. 144(2), 305-312 (2012). 8. Ossowska, G., Danilczuk, Z., KlenkMajewska, B., Czajkowski, L., & ZebrowskaŁupina, I. Antidepressants in chronic unpredictable mildstress (CUMS)-induced deficit of fighting behavior. Pol J Pharmacol. 56, 305–311 (2004). 9. Qi, X., Lin, W., Li, J., Li, H., Wang, W., Wang, D., & Sun, M. Fluoxetine increases the activity of the ERK-CREB signal system and alleviates the depressive-like behaviour in rats exposed to chronic forced swim stress. Neurobiol Dis. 31, 278-285 (2008). 10. Taupin, P. BrdU immunohistochemistry for studying adult neurogenesis: paradigms, pitfalls, limitations, and validation. Brain Res Rev. 53, 198-214 (2007). 11. Yang, Y., Chen, Y., Zhou, X., Hong, C., Li, C., & Guo, J. Beta-asarone, a major component of acorus tatarinowii schott, attenuates focal cerebral ischemia induced by middle cerebral artery occlusion in rats. BMC Complem Altern M. 13, 236. (2013).
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Stay Awake and Stressed for Better Memory
Authors: Chuen, Victoria., Lee, Michelle., Zhang, Ifan (Audrey). University of Toronto - HMB420 October 27, 2014
Introduction Sleep-associated memory consolidation is not a new concept in scientific literature; there is a rich pool of research on sleep and its effects on memory, as well as which neural pathways are involved. These neural pathways and networks have long been areas of interest for researchers, both for the expanding field of neuroscience and possibilities for enhancing human cognition—after all, improved memory means improved learning capability.
Background Currently, it is hypothesized that memory consolidation occurs through the reactivation of neural activity patterns to stabilize labile representations in the hippocampus into engrams within neocortex1. It has been suggested that this process occurs during sleep, where declarative memories and non-declarative memories are consolidated during slowwave and rapid eye movement (REM) sleep respectively. Conversely, wakefulness promotes learning but not necessarily memory consolidation. This ensures that the memory encoding and consolidation phases do not interfere with one another2.
More recently there has been a growing interest in understanding how glucocorticoids (GC) fit in the overall picture of memory and sleep, especially due to the high concentration of hippocampal GC receptors. It is known that GCs can impair memory retention, but enhance learning— it is dependent on the memory phase4. During wakefulness, corticosterone, a GC found in rats, activates the HPA axis pathway and increases AMPA receptors at the synapse to improve memory consolidation3. Manipulation of exogenous GCs in rats has supported the idea that GCs can enhance memory consolidation. Together, these studies have emphasized the potential role of GCs in the interplay between memory and sleep processes4. There was an understanding that both memory processing and physiological levels of GCs were state dependent- GC levels are minimal during sleep and maximum during wake. However, although studies have investigated the enhancing effects of GC during wake states, they have failed to acknowledge the potential differential effects of GCs during sleep and wake states. It was suggested in 2011 by Wilhelm, I. et al.5, that there is a state-dependent effect of cortisol levels on memory consolidation in humans. Systemic injections of cortisol during either the wake or sleep state
14 indicating that cortisol impaired memory consolidation during sleep, but enhanced it during wakefulness. In this review, we will discuss a recent study performed by Kelemen, E. et al.6, which aimed to recapitulate the results obtained from the human study5, in rats. The purpose was to investigate whether the effects of GC on memory consolidation are localized to hippocampal networks, and if these effects are dependent on their administration during the sleep or wake state. In addition to validating previous findings of statedependent effects of GCs on memory consolidation in humans, this study also introduces a new basis to explore the cellular and molecular mechanisms underlying this interesting effect. We will review various limitations and issues to consider as the field of neuroscience continues on with this research.
Results Rats were tested on in an object-place and novel-object recognition task. The rats were first placed in an open field and presented with two novel objects. Subsequently, in the following 80 minutes, the researchers investigated the effects of sleep and sleep deprivation, in addition to the effects of intrahippocampal corticosterone (10ng/0.5uL saline) or vehicle infusion (0.5uL saline) during the sleep or wake state. A subsequent retrieval phase tested for either the object-place or novel-object memory. As expected, in the object-place recognition task, which is hippocampal dependent, results from the vehicle groups showed state-dependent effects on memory consolidation. Rats who were allowed to sleep in the retention phase
Figure 1. Experimental paradigm: Rats were trained in either an object-place recognition task or a novel-object recognition task to investigate the effects of an intrahippocampal infusion of corticosterone or saline into the dorsal hippocampus. Drug infusions were administered during an 80 minute retention period during either the sleep or wake state.
