Cook, MN*, Hartless, CB and Lester, DB.

Stress may contribute to the development and/or emergence of neuropsychiatric disorders, including those related to changes in reward and motivation. Anhedonia is a common outcome of chronic unpredictable stress (CUS) paradigms and may be indicator of resilience or vulnerability to the effects of chronic stress. Here, we exposed male and female C57BL/6J (B6) and DBA2/J (D2) mice to four weeks of chronic unpredictable stress, then used fixed potential amperometry to examine stimulated dopamine (DA) release, half-life, and auto receptor functioning. Overall, D2 mice exposed to CUS tended to have increased dopamine release compared to their control counterparts or B6 mice. D2 mice exposed to stress also had a shorter DA half-life than D2 control mice. CUS tended to decrease DA autoreceptor functioning in B6 but not D2 mice. Finally, CUS increased DA half-life in D2 mice following administration of nomifensine, a DA agonist. These strain-related differences in the effects of CUS on autoreceptor functioning and DA half-life may help in understanding reward- and motivation-related changes following exposure to chronic unpredictable stress.

Department of Psychology, The University of Memphis, Memphis, TN USA.

Funding support: Dunavant Professorship

Renee Papaluca1,2, Eva Guerrero-Hreins1,2, Aneta Stefanidis4, Claire Foldi4, Brian Oldfield4, Priya Sumithran*3, Robyn Brown*1

Bariatric surgery is the most effective long-term obesity treatment, reducing appetite and improving glycaemic control. These benefits are due to profound changes to the gut-brain axis. However, a small patient subset experience adverse mental health outcomes post-surgery, such as depression. While longitudinal studies begin to explore this, research exploring how gut-brain axis alterations could mechanistically contribute to these adverse mental health outcomes post-surgery is lacking. Therefore, this study investigates individual differences in depressive-like behaviour after vertical sleeve gastrectomy (VSG) in mice. Methods: Male and female C57BL/6 mice (n=57) were fed a high-fat, high-sugar diet (11 weeks) before VSG or sham surgery. Depression-like behaviour was assessed pre- and post-surgery. Based on test performance, VSG mice were then separated into tertiles and the upper and lower tertiles were used as ‘depression-susceptible and ‘depressionprone’ mice respectively. From these subgroups, RNA was extracted from tissue punches of the dorsal striatum and nucleus accumbens, key regions implicated in major depressive disorder and subjected to bulk RNA-sequencing. Results: VSG led to significant, sustained weight loss and reduced food intake. Differential gene expression analysis revealed downregulated neuroinflammation and tight junction-related genes in VSG depression-susceptible mice. Further gene set enrichment analysis displays an upregulation of TNF-α-NF-κB and PI3K-AKT/mTOR pathways. Additional chow-diet cohort data is pending, with preliminary findings to be presented at the meeting. Conclusion: This study provides insight into the possible neurobiological mechanisms underlying post-bariatric surgery mental health outcomes, shedding light on potential new biomarkers for depression risk associated with this common weight-loss procedure.

1University of Melbourne, School of Biomedical Sciences, Department of Biochemistry and Pharmacology, 2Florey Institute of Neuroscience and Mental Health, 3Department of Surgery, School of Translational Medicine, Monash University, 4Department of Physiology, Monash University.

*equal contribution Introduction:

CA Moul, 1

While there is considerable evidence for a role of the serotonin system in the aetiology of antisocial and aggressive behaviour, parsing out the mechanisms responsible for this relationship have remained elusive. The body of work presented here focusses on evidence to link the serotonin 1B receptor gene with callous-unemotional (psychopathic) traits and proposes a cognitive mechanism that may explain the relationship.

First, evidence is provided from genetic and epigenetic studies of clinical samples of children and adults with antisocial behaviour problems to demonstrate an association between HTR1B and callous-unemotional (psychopathic) traits. Next, a theory of amygdala function in psychopathy is used to provide the rationale for animal studies that tested the role of serotonin 1B in basic aspects of associative learning (response-reversal). Finally, the translation of this to a human study shows how callous-unemotional (psychopathic) traits in healthy, non-criminal young adults are associated with a deficit in the allocation of attention to, and concurrent encoding of, outcome stimuli.

