Anterior Cingulate Cortex as Therapeutic Target for Autism Spectrum Disorder

Anterior Cingulate Cortex as Therapeutic Target for Autism

Yunqing Zhu

Autism spectrum disorder is a neurodevelopmental disorder in which social deficits is one of the core symptoms. However, there is currently no effective treatment targeting this symptom. In the study by Guo et al. (2019), the author used Shank3 knockout mice as animal model for autism, and observed the structural and functional changes in the anterior cingulate cortex. They found that Shank3 mutations caused morphological and functional change in pyramidal neurones of the region, and led to social deficits (Guo et al. 2019). They then enhanced the pyramidal neurone activity in anterior cingulate cortex using optogenetics, chemogenetics and a pharmacological compound, and were able to rescue the social impairments (Guo et al. 2019). These findings provide a causal link between hypoactivity of the anterior cingulate cortex and social impairments in autism, and proves that the area could be a potential therapeutic target (Guo et al. 2019).

Background and Introduction

Autism spectrum disorder (ASD) is a neurodevelopmental disorder that affects about 1% of individuals globally (Lai, Lombardo, and Baron-Cohen 2014). Although it greatly impacts the quality of life of affected individuals, there is currently no effective pharmacological treatment for the core symptoms of the disorder. This is in part due to the heterogeneity and lack of understanding of the underlying mechanisms of the disorder. Instead, available treatments focus on socio-behavioural interventions or treating symptoms of comorbid disorders.

Social communication deficit is one of the core diagnostic criteria of ASD. Successful social interactions and behaviour require effective processing and integration of social inputs from various sources (Guo et al. 2019). The anterior cingulate cortex (ACC) is an association cortex involved in multiple cognitive processes including learning and error monitoring (Apps, Rushworth, and Chang 2016). Anatomically, the ACC is highly conand information processing (Apps, Rushworth, and Chang 2016). Numerous neuroimaging studies have demonstrated altered neural circuitry in ACC of ASD patients compared to healthy controls ( Balsters et al. 2016; Zhou et al. 2016; Zikopoulos and Barbas 2013). Furthermore, studies in both humans and monkeys found that the ACC is activated when engaging in economic games involving the need for social cooperation, decision making based on social information and making predictions based on others’ behaviours (Gabay et al. 2014; Haroush and Williams 2015). Laidi et al. (2019) also found through a magnetic resonance imaging (MRI) study that adults with ASD display decreased cortical thickness in ACC. Thus, it can be concluded that ACC is highly implicated in various social behaviours.

Major Findings Global deletion of Shank3 caused morphological and functional deficits in excitatory pyramidal neurones of the ACC, resulting in social deficits

Using fluorescent in situ hybridization and immunofluorescent staining, Guo et al.(2019) found that Shank3 mRNA is mainly expressed in the excitatory pyramidal neurones in the ACC. Therefore, the authors focused on the morphological and function changes caused by Shank3 mutation in these ACC pyramidal cells in their study. Green fluorescent protein (GFP) was delivered to the ACC of wild-type (WT) and Shank3 KO mice to nected to regions in the brain important for social behaviours

visualize the pyramidal neurones in the region(Guo et al. 2019). It was found that the neurones in KO mice showed decreased dendritic complexity, spine density as well as reduced number of mushroom-shaped spines compared to WT(Guo et al. 2019). In addition, electron microscopy showed decreased average length and thickness of postsynaptic densities in the pyramidal cells in ACC of KO mice(Guo et al. 2019).

Guo et al. then measured the post-synaptic activities of the ACC pyramidal neurones in KO and WT mice using whole cell patchclamp recording. Results showed weaker a-amino-3-hydroxy-5- methylisoxazole-4-propionic acid receptor (AMPAR)-mediated postsynaptic transmissions in KO mice as well as a reduced AMPAR/N-methyl-D-aspartate receptor (NMDAR) ratio(Guo et al. 2019). Higher intensities of presynaptic stimulations were required to generate an action potential postsynaptically(Guo et al. 2019). Western blot analysis also showed reduced expression of the AMPAR subunit GluR1 (Guo et al. 2019). These results suggest that AMPAR dysfunctions are the predominant cause of functional deficits in the ACC pyramidal neurones of KO mice (Guo et al. 2019). Furthermore, paired training paradigm failed to induce long-term potentiation (LTP) in KO mice as it did in the WT (Guo et al. 2019).

Previous studies have established the important role ACC plays in integration social information and regulating behaviours. However, the underlying neural mechanisms that cause the social deficits are unknown. Moreover, previous studies had not been able to establish a causal link between specific ACC dysfunctions with ASD-related social deficits. Therefore, Guo et al. (2019) used a Shank3 mice model for ASD to test whether dysfunctions in ACC directly leads to social deficits. The team found that Shank3 knockout (KO) mice showed aberrations in neuronal morphology as well as synaptic functions in their ACC excitatory pyramidal neurones and that selective deletion of Shank3 in the ACC was sufficient to generate the social deficits shown in KO mice (Guo et al. 2019). Furthermore, they found that increasing the excitatory activities in the ACC by using optogenetic and chemogenomic techniques as well as pharmacological agents successfully rescued the social deficits (Guo et al. 2019). As a result, the author concluded that hypoactivity in the ACC directly causes social impairments in ASD, and that the ACC should be further researched as a potential therapeutic target for ASD (Guo et al. 2019). Behavioural tests were used on the KO and WT mice. In particular, three-chamber test and home cage social interaction test both showed significant social deficits in KO mice compared to WT (Guo et al. 2019). Elevated plus maze and open field tests also resulted in higher anxiety levels in the KO mice (Guo et al. 2019). Then, the social interaction task was repeated with fiber photometry systems implanted into the mice to measure the real-time calcium activities as the mice approached the novel mice in their home cage. KO mice showed decreased ACC excitatory activity during this task compared to WT (Guo et al. 2019).