15 showed stronger memories than rats that were sleep deprived. However, this effect was reversed when an intrahippocampal infusion of corticosterone was administered. Researchers observed that the effects of corticosterone on memory consolidation are state dependent. When administered during sleep and in contrast to the vehicle, corticosterone completely abolished the ability to consolidate the object-place memory. Interestingly however, the effects of corticosterone during wakefulness showed a positive effect on consolidation, showing enhanced object-place memory. Conversely, researchers did not find any differential effects on memory consolidation when corticosterone was administered in a novel-object recognition task. As such, researchers concluded that (1) intrahippocampal infusions of corticosterone affect memory consolidation in a state-dependent manner and (2) the effect can be localized to the hippocampal networks.
Conclusion Despite the novel findings of the paper, we must consider the various limitations in their methodology and conclusions. In addition to having limited direct experimental evidence to support their conclusions, many assumptions were also made throughout the experiment. The decision to use a 10ng dose of corticosterone was based solely on the findingsofMedinaetal.wherecorticosterone was administered to the dorsal striatum for an inhibitory avoidance task7. The differences in the behavioral paradigm and brain structures of interest may make this dose unsuitable. In addition, it is possible
the use of “gentle handling� to promote wakefulness during the retention period could increase endogenous corticosterone levels. They relied on results from Meerlo et al., which stated corticosterone levels following such protocols are only observed after 6 hours, thus assumed an 80 minute period would produce negligible effects of endogenous corticosterone8. Furthermore, the researchers used isoflurane during their surgical procedures, which has been suggested to have detrimental effects on spatial memories. This could confound the results obtained in their object-place recognition task9. To solidify their conclusions, the researchers could perform more experiments to look at the molecular basis of the behaviours. Pharmacologically, blocking the GC receptor using RU38-486 (Mifepristone) could block enhanced spatial learning, memory and AMPAR expression10. In order to distinguish between genomic and non-genomic actions of GC receptor expression, we suggest a longer latency period between sampling and retrieval phase. However, if this is implemented, the researchers should account for endogenous increases in GC, which could confound results. Surgical removal of the adrenal cortex could inhibit the production of endogenous GC, or alternatively, Metyrapone, a CORT synthesis blocker could be used to prevent GC production11. In order to further support conclusions that the GC effects were localized to the hippocampus, injection into another region should produce no effects. We suggest injection of GC into the perirhinal cortex, where novel-object recognition has been demonstrated to take place12. We also suggest single-cell electrode recording, as performed by Whitlock et al., to assess
the electrophysiological changes resulting from learning in either the objectplace or novel-object recognition task13. Ultimately, while the results presented in this study are both novel and interesting, much research needs to be done to strengthen our knowledge about the state-dependent effect of corticosterone on memory consolidation. However, these findings may have significance in future clinical applications such as for post traumatic stress disorder (PTSD) patients. Manipulation of GC levels during sleep or wake states could offer an interesting treatment approach to distinguishing or reducing the strength of adverse emotional memories.
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3. Krugers, H. J., Hoogenraad, C. C., & Groc, L. Stress hormones and AMPA receptor trafficking in synaptic plasticity and memory. Nature Rev Neurosci 11(10), 675-681 (2010) 4. Roozendaal, B. Stress and memory: opposing effects of glucocorticoids on memory consolidation and memory retrieval. Neurobiol Learn Mem 78, 578-595 (2002) 5. Wilhelm, I., Wagner, U., & Born, J. Opposite effects of cortisol on consolidation of temporal sequence memory during waking and sleep. J Cognitive Neurosci, 23(12), 37033712 (2011) 6. Kelemen, E., et al. Hippocampal corticosterone impairs memory consolidation during sleep but improves consolidation in the wake state. Hippocampus, 24(5), 510-515 (2014) 7. Medina, A. C. et al. Glucocorticoid administration into the dorsal stratium facilitates memory consolidation of inhibitory avoidance training but not of the context or footshock components. Learn. Mem 14, 673-677 (2007)
Figure 2. Site of suggested corticosterone or saline infusions. Blue arrows indicate infusions into the dorsal hippocampus. Green arrows indicate suggested infusions into the perirhinal cortex14.