Data from these three lines of work will be presented and entwined with a narrative that seeks to emphasise the importance of cognitive endophenotypes in genetic association studies of psychopathology. These data will illustrate how deviations in simple cognitive mechanics (e.g. attention and associative learning) can cause potentially dramatic variations in social behaviour. Missing pieces of the puzzle and future directions will be discussed.

1 School of Psychology, University of Sydney

Funding Support: Australian Research Council DP200102371

AR McEwan 1, G Hutchings 1, A Rattray 1, C Murgatroyd 2, A MacKenzie 1

Human development and health depend on the precise expression of genes in specific cells, at the right times, and in response to appropriate stimuli, known as context-dependent gene regulation. This regulation involves a variety of DNA elements called context-dependent cis-regulatory elements (cdCREs), which may constitute up to 10% of the genome. These include enhancers, silencers, and promoters that are crucial for complex multicellularity. Vertebrates share many phenotypic similarities, cell types, and gene expression patterns, often regulated by conserved cdCREs. The regulation of these genes has been implicated in the progression of disorders in humans. Obesity and anxiety have significantly impacted society during the COVID-19 pandemic. Understanding the role of cdCREs behind these conditions can improve our quality of life and resilience. We identified the BE5.1 region within the BDNF gene as of interest, being associated with obesity (rs10767664; p = 4.69 × 10^–26). The major T-allele of BE5.1 acted as an enhancer, while the obesity-linked A-allele did not. Deleting BE5.1 in mice showed no significant effect on BDNF expression in the hypothalamus or weight gain but increased anxiety-like behaviour in female mice, reversible by diazepam. Human studies also linked the rs10767664 polymorphism to anxiety. The findings suggest a novel mechanism regulating BDNF expression, affecting mood and anxiety in both mice and humans.

1. Institute of Medical Sciences, University of Aberdeen, Ashgrove Road West, Aberdeen, Scotland. AB25 2ZD, United Kingdom
2. Manchester Metropolitan University, Ormond Building, Lower Ormond Street, Manchester, M15 6BX, United Kingdom

J. Almeida¹, H. Chen¹, E. Legge¹, M. Sutter², and A.W. Lasek¹.

Perineuronal nets (PNNs) are specialized extracellular matrix structures that primarily surround parvalbumin (PV)-expressing GABAergic interneurons in cortical regions. We previously showed that binge-like ethanol consumption resulted in increased accumulation of PNNs in the insular cortex (insula) and that PNNs in the insula regulate aversion-resistant ethanol consumption, as measured by adulteration of ethanol with quinine, in male mice. To investigate mechanisms contributing to increased brevican accumulation following chronic ethanol exposure, C57BL/6J mice underwent 6 weeks of binge-like drinking in the drinking in the dark (DID) procedure. Western blots of insula protein lysates showed reduced brevican cleavage product following chronic DID, suggesting reduced proteolytic activity of matrix metalloproteinases (MMPs). We performed qPCR to measure transcript levels of MMPs and their endogenous inhibitors (TIMPs). Timp3 transcript levels were increased in the insula after chronic DID, consistent with reduced activity of MMPs. We next investigated whether brevican is involved in ethanol consumption. Since brevican cell-autonomously regulates the excitability of PV neurons, we generated mice with knockout of the brevican gene (Bcan) in PV neurons (Bcan^PVKO) to test the hypothesis that brevican may regulate aversion-resistant ethanol consumption. Male Bcan^PVKO exhibited decreased aversion-resistant ethanol consumption when compared to their wild-type counterparts. We also found that knockout of Bcan in anterior insula neurons (Bcan^insnKO) by injecting AAV-hSyn-EGFP-Cre into the insula of male homozygous Bcan floxed mice resulted in reduced aversion-resistant ethanol consumption. Interestingly, female Bcan^PVKO and Bcan^insnKO mice did not exhibit this phenotype, suggesting a sex-specific role for brevican in insular cortex neurons in aversion-resistant ethanol consumption.

¹Department of Pharmacology and Toxicology, Virginia Commonwealth University, Richmond, VA, 23298, USA.

²Department of Psychiatry, University of Illinois at Chicago, Chicago, IL, 60612, USA.

Funding support: NIAAA U01AA020912 and R01AA027231.

CL Barr 1,2,3, Y Feng 1, KG Wigg 1G

Genome wide association studies (GWAS) have implicated the OTX2 (Orthodenticle homeobox 2) gene locus in major depressive disorders (MDD) as well as genetically correlated traits (alcohol consumption, smoking initiation, chronotype). Of the genes identified by MDD GWAS, the gene for the transcription factor OTX2 stands out as it is responsible for both opening and closing of critical and sensitive brain periods. These are developmental periods where the brain is more sensitive to environmental input and are critical for normal brain development. Evidence suggests that the brain may also be more sensitive to negative environmental impact during sensitive periods. Critically, human and animal models both specifically implicate OTX2 gene expression in the response to stress and risk for depression. Based on the genetic findings, and the potential role of OTX2 as a mediator of environmental risk for depression, we identified genes regulated by OTX2 in human neural precursor cells (NPCs) using CRISPR activation (CRISPRa) to increase expression. We identified 17 significantly differentially expressed genes, including OTX2 which was increased 4-fold. In addition to OTX2, 4 genes of the 17 have been directly implicated in depression/depressive behaviours from human and animal studies (GPER1, VGF, TAFA5, P3H2). GPER1 has been indicated as serum biomarkers for depression. Additional differentially expressed genes are involved in processes implicated in depression (e.g. neurogenesis, neuroplasticity, response to stress). These novel findings link OTX2 expression with genes previously implicated in depression from human and animal studies, suggesting OTX2 as a master regulator of depression risk.

1 Division of Experimental and Translational Neuroscience, Krembil Research Institute, University Health Network, Toronto, Ontario, Canada

2 Program in Neuroscience and Mental Health, Hospital for Sick Children, Toronto, Ontario, Canada

3 Departments of Psychiatry and Physiology, University of Toronto, Toronto, Ontario, Canada

Funding Support: Labatt Family Chair in Depression Biology in Children

Youssef S¹, Johal K¹, Stefanelli G¹.

Neurogenesis and neurodevelopment are complex processes that heavily rely on dynamic epigenetic mechanisms and their ability to regulate gene expression. Recently, a role for histone variants and their dynamics has been shown to be crucial for neurodevelopment. Our lab is focused on Anp32e, a histone chaperone specific for H2A.Z variant. Notably, Anp32e has been shown to regulate H2A.Z levels in the genome and contribute to higher brain functions like memory formation. In addition, the lack of Anp32e has been associated with impaired dendritic morphology and downregulation of genes involved in neuronal development, highlighting its critical role in development. Yet, Anp32e role in neurodevelopment remains unexplored. Here, we use a knock-out mouse model to investigate how Anp32e regulates H2A.Z distribution and gene expression during critical stages of embryonic cortical development. We show that knock-out of Anp32e result in accumulation of H2A.Z throughout cortical development, severely impacting gene expression. In addition, we will perform behavioral assays to investigate if the epigenomic and transcriptomic alterations result in altered adult behaviors. Overall, our studies investigate for the first time the contribution of a new epigenetic mechanism involving H2A.Z and Anp32e to neurodevelopment and the regulation of adult behaviors.