Conditional Knockout (CKO) of Shank3 gene in the ACC region alone was sufficient to generate hypoactivity in pyramidal neurones and social deficits

Clustered regularly interspaced short palindromic repeats (CRISPR)-Cas9 genome-editing technique was used to selectively knock out Shank3 in the ACC of the mice models (Guo et al. 2019). As a result, this generated similar patterns of aberrations 202

in synaptic activity in the pyramidal cells as in KO mice, including decreased postsynaptic activities, decreased AMPAR/ NMDAR ratio, as well as decreased expressions of AMPAR and NMDAR subunits (Guo et al. 2019). Three-chamber test and social interaction tests were again used on the CKO, and both showed significant social deficits (Guo et al. 2019). These results indicate that deletion of Shank3 in ACC alone was sufficient to cause functional and social deficits in mice models, suggesting causal link between decreased excitatory activities in ACC and social deficits seen in ASD (Guo et al. 2019). Guo et al. (2019) found their results of abnormal morphology in the ACC to be consistent with the previous studies that found altered anatomical characteristics, decreased volume or cortical thickness in the ACC of ASD patients (Haznedar et al. 2000; Laidi et al. 2019). It was well established that ACC plays an important role in social behaviours and that the general region is implicated in ASD. However, Guo et al. (2019) expanded beyond these findings in that they identified the specific neural substrates involved in the ACC changes, and proved experimentally that these dysfunctions directly cause the social impairments seen in ASD. They also demonstrated that these deficits could be rescued by enhancing the activities of ACC pyramidal neurones (Guo et al. 2019).

Critical Analysis

In the study by Guo et al. (2019), the authors looked at whether dysfunctions in ACC pyramidal neurones were sufficient to cause ASD-like social communications symptoms in Shank3 model mice and established that reactivation of the ACC pyramidal neurones ameliorated these symptoms. However, Guo et al. (2019) recognised that the level of photostimulation they applied using optogenetics in this study may be outside physiological levels of stimulation in the brain. As they found that lowest levels of photostimulation did not achieve significant imenough to ameliorate symptoms in human brain might not be

Enhancement of ACC excitatory activities rescued synaptic functional and social deficits

As the causal link between hypoactivity of ACC and ASD-like social deficits were established, the authors then explored whether using optogenetic, chemogenetic and pharmacological methods to enhance the activity of ACC pyramidal neurones would rescue these social deficits. The results showed that optogenetically enhancing ACC excitatory activities led to increased sociability shown in behavioural tests in both KO and WT mice as well as reduced anxiety (Guo et al. 2019). On the other hand, optogenetic inhibition of the same neurones led to social deficits in WT mice similar to those exhibited by KO mice (Guo et al. 2019). Selective activation of ACC pyramidal neurones using designer receptors exclusively activated by designer drugs (DREADDs) also improved social behaviours (Guo et al. 2019). In addition, both selective re-expression of Shank3 in ACC and the injection of AMPAR-positive allosteric modulator (PAM) CX546 improved synaptic functions on top of sociability est in ASD research. Now that the causal relationship between ACC under-excitation and social deficits is established, future volved in the area. As Guo et al. (2019) pointed out that AMPAR

(Guo et al. 2019).

Conclusion/Discussion

Guo et al. ( 2019) concluded that mutations in Shank3 cause dysfunctions in pyramidal neurones of ACC, directly leads to hypoactivity in the area and in turn causes social impairments. However, optogenetically and chemogenetically activating the pyramidal neurones of ACC effectively rescued these social deficits. Since AMPAR-related dysfunctions was the main cause of synaptic hypoactivity, using an AMPAR-PAM also significantly ameliorated the social impairments. Thus, ACC can be consid

provements in sociability, it might be that effective intensity of stimulation symptoms of the pyramidal neurones which is attainable (Guo et al. 2019).

Furthermore, Guo et al. (2019) mentioned that AMPAR could be a potential therapeutic target as hypoactivity in the ACC due to AMPAR dysfunctions lead to the social deficits. However, ASD is also characterised by over-excitation of neurones in many other brain regions which may lead to symptoms such as hypersensitivity to sensory inputs. Although an AMPAR-PAM might be useful in enhancing activity of the ACC, if applied to humans, it would need to be delivered to target the ACC specifically to avoid overstimulating other areas of the brain.

Future Direction

The study by Guo et al. (2019) points to ACC as an area of interstudies could look at the specific molecular mechanisms inered as a potential therapeutic target for ASD.

is a main factor in the ACC dysfunctions, genomic sequencing could be done to screen for potential over- or underexpression of genes related to the receptor. It would also be useful to map out the circuitry that the pyramidal neurones in ACC are involved in. This would include looking at what other areas and the types of neurones that these pyramidal neurones are connected to.

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