References 1. Mehta, M. R. Cortico-hippocampal interaction during up-down states and memory consolidation. Nature Neurosci 10, 13-15 (2007) 2. Diekelmann, S. & Born, J. The memory function of sleep. Nature Rev Neurosci 11: 114-126 (2010)
8. Meerlo, P. et al. Differential effects of chronic partial sleep deprivation and stress on serotonin-1A and muscarinic acetylcholine receptor sensitivity. J. Sleep. Res. 15, 386-394 (2007) 9. Culley, D. J. et al. Impaired acquisition of spatial memory 2 weeks after isoflurane and isoflurane-nitrous oxide anesthesia in aged rats. Anesth. Analg. 99, 1393-1397 (2004) 10. Oitzl M.S., de Kloet, E.R. Selective corticosteroid antagonists modulate specific aspects of spatial orientation learning. Behav. Neurosci. 106, 62-71 (1992)
1. Marin, M. et al. Metyrapone administration reduces the strength of an emotional memory trace in a long-lasting manner. J Clin Endocrinol Metab 96, E1221–E1227 (2011) 2. Albasser, M. M., Poirier, G. L., & Aggleton, J. P. Qualitatively different modes of perirhinal–hippocampal engagement when rats explore novel vs. familiar objects as revealed by c‐Fos imaging. Eur J Neurosci, 31(1), 134-147 (2010)
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Functional Connectivity as a Predictor of rTMS Treatment Response in Major Depressive Disorder Authors: Fettes, Peter; Ceniti, Amanda; Burke, Charles University of Toronto - HMB420 October 27, 2014
Introduction and Background Major depressive disorder (MDD) is a prevalent and debilitating disorder, making it the third leading contributor to the global disease burden1. The DSM-V characterizes MDD by depressed mood, changes in sleep and activity patterns, suicidality, and change in weight amongst others. Its effects are felt both on the individual and on society. However, despite its prevalence, up to 15% of MDD patients present with treatment resistant depression2. This has lead to a call for more effective and cost-efficient treatments in these individuals living with the disease1. Previous studies have shown differences in functional connectivity between refractory and nonrefractory depression3. Salomons et al examined how functional connectivity could be used as a predictor of treatment outcome for repetitive transcranial magnetic stimulation (rTMS) therapy4. This includes an examination of baseline connectivity as well as changes to connectivity after treatment. Leone et al first described the efficacy of rTMS as a non-invasive option in treating treatment resistant depression by stimulating the dorsal lateral prefrontal cortex (dlPFC)5. However, recent evidence suggests that the dorsal medial prefrontal cortex (dmPFC) is a more promising target as it is more pivotal in emotional regulation6.
Salomons et al aimed to primarily investigate the role of functional connectivity between the dmPFC and the subgenual cingulate cortex as a predictor of rTMS treatment outcome4. Secondarily, they aimed to investigate the changes in connectivity between these regions and its relation to treatment response. Thus, the focus of this review is to discuss the significance of the results obtained by Salomons et al in predicting treatment outcomes and the role this may play in improving treatment efficiency and effectiveness.
Introduction and Background Over the course of 4 weeks, 25 treatment-resistant individuals with unipolar (n=21) or bipolar (n=4) depression underwent 20 rTMS sessions of 10Hz stimulation to the dmPFC. Pre- and post-treatment fMRI analysis was performed, and the Hamilton Depression Rating Scale (HAMD17) was used to assess depressive symptoms. Following treatment with rTMS, there was a staggering 45% decrease in mean HAMD17 scores, from 21.3 to 12.0. Response prediction using baseline connectivity Higher baseline connectivity between the dmPFC and subgenual cingulate cortex was positively correlated with greater improvement in HAMD17 scores following
treatment (Figure 1). In contrast, there was a negative correlation between dmPFC-thalamus and dmPFC-putamen baseline connectivity and positive rTMS treatment response.
