Conference Agenda

Christopher P. King^1,2, Nana K. Amissah¹, Brady M. Thompson¹, Elizabeth A. Rakowski¹, Luke T. Hannan¹, Apurva S. Chitre³, Thiago Sanches³, Beverly Peng³, Oksana Polesskaya³, Leah C. Solberg Woods⁵, Abraham A. Palmer^3,6, and Paul J. Meyer¹

Understanding the genetic factors that influence problematic drug use can identify neurogenetic targets for treating substance use disorder (SUD). We used an intermittent-access self-administration procedure in rats that triggers intense cocaine motivation (i.e., incentive sensitization) after only limited amounts of drug exposure and conducted a genome-wide association study (GWAS) of key traits obtained in this paradigm. We identified multiple genomic regions associated with escalation of cocaine intake and response to cocaine-cues, each containing several candidate genes. In a related project, we conducted a GWAS of food-cue reactivity, which we found to be phenotypically and genetically correlated with measures of cocaine and nicotine motivation. We then used genetic correlations, phenome-wide association studies (PheWAS), and transcriptomics to identify understudied genetic variants influencing traits in these GWASs. A region on chromosome 1 influenced responding for a cocaine-associated cue following two weeks of abstinence, which contains several genes that are expressed in the central nervous system (e.g., Far1) and dopaminergic midbrain (e.g., Spon1). PheWAS indicated that this region also influenced motivation to self-administer cocaine during a progressive ratio test. One gene that associated with food-cue reactivity (Tenm4) was of particular interest due to its identification in human GWASs of SUD and its involvement in synaptic connectivity. Intracranial administration of a Tenm4-associated protein reduced several measures of cocaine motivation, confirming a pleiotropic effect of this gene on cocaine motivation and cue-reactivity. Thus, these studies demonstrate the utility of using HS rats to identify genes associated with complex vulnerability traits related to SUD.

¹Department of Psychology, University at Buffalo, Buffalo, USA.

²Clinical and Research Institute on Addictions, Buffalo, USA.

³Department of Psychiatry, University of California San Diego, La Jolla, USA.

⁴Department of Pharmacology, Addiction Science and Toxicology, University of Tennessee Health Science Center, Memphis, TN

⁵Department of Internal Medicine, Molecular Medicine, Center on Diabetes, Obesity and Metabolism, Wake Forest School of Medicine, Winston-Salem, USA.

⁶Institute for Genomic Medicine, University of California San Diego, La Jolla, USA.

Funding Support: National Institute of Drug Abuse (DA037844); Office of the Director, National Institutes of Health; and the National Institute on Alcohol Abuse and Alcoholism (AA024112).

KM Loedige¹, HHA Thorpe¹, JY Khokhar¹

Cell Adhesion Molecule 2 (CADM2) is a gene encoding a synaptic cell adhesion molecule primarily expressed in the mesolimbic system, crucial for synaptic organization, structural stability and signaling. Previous Genome-Wide Association Studies (GWAS) have linked Cadm2 polymorphisms with human behavioural traits including impulsivity, substance use, and executive function. Our prior investigation using a Cadm2 transgenic mouse model showed that Cadm2 expression influences cannabis self-administration, impulsive behaviours, and alters electrophysiological properties of prelimbic cortical neurons. To elucidate the molecular underpinnings of CADM2-mediated phenotypic changes, this study aims to assess transcriptomic alterations associated with Cadm2 knockout in mesolimbic regions using a transgenic mouse model.

Bulk RNA sequencing was employed on whole brain, frontal cortex, and striatal tissues of male and female Cadm2-/- (KO) and Cadm2+/+ (WT) C57BL/6J mice to identify differentially expressed genes (DEGs) within the mesolimbic system. Preliminary findings reveal DEGs encompassing proteins involved in glutamatergic, GABAergic, cholinergic, serotonergic and dopaminergic neurotransmission, as well as voltage-gated and calcium-activated ion channels (p_adj ≤ 0.05). These DEGs are accompanied by enriched gene sets associated with synaptic structure and transmission, along with enriched pathways related to neurotransmitter synthesis (p_adj ≤ 0.05). To validate potential mechanisms underlying the contribution of Cadm2 expression to impulsivity and addiction vulnerability, a targeted analysis of DEGs will be conducted using rt-qPCR and western blotting, followed by behavioural assays probing for genotype-specific differences in executive function and reward-seeking behaviours. By delineating this neurobiological mechanism, this research holds promise for informing the development of novel strategies to address substance use disorders.

¹Department of Anatomy and Cell Biology, University of Western Ontario, London, ON, Canada

Funding Support: JYK is funded by a Canada Research Chair in Translational Neuropsychopharmacology from CIHR. HT is funded by a post-doctoral fellowship from CIHR.

Marcelo R. Zimmer1, Gustavo M. Santana2,3, Marcelo O. Dietrich1,4

The infant's relationship with their mother is vital for the survival of the offspring. The mother provides nutrients and sensorial stimulation necessary for normal development. Upon separation from the mother, mouse infants, like human babies, respond with distress, emitting vocalizations and recruiting thermoregulatory and stress responses. Here, we focus on understanding the variability of distress responses in founder strains of the Collaborative Cross Consortium, a panel of genetically diverse mouse lines. Upon maternal separation, these strains display highly diverse vocal profiles. For example, wildderived PWK/PhJ pups exhibit the lowest rate of distress calls throughout separation among all strains. Moreover, we identified different vocal signatures using supervised and unsupervised machine learning methods, with wild-type derived WSB/EiJ pups emitting more than 40% of chevron syllables compared to 8% in B6 pups. Importantly, vocal behavior does not correlate with the activation of the HPA axis. C57BL/6J pups, which have a relatively high rate of vocalizations, have low levels of corticosterone. Conversely, WSB/EiJ pups show higher levels of corticosterone despite exhibiting a low rate of distress calls. WSB/EiJ pups also display an enhanced thermogenic phenotype, suggesting stress-induced hyperthermia, contrasting with the C57BL/6J-suppressed thermogenesis. Our ongoing studies are integrating these behavioral and physiological readouts with whole-brain mapping of activated neurons. Our preliminary mapping data indicates that WSB/EiJ pups exhibit the highest levels of whole-brain activation, suggesting their brains are more responsive to separation. This comprehensive approach will help us understand how genetic diversity influences brain network activity and behavioral and physiological responses in infant mammals.

1 Department of Comparative Medicine, Yale University, New Haven, CT, USA 2 Interdepartmental Neuroscience Program, Yale University, New Haven, CT, USA 3 Program in Physics, Engineering and Biology, Yale University, New Haven, CT, USA 4 Department of Neuroscience, Yale University, New Haven, CT, USA

Suelen Baggio¹, Andrzej Nagalski¹, Kamil Koziński¹, Łukasz M. Szewczyk¹, Marcin A. Lipiec¹, Ksenia Meiza², Anna Goncarzewicz², Ludwika Szczepańska², Ewelina Knapska², Marta B. Wiśniewska¹

TCF7L2 is a transcriptional effector of the Wnt/β-catenin signaling pathway which controls developmental and homeostatic processes and is a risk gene for autism spectrum disorder (ASD). TCF7L2 is highly expressed in the developing and adult thalamus. Our previous research discovered the role of TCF7L2 in regulating the establishment of thalamocortical connections and electrophysiological maturation of thalamic neurons. The extent to which thalamic dysfunction influences the development of ASD remains little explored. We hypothesized that postnatal disruption of thalamocortical circuit due to TCF7L2 deficiency leads to autism-like behaviors. We investigated these hypotheses using mice with Tcf7l2 knockout (Tcf7l2 cKO) mediated by Cre recombinase whose expression was induced postnatally in thalamic neurons. We analyzed the behavioral profile of the Tcf7l2 cKO mice to assess cognition (Intellicage), anxiety (open field, grooming, dark-light box), repetitive behaviors (marble-burying test), and social performance (Eco-HAB and Three-chamber). Tcf7l2 cKO mice showed no cognitive deficits, nor did they display differences in compulsive behavior. However, they exhibited a decrease in social interest, as evidenced by reduced preference to interact with a mouse over an object, and spending less time near the social scent. This behavior does not seem to be driven by anxiety because Tcf7l2 cKO mice showed a significant decrease in grooming behavior and a tendency to spend more time in an open arena. These results corroborate a hypothesis that thalamic dysfunctions originating from perinatal development can be a primary cause of social deficits, and impairments in thalamocortical circuits play a role in the pathogenesis of ASD.

¹ Laboratory of Molecular Neurobiology, Centre of New Technologies, University of Warsaw, Warsaw, Poland

² Laboratory of Emotions Neurobiology, Nencki Institute of Experimental Biology, Warsaw, Poland

Tengfei Wang¹, Thiago Sanches², Angel Garcia Martinez¹, Apurva S Chitre², Shangying Leng¹, Caroline Jones¹, Tomomi Green¹, Katie Holl³, Celine St Pierre², Hannah Bimschleger², Jianjun Gao², Riyan Cheng², Oksana Polesskaya², Leah C Solberg Woods³, Abraham A Palmer^2,4, Hao Chen¹

The social environment significantly influences cigarette smoking behavior. To model this in rodents, we established a socially acquired nicotine intravenous self-administration (IVSA) procedure in adolescent rats. In this model, nicotine delivery is paired with a contingent flavor cue that includes both odor and taste. The transfer of the nicotine-associated odor cue is crucial for initiating nicotine self-administration. We conducted a genetic association study on socially acquired nicotine IVSA using a population of outbred adolescent heterogeneous stock rats (n=1995, balanced for sex, IVSA started on postnatal day 41). Each rat was genotyped at 5,489,880 sites. We analyzed 63 phenotypes (heritability range: 0.05 to 0.26, mean: 0.165, SD: 0.04) and found 50 genome-wide significant loci linked to various aspects of nicotine IVSA. Among them, 30 contain genes with human orthologs related to smoking behaviors. For instance, total nicotine intake in rats is associated with Chr16:78175034, which harbors Arhgef7, associated with PackYears (lifetime cigarette consumption) in human GWAS. Additionally, PheWAS revealed shared genetic influences between nicotine motivation and social or anxiety-related phenotypes. For example, chr4:151349337 was associated with time spent in a social zone as well as nicotine intake during the first three IVSA sessions. These findings shed light on the intricate genetic control of nicotine motivation within a social context. Establishing the causality of these candidate genes could pave the way for successful treatments for nicotine addiction. This research was supported by NIDA grant P50DA037844.

1 Department of Pharmacology, Addiction Science and Toxicology, University of Tennessee Health Science Center, Memphis, TN, United States.

2 Department of Psychiatry, University of California San Diego, La Jolla, CA, United States.

3 Department of Internal Medicine, Wake Forest School of Medicine, Winston Salem, NC, United States.

4 Institute for Genomic Medicine, University of California San Diego, La Jolla, CA, United States.