¹ University of Ottawa, Department of Biology, Ottawa, ON, Canada

WC Booher^1,2, K Lanier¹, K Doenges¹, J Manke¹, M Armstrong¹, L Saba^1, N Reisdorph¹, RA Radcliffe^1,2

Oral consumption of Cannabis carries many of the same well-documented negative consequences associated with smoking Cannabis, along with additional health concerns specific to oral ingestion. It is essential to gain a deeper understanding of the factors contributing to the health risks of oral consumption, including individual differences in how people metabolize delta-9-tetrahydrocannabinol (∆9-THC), which is likely influenced by genetic factors. We hypothesized that genetically distinct mouse strains would exhibit variations in metabolism of ∆9-THC. To test this hypothesis, we administered 10 or 15 mg/kg ∆9-THC via oral gavage to four mouse strains (A/J, C57BL/6J, CAST/EiJ, and NZO/HILtJ) that are among the founders of two well-characterized experimental genetic mouse populations, the Collaborative Cross recombinant inbred strains and the Diversity Outbred heterogenous stock mice. Blood was collected at 60-, 90-, and 240-minutes post administration and plasma levels of ∆9-THC and its primary metabolites were measured using mass spectrometry. There were significant differences between the A/J and C57BL/6J ∆9-THC plasma levels and some of the metabolites, with few notable sex differences. The overall pattern of these differences suggest that C57BL/6J mice metabolize ∆9-THC more rapidly than A/J mice. CAST/EiJ and NZO/HILtJ mice exhibit significant differences in ∆9-THC plasma concentrations, but not other metabolites. To our knowledge, this is the first study to include A/J, CAST/EiJ, and NZO/HILtJ mice in a ∆9-THC oral administration mouse model. These findings underscore the need for further research to explore genetic differences in the pharmacokinetics of ∆9-THC in preclinical models of oral Cannabis consumption.

¹Skaggs School of Pharmacy and Pharmaceutical Sciences, University of Colorado Anschutz Medical Campus, Aurora, CO 80045

²Institute for Behavioral Genetics, University of Colorado, Boulder, CO

Funding support: Skaggs School of Pharmacy and Pharmaceutical Science ADR Seed Grant Program, Institute of Cannabis Research, and NIH-NCRR grant 1S10OD010366-01A1

Emmanuel S. Onaivi1 , Anna Bukiya2 , Venkat Sharma1 , Shilpa Buch3

There is an explosion of new knowledge that the effect of cannabis is encoded in our genome on chromosomes 1 and 6, and the biological effects is mediated by action on human endocannabinoid system (ECS) that are involved in reproduction and implantation of fertilized egg in the uterus. The identification of additional lipid mediators, enzymes and receptors is expanding the ECS, to endocannabinoidome (eCBome). Together with the ECS, eCBome are composed of genes coding for CB1 and CB2 cannabinoid receptors (CB1Rs, CB2Rs), endocannabinoids (eCBs), and their metabolic enzyme machinery for their biosynthesis and catabolism. The remarkable progress and new understanding are unravelling the role of the eCBome genetics, influence on behavior and providing deeper insights on the biological actions of cannabinoids in many disorders. CB2R conditional knockout (cKO) mice with deletion of CB2Rs from dopamine neurons, DAT-Cnr2 and those with deletion from microglia Cx3Cr1- Cnr2 were used and they displayed differential phenotypes. CB2Rs in dopamine neurons and microglia upregulated the expression of NLRP3 inflammasome pathway including NLRP3, and increased expression of proinflammatory cytokines, TNF-α, IL-6, IL-1α and IL-1β in the frontal cortices of the cKO mice following subacute treatment with 8% alcohol. The molecular pathways of fetal damage by alcohol, from cerebral artery to developmental delay highlights the functional presence and interaction of the ECS in baboons at different stages of development, including the effects of simultaneous alcohol and tetrahydrocannabinol on cerebral artery. An insight into the emerging eCBome genetics and neuroimmune crosstalk offers opportunities for continued preclinical and clinical studies.

1Biology department, College of Science and Health, William Paterson University, Wayne NJ, 2University of Tennessee Health Science Center, Memphis, TN, 3University of Nebraska Medical Center, Omaha, Nebraska.