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Figure 1. Comparison of baseline functional connectivity from the dmPFC and improvement in HAMD17 scores.
As shown in Figure 2, positive treatment outcome was correlated with higher baseline functional connectivity between the sgACC and dlPFC. In addition, lower baseline functional connectivity between the sgACC and the insula and amygdala was shown to underlie improvement in HAMD17 scores following rTMS treatment.
Figure 2. Comparison of baseline functional connectivity from the sgACC and improvement in HAMD17 scores.
Changes in functional connectivity over the course of treatment A decrease in functional connectivity between the dmPFC and the insula over the course of treatment was correlated with
successful response to rTMS. Conversely, an increase in functional connectivity between the dmPFC and the thalamus was shown to result in increased HAMD17 scores following treatment. (Fig. 3).
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Figure 3. Correlation between changes in functional connectivity with the dmPFC as a result of rTMS treatment and improvement on HAMD17 scores following treatment.
Additionally, decreased functional connectivity between the sgACC and both the midcingulate cortex (MCC) and the caudate was associated with more positive rTMS treatment outcomes (Fig. 4).
Figure 4. Correlation between changes in functional connectivity with the sgACC as a result of rTMS treatment and improvement on HAMD17 scores following treatment.
Significance/Future Directions: Due to the debilitating nature of MDD, it is prudent that effective treatments be developed in order to facilitate patient recovery. The emergence of personalized medicine in the treatment of depression has created an option for those patients with refractory depression, who have failed at least two previous rounds of pharmacological therapy. Salomons et
al were able to show that the baseline functional connectivity between specific regions of the brain may be used as biomarkers, as the strength of the connectivity between these areas was able to predict the efficacy of rTMS treatment to the dmPFC. Moreover, it was shown that certain alterations in connectivity following rTMS therapy were associated with the greatest improvements in depressive symptomatology. As patients typically receive and fail numerous treatments
before finding an effective one7, the use of biomarkers to develop a therapy tailored for the specific patient provides a window of opportunity to decrease the number of patients who present with treatment resistant depression. Removing the speculation of treatment efficacy would ultimately reduce patient wait times, and help to minimize the global disease burden of depression. In order to provide the most comprehensive picture of the use of biomarkers in the treatment of MDD, future studies should include a population of patients with exclusively unipolar depression. Additionally, it has been demonstrated that rTMS stimulation with a frequency of 10 hertz produced a high degree of inter-individual variability between inhibition and facilitation of populations of neurons8. Thus, the use of different frequency parameters (1 hertz, 15 hertz and 20 hertz) should be examined in future studies to examine if the specific stimulation frequency used has an impact on the overall efficacy of the rTMS treatment.
References 1. Collins, P. Y. et al. Grand challenges in global mental health. Nature, 475, 27–30 (2011). 2. Berlim, M. T., & Turecki, G. Definition, assessment, and staging of treatmentresistant refractory major depression: a review of current concepts and methods. Can. J. Psychiat. 52(1), 46–53 (2007). 3. Lui, S. et al. Resting-state functional connectivity in treatment-resistant depression. Am. J. Psychiat 168, 642-648 (2011).
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4. Salomons, T. V. et al. Resting-state cortico-thalamic-striatal connectivity predicts response to dorsomedial prefrontal rTMS in major depressive disorder. Neuropsychopharmacol. (August), 1–39 (2013). 5. Leone, A. et al. Early report Rapid-rate transcranial magnetic stimulation of left dorsolateral prefrontal cortex in drugresistant depression. Lancet 348, 233–237 (1996). 6. Downar, J., & Daskalakis, Z. J. New targets for rTMS in depression: a review of convergent evidence. Brain. Stimul. 6(3), 231-240 (2012). 7. Spielmans, G. I., Berman, M. I., & Usitalo, A. N. Psychotherapy versus secondgeneration antidepressants in the treatment of depression: a meta-analysis. J. Nerv. Ment. Dis. 199(3), 142–9 (2011). 8. Maeda, F. et al. Interindividual variability of the modulatory effects of repetitive transcranial magnetic stimulation on cortical excitability. Exp. Brain. Res. 133(4), 425–430 (2000).
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