Conference Agenda

Cheryl Reed¹, Grant Moen¹, Sahana Srinivasan¹, Shauna Rakshe², Suzanne Fei², Jason Erk¹, Tamara Phillips^1,3

Selective breeding for methamphetamine (MA) intake identified a null mutation in the trace amine-associated receptor 1 (Taar1) gene that negates TAAR1 function and increases genetic risk for MA intake in mice. A knock-in (KI) was produced on an isogenic C57BL/6J background, replacing the reference Taar1⁺ allele with the mutant Taar1^m1J allele. Increased MA intake in Taar1^m1J mice was verified. Here, we examined the effects of Taar1^m1J on MA-induced locomotor stimulation and conditioned taste aversion (n=15-16 and 6-9 mice/genotype/sex/dose, respectively) and characterized the transcriptome using RNA-sequencing data from MA-naïve mice (n=12 mice/genotype/sex) for the nucleus accumbens, prefrontal cortex, and ventral midbrain (VMB). The allele replacement did not alter MA-induced locomotor stimulation but attenuated MA-induced conditioned taste aversion (2 and 4 mg/kg MA; ps<0.001). Transcriptome analysis identified 1,326 differentially expressed genes (p<0.05), with 56 shared across brain regions. QIAGEN Ingenuity Pathway Analysis identified 265 enriched pathways, three of which included Taar1 (NF1 RAS Signaling Pathway, Phagosome formation, and Cellular Effects of Sildenafil). Weighted gene co-expression network analysis identified modules that differed (ps < 0.05) between Taar1 genotypes only in the VMB. Enriched gene ontology processes included: neuron ensheathment, myelination, glial cells, synaptic transmission, synaptic assembly, synaptic regulation, and neural development/regulation. Nineteen hub genes were identified including: Litaf, Trp53inp2, Insc, Gltp, Tmprss5, Gpr37, Car14, Ids, Lonrf2, Camsap2, Dzank1, Ppp1r9a, Arhgap20, Cmtm4, Sox10, Fa2h, Plekhg3, Tmem63a, and Jup. The underlined hub genes were previously found to be involved in MA-related behavioral and molecular changes. These processes may underlie TAAR1-mediated MA aversion impacting MA intake.

1. Department of Behavioral Neuroscience and Portland Alcohol Research Center, Oregon Health & Science University, Portland, OR, United States

2. Bioinformatics & Biostatistics Core, Oregon National Primate Research Center, Oregon Health & Science University, Beaverton, OR, United States

3. VA Portland Health Care System, Portland, OR, United States

Support provided by NIH/NIDA R01DA046081, R01DA057420, and U01DA041579; NIH/OD P51OD011092 and S10OD034224; Department of Veterans Affairs IK6 BX006342

AW Lasek1, M Galan-Llario1, H Chen1, E Legge1, J Almeida1, L Carvalho1, CM Erikson2, R Vlkolinsky2, M Bajo2, and M Roberto2

STAT3 is a transcription factor activated downstream of cytokine, chemokine, and growth factor receptor stimulation. It plays an important role in the innate immune response by promoting astrocyte reactivity and neuroinflammation in response to tissue injury. STAT3 is activated in several brain regions following chronic ethanol exposure in mice, rats and humans. To determine if astrocyte-expressed STAT3 regulates ethanol consumption, we generated conditional astrocyte Stat3 knockout (Stat3aKO) mice. Male, but not female, Stat3aKO consumed less ethanol than controls. To determine potential mechanisms contributing to decreased ethanol consumption in the Stat3aKO mice, we recorded glutamate receptor-mediated spontaneous excitatory post-synaptic currents (sEPSCs) in prelimbic pyramidal neurons in male Stat3aKO and control mice. We found a significant increase in the mean baseline sEPSC amplitude, but not frequency, in Stat3aKO mice. As astrocytes are involved in uptake of synaptic glutamate, we hypothesized that STAT3 might regulate the expression of glutamate transporters and measured transcript levels of Slc1a2, Slc1a3, Slc7a11, and Slc17a8 in Stat3aKO and control mice. Slc1a2 and Slc17a8 were significantly decreased in both male and female Stat3aKO mice, although the magnitude of the decrease was greater in males. These results suggest impaired glutamate clearance in male Stat3aKO mice, and that STAT3 in astrocytes promotes binge drinking. In conclusion, this study links neuroimmune signaling in astrocytes to cortical excitatory neurotransmission and ethanol consumption.

1Department of Pharmacology and Toxicology, Virginia Commonwealth University, Richmond, VA, USA, 2Department of Translational Medicine, The Scripps Research Institute, La Jolla, CA, USA

Funding support: NIAAA U01AA020912 and R01AA027231 to AWL; U01AA013498 and P60AA006420 to MR; T32 AA007456 to CME.

Meshi Micheala¹, Levi Mali¹ & Shohat-Ophir Galit¹

Motivation is an internal representation of physiological needs that drives goal-directed behavior, ensuring that actions are expressed at the appropriate time, intensity, and context. Although the neurobiology of motivation has been extensively studied, the mechanisms by which motivational drives accumulate over time and are translated into sustained behavioral states remain poorly understood. Konrad Lorenz’s hydraulic model proposes that motivation builds gradually like pressure in a reservoir, yet the existence and nature of a corresponding biochemical mechanism remain unclear. In this study, we tested the hypothesis that drive buildup is encoded by the accumulation of prion-like proteins within neurons that function as motivational hubs. We focused on the Drosophila neuropeptide F (NPF) system, which is homologous to the mammalian NPY system, and examined the role of the prion-like RNA-binding protein Orb2, which undergoes state-dependent oligomerization and regulates local protein synthesis at synapses. Using microscale thermophoresis, western blotting, and confocal imaging, we quantified Orb2 monomeric and oligomeric states in vivo under distinct internal drives. The result demonstrate that prolonged starvation, but not acute water deprivation, induced increased Orb2 accumulation and a shift toward oligomerization in NPF neurons, with enrichment at synapses of a subset of neurons projecting to the fan-shaped body and suboesophageal zone. This accumulation was reversible upon refeeding. In contrast, neurons expressing the NPF receptor showed reduced Orb2 accumulation during food deprivation, consistent with reciprocal signaling. Functionally, Orb2 oligomerization was required for starvation-induced synaptic localization of neuropeptide F released from NPF neurons. Together, these findings support a model in which Orb2 acts as a reversible molecular integrator of metabolic state, linking internal energy deficit to sustained neuromodulatory remodeling in feeding-related circuits, and providing a framework for future work on graded drive and downstream RNA targets.

1 The Mina & Everard Goodman Faculty of Life Sciences, The Multidisciplinary Brain Research Center and, Institute
for Nanotechnology and Advanced Materials, Bar-Ilan University, Israel

Nathan Bolen, Colton J. Treadway, Michael Leonardo, Wes Tackett, Aneesh Gupta, Joshua Sisco, Travis B. Salisbury, James Denvir, Sadia Akter, Vini Magalhaes Borges, Alejandro Q. Nato, Jr., Brandon J. Henderson, Price E. Dickson

In 2024, 47,735 people in the United States died from synthetic opioid overdose, primarily due to fentanyl (CDC, 2026). This striking statistic underscores the urgent need to discover and characterize the genetic underpinnings of fentanyl addiction. To this end, we are using genetically diverse mouse strains (N = 40) from the Collaborative Cross (CC) and their founders to discover genetic, transcriptomic, epigenomic, and neurophysiological mechanisms underlying fentanyl addiction-like behaviors and addiction endophenotypes. Notably, the CC founder strains encompass ~90% of known genetic diversity in laboratory mice originating from Mus musculus. We are testing mice on four behavioral assays: intravenous fentanyl self-administration, sign-tracking/goal-tracking, fentanyl locomotor sensitization, and operant sensation seeking. Machine learning enables nuanced analysis of these mouse behaviors. We are collecting nucleus accumbens punches following sign-tracking/goal-tracking and fentanyl locomotor sensitization; gene expression using RNA-seq will be quantified from this tissue. We are using fast-scan cyclic voltammetry, patch-clamp electrophysiology, spatial transcriptomics, single-cell multiomics, and long-read sequencing to characterize CC founder strains, extreme CC strains, or both. Data collection for this study will occur over five years. Here, we report results after ~20 months of phenotyping. Briefly, interim results reveal significant strain effects, GxE effects, and sex effects on many phenotypes. Over the next several years, we will integrate QTL mapping, eQTL mapping, and genetic correlations spanning multiple biological levels. Collectively, these data will provide a foundation for future deep characterization of identified mechanisms, a lasting community resource, and will ultimately contribute to the development of novel, more effective addiction treatments.

Department of Biomedical Sciences,, Joan C. Edwards School of Medicine,, Marshall University,, 1700 3rd Ave.,, Huntington, WV, 25703, USA.

Aijun Zhang1,2, Lukmon Raji2, Zhiping Wu4, Sufiya Khanam1, Dehui Kong1,2, Ling Li2, Junmin Peng4, Bob Moore3, Xusheng Wang1,2, Megan Mulligan2

Cannabis is the most used federally illegal drug in the United States. Over the past years, cannabis usage has increased among adults. Growing research shows that high levels of ∆9-tetrahydrocannabinol (THC), the major psychoactive component of cannabis, impairs memory, learning, attention, cognitive performance, motivation, and is also associated with brain alterations. Accordingly, there is increasing demand to define targets of cannabinoids and to understand how individual genetic differences moderate response to THC. To address this gap, we developed a holistic approach to quantify the effects of acute THC on brain signaling by integrating multi-omics data collected from genetically divergent C57BL/6J and DBA/2J mice. Both sexes (n=4) were injected with 10 mg/kg THC or vehicle (i.p). Cortex was collected 60 mins post-injection. RNA and protein were extracted from each hemisphere, followed by RNA-seq, proteomics, and phosphoproteomics profiling. The results show that the DBA/2J strain exhibits a larger acute response to acute high dose THC than C57BL/6J, with most differences observed at the phosphosite level. Moreover, multi-omics analysis indicates that the cortical phosphoproteome responds robustly to THC at 60 mins, while whole proteome and transcriptome are less responsive. This is consistent with expectations that phosphorylation acts as an early signaling event preceding downstream transcriptional remodeling and changes in protein abundance. Our preclinical study reveals new molecular targets and signaling pathways underlying individual differences in acute responses to THC that may provide insight into genetic differences in adverse health risks in humans.

1. Department of Neurology, University of Tennessee Health Science Center, Memphis, TN 38163 USA 2. Department of Genetics, Genomics and Informatics, University of Tennessee Health Science Center, Memphis, TN 38163 USA 3. Department of Pharmaceutical Sciences, University of Tennessee Health Science Center, Memphis, TN 38163 USA 4. Department of Structural Biology, St. Jude Children's Research Hospital, Memphis, TN 38105 USA

Alanna Mayberry1,2, Eamonn Duffy1,3,4,7, Johnson Ajanaku1,3,4, Laura Saba6,6, Ryan Bachtell2,3,4, Marissa Ehringer1,3,4

Chronic opioid exposure can paradoxically enhance pain sensitivity, expressed behaviorally as opioidinduced mechanical allodynia. This effect is thought to reflect maladaptive nociceptive plasticity, yet the genetic and biological determinants of vulnerability remain incompletely defined. We examined strain- and sex-dependent variation in oxycodone-induced mechanical allodynia, mass and its trajectory, and voluntary opioid intake using a genetically diverse rat panel and applied multivariate analyses to resolve latent dimensions of opioid response. Adult male and female rats (n = 410) from 15 strains of the Hybrid Rat Diversity Panel underwent intravenous oxycodone or saline self-administration. Mechanical withdrawal thresholds (von Frey testing) and body mass were assessed before and after oxycodone exposure. Univariate analyses, principal component analysis, unsupervised k-means clustering, and broad-sense heritability estimation were performed. Oxycodone self-administration produced robust mechanical allodynia and disrupted normal weight gain. Both effects were strongly strain-dependent, whereas sex significantly moderated change in body weight but not allodynia magnitude. Principal component analysis identified four orthogonal dimensions explaining 94% of total variance, dissociating baseline mechanical sensitivity from opioid-induced plasticity and global physiological state. Unsupervised clustering revealed four biologically coherent subtypes distinguished by opioid intake, severity of mechanical hypersensitivity, and body mass. Cluster membership was non-randomly distributed across strain, sex, and treatment condition. Baseline traits demonstrated moderate-to-high heritability, while opioid-induced changes showed reduced heritability, consistent with strong environmental modulation during drug exposure. Oxycodone induces genetically constrained yet heterogeneous neurobehavioral adaptations. Multivariate phenotyping reveals distinct biological routes to opioid vulnerability, underscoring the importance of systems-level approaches in addiction and pain research.

1Department of Integrative Physiology, 2 Department of Psychology and Neuroscience, 3 Institute of Behavioral Genetics, 4 University of Colorado, Boulder, Colorado, USA; 5 Department of Pharmaceutical Sciences, 6 University of Colorado Anschutz, Aurora, Colorado, USA; 7 Northeastern University, Boston, Massachusetts, USA

Carlos Novoa 1, Thomas J. Gould 1 1 Department of Biobehavioral Health

Adolescence is a sensitive developmental window characterized by profound changes in brain circuitry, amplifying both adaptability and vulnerability to environmental influences. Social stress during this period is a prominent risk factor for adverse psychiatric outcomes later in life. This study investigates the interplay between adolescent social stress and genetic background in determining anxiety-related behaviors and nicotine sensitivity. Using male C57BL/6J and C57BL/6NJ mice, we assessed the physiological and behavioral impacts of adolescent social instability stress (SIS). Our results demonstrate that adolescent SIS significantly impairs body weight gain throughout late adolescence. Regarding behavioral indicators, SIS increases spontaneous locomotion and anxiety-like behaviors in the elevated plus maze. Furthermore, physiological assessments revealed that SIS induces the accumulation of hair corticosterone (CORT) while the HPA response to an acute pharmacological challenge with nicotine was attenuated. Adolescent stress altered subsequent nicotine sensitivity, resulting in a reduced recovery from acute nicotine hypolocomotor effects. Finally, genetic background significantly influenced baseline behavior and drug reactivity, with distinct straindependent differences observed in both anxiety and locomotor activity. These findings underscore the critical role of genetic factors in shaping individual vulnerability to mental health challenges and substance use following adolescent stress.

The Pennsylvania State University

LM Raji1, S Saxena1, EA Duecker1, S Khanam1, BM Moore II2, MK Mulligan1

We compared behavioral and cortical transcriptional responses between sexes and genotypes to identify shared and unique pathways that play a role in tolerance to repeated THC (10 mg/kg, i.p.). C57BL/6J and DBA/2J mice of both sexes (n=4) were assigned to THC or vehicle (VEH). Following injection of VEH on day 0, mice received four consecutive daily injections of THC or VEH. THC initially induces antinociception, hypomobility, and hypothermia and these traits were quantified daily to track tolerance. On the 5^th day, cortex was collected and RNA extracted for RNAseq analysis. DSeq2 and GSEA were used to identify differentially expressed genes (DEGs) within groups due to treatment and to identify enriched pathways.

Behavioral trait PCA revealed weaker contributions of treatment-associated PCs over exposure days. More treatment DEGs (p<0.05) were detected in C57BL/6J (1,291) and DBA/2J (786) females relative to C57BL/6J (216) and DBA/2J (517) males. There was little overlap between groups. Comparison of top enriched genes and pathways between groups revealed shared and unique responses. Ribosome and protein translation was upregulated in C57BL/6J and male DBA/2J but downregulated in female DBA/2J. Mitochondrial metabolic processes were upregulated in C57BL/6 and downregulated in female DBA/2J. Cellular components of neurons were downregulated in C57BL/6. ERK, GTPase, glucocorticoid receptor, and TNFA-NFKB signaling was upregulated in females or DBA/2J females and downregulated in males or C57BL/6 males. Vascular processes were downregulated in males. Identification of individual and shared mechanisms underlying drug tolerance is important for treatment efficacy and understanding the progression towards problem use and addiction.

1Department of Genetics, Genomics and Informatics, College of Medicine, University of Tennessee Health Science Center, Memphis, TN, USA, 2Deparment of Pharmaceutical Sciences, College of Pharmacy, University of Tennessee Health Science Center, Memphis, TN, USA

Funding Support: NIH/NIDA R01DA056523

Justin Q. Anderson1,2, R. Dayne Mayfield3, Ethan Hagmann2,4, Bryan E. Jensen2, Marissa Borrego1,2, Antonia M. Savarese1,2, Olga Ponomareva3, Elizabeth Osterndorff-Kahanek3, Emma Erickson3, Anna Warden3, Nihal Salem3, Molly Matty4, John C. Crabbe1,2, Angela Ozburn1,2

Alcohol use disorders (AUDs) are known to be polygenetic diseases with high (~0.8) heritability for risk, suggesting that dysregulation of gene expression preceding alcohol exposure may underlie risk. Moreover, there may exist multiple and distinct patterns of gene dysregulation that lead to similar risk for AUDs, thus necessitating the study of multiple, distinct models. We directionally selected the High Drinking in the Dark-1 (HDID-1; 44 generations) and a separate, replicate line (HDID-2; 37 generations) for high blood alcohol following a period of binge-like alcohol drinking from the same genetically heterogeneous progenitor stock (HS/Npt). We report the successful inbreeding of both lines, each starting from their 26th respective selected generation, and the preservation of the selected phenotype (n = 10/sex/fluid/strain; 280 mice). The inbred HDID (iHDID-1&2) lines therefore represent two independently derived, genetically stable models for binge-like alcohol drinking. The two replicate inbred lines are also found to be as transcriptionally distinct as they are similar, relative to their founders. We present differential expression results from alcohol naive mice from both inbred strains and their founders. RNA TAG-seq data were analyzed from six brain regions important for addiction: nucleus accumbens, central nucleus of the amygdala, ventral tegmental area, bed nucleus of the stria terminalis, medial prefrontal cortex, and basolateral amygdala (n = 20/sex/strain/region; 120 mice, 720 samples). We discuss common and unique genetic signatures that are consistent across all six brain regions and their upstream regulators as the most likely targets for therapeutic intervention.

1Department of Behavioral Neuroscience, Portland Alcohol Research Center, Oregon Health and Science University, Portland, OR 97239, United States,2Portland Veterans Affairs Medical Center, Research and Development Service, Portland, OR 97239, United States, 3The Waggoner Center for Alcohol and Addiction Research, The University of Texas at Austin, Austin, TX 78705, United States, 4University of Portland, Portland, OR 97203, United States

Tori Keefauver^1,2, Nicole L. Horan¹, Lillian Morgan¹, Anisha Saxena¹, Ryan W. Logan^3,4, Gregg E. Homanics², Marianne L. Seney¹, Sean P. Farris², Kyle D. Ketchesin¹

Alcohol misuse is a leading cause of preventable death worldwide. Chronic alcohol is associated with disrupted circadian rhythms, yet molecular mechanisms linking circadian rhythm dysregulation and alcohol consumption are poorly understood. Current FDA-approved treatments for alcohol use disorder (AUD) do not target molecular rhythms or sleep-wake cycles. Mammalian circadian rhythms are regulated by transcription-translation feedback loops that regulate the expression of ‘clock genes’ (e.g. Clock, Per, Arntl encoding for BMAL1). Both human and rodent studies have shown clock gene variants are associated with significant changes in reward-seeking behavior. Evidence suggests astrocytes, non-neuronal brain cells with cell-autonomous rhythms, may regulate both circadian rhythms and reward. In the nucleus accumbens (NAc), a region responsible for modulating alcohol- and reward-related behavior, over 43% of the astrocyte transcriptome is expressed rhythmically. No studies to date have investigated the role of NAc astrocyte rhythmicity in regulating alcohol drinking. We used AAV8-Gfap-Cre to functionally ablate molecular rhythms in NAc astrocytes of BMAL1 floxed mice (n = 6/group/sex). Continuous two-bottle choice (2BC), every other day 2BC, and drinking in the dark assessed alcohol drinking. Sucrose preference, social interaction, and light/dark locomotor boxes assessed reward, social, and locomotor behaviors. BMAL1 functional ablation was not associated with changes in any drinking or behavioral paradigm. Future studies will use operant alcohol self-administration to investigate the role of astrocyte molecular rhythms in motivated alcohol-seeking behavior. Understanding bidirectional relationships between astrocyte molecular rhythms and alcohol consumption will elucidate novel mechanisms of diurnal rhythmicity and inform development of targeted circadian therapeutics for AUD.

¹ Department of Psychiatry, Translational Neuroscience Program, Center for Neuroscience, University of Pittsburgh, Pittsburgh, PA 15261, USA.

² Department of Anesthesiology and Perioperative Medicine, Center for Neuroscience, University of Pittsburgh, Pittsburgh, PA 15261, USA.

³ Department of Psychiatry, University of Massachusetts Chan Medical School, Worcester, Massachusetts

⁴ Department of Neurobiology, University of Massachusetts Chan Medical School, Worcester, Massachusetts

Funding Support: NIAAA R21 (AA031074; KDK, MLS, SPF, GEH, RWL), NIMH K01 (MH128763; KDK), and predoctoral fellowship to TK funded by National Institute of Neurological Disorders and Stroke (NINDS: T32 NS141747).

A Bahi1,2, JL Dreyer3

Anxiety disorders frequently co-occur with alcohol use disorder (AUD), and mesolimbic cholinergic signaling has been implicated in the regulation of affective and reward-related behaviors. The muscarinic M4 receptor (M4R) is highly expressed in striatal circuitry, including the nucleus accumbens (NAc), where it may function as an inhibitory modulator of behavioral output. However, its role within the NAc in regulating anxiety-like and ethanol-related behaviors remains unclear. Adult mice received bilateral intra-accumbal injections of viral vectors expressing shRNA targeting M4R or a control construct. Anxiety-like behavior was assessed using the elevated plus maze (EPM) and open field (OF) tests. Voluntary ethanol intake and preference were subsequently measured using a two-bottle choice paradigm. Endpoint M4 mRNA expression within the NAc was quantified using real-time PCR, and expression levels were correlated with behavioral parameters. M4R loss-of-function in the NAc significantly increased anxiety-like behavior, indicated by reduced open-arm exploration in the EPM and decreased center time in the OF test. Following anxiety assessment, accumbal M4-deficient mice exhibited elevated voluntary ethanol intake and preference compared to controls. Saccharin and quinine consumption were unchanged, suggesting enhanced ethanol drinking was not attributable to altered taste sensitivity. Importantly, the degree of M4R mRNA reduction correlated positively with anxiety-like measures and ethanol intake. These findings demonstrate that M4R in the NAc serve as a critical inhibitory regulator of anxiety-like behavior and ethanol consumption, identifying accumbal M4R signaling as a potential therapeutic target for comorbid anxiety disorders and AUD.

1Department of Basic Medical Sciences, College of Medicine, Ajman University, Ajman, UAE

2Center of Medical & Bio-Allied Health Sciences Research, Ajman University, Ajman, UAE

3Division of Biochemistry, Department of Medicine, University of Fribourg, Fribourg, Switzerland

Lauren May¹, Alexander Khortus², Jasmohan S Bajaj³ and Jennifer T. Wolstenholme¹

Excessive alcohol consumption decreases gut microbiota diversity and is associated with microbial dysbiosis, defects in the intestinal mucosal barrier, and changes in immune responses. In a germ-free mouse model, our prior work has shown that the beneficial effects of a fecal microbiome transplant (FMT) from treated patients can be transmitted into mice to reduce ethanol intake. Importantly, live microbiota associated with butyrate production, but not their germ-free supernatants, were necessary to reduce drinking. These findings suggest that altering the microbiome in favor of short-chain fatty acid-producing bacteria may be a promising avenue for alleviating the symptoms associated with alcoholic liver disease and drinking. The goal of the current study is to determine what characteristics of FMT donors (bacterial diversity, engraftment efficiency) correlate with the greatest reduction in ethanol consumption. The endogenous gut microbiome was depleted in male and female C57BL/6 mice, followed by administration of one of three healthy human FMT samples and ethanol intake and preference was assessed. We found a robust sex difference in response to cross-species FMT. Two of the three donors significantly reduced ethanol intake and preference in female, but not male, mice. Ongoing metagenomic studies are evaluating microbiota chages during the course of engraftment and drinking to determine the specific characteristics of healthy FMT samples which may most contribute to lasting engraftment and reductions in ethanol drinking. These models of immunocompetent mice with cross-species FMT will provide a foundation for mechanistic studies on gut microbiome modulation as a way to influence ethanol consumption.

¹VCU-Alcohol Research Center and Department of Pharmacology and Toxicology, Virginia Commonwealth University, Richmond, VA 23298

²Department of Medicine, University of Minnesota, Minneapolis, MN 55455

³Department of Internal Medicine, Virginia Commonwealth University, Richmond, VA 23298

Funding Support: NIAAA R21 AA031293

David L. Haggerty^1,2, Caleb B. Darden¹, David M. Lovinger¹

Accurate pose estimation underpins quantitative analysis of behavior, yet many deep learning-based tracking tools remain optimized for offline workflows that rely on fragmented software pipelines, workstation-grade GPUs, or external middleware for real-time deployment. Here, we present an integrated software-hardware ecosystem for pose estimation that spans dataset creation, model training, offline analysis, and real-time deployment on embedded edge-computing devices. SqueakPose Studio provides a unified software suite for whole-frame, deep learning-based pose estimation that integrates dataset creation, manual and model-assisted labeling, model training, validation, and large-scale offline inference. For experiments requiring continuous recording and synchronized data acquisition, SqueakView enables real-time model deployment, video capture, and sensor logging on embedded hardware. In parallel, MouseHouse provides a compact, modular enclosure for home cage-based experiments that integrates embedded GPU compute, microcontroller-based timing, and peripheral I/O. A shared data format and deterministic timing architecture ensure consistency between offline analysis and real-time experimentation. Together, SqueakPose Studio, SqueakView, and MouseHouse provide a unified and scalable platform for pose estimation and embedded behavioral experimentation without reliance on workstation-grade hardware or external middleware. This framework enables discovery of previously unobserved behavioral motifs in real time and supports in silico modeling of behavioral sequences relevant to health and disease states.

¹ Laboratory for Integrative Neuroscience, National Institute on Alcohol Abuse and Alcoholism, National Institutes of Health, Bethesda, MD 20892, USA

² PRAT Fellow, National Institute of General Medical Sciences, National Institutes of Health, Bethesda, MD 20892, USA

Funding Support:

NIGMS – NIH: 1FI2GM154674 (DLH)

NIAAA – NIH: 1ZIAAA000407 (DML)

Reza Azanchi¹, Arianna Bouchie¹, Miauaxochitl Haskie¹, Amanda Waterman¹, Mukulika Ray^2,3, Kate O’Connor-Giles¹, Erica Larschan³ and Karla R. Kaun¹

The last 20 years of Drosophila addiction research has revealed remarkable similarities between flies and mammals in alcohol and drug-induced behavior and the neural and molecular mechanisms underlying these behaviors. Drosophila show parallels to mammals in nicotine-induced behavior and the neural and molecular mechanisms underlying this. Low doses of volatilized nicotine induce an acute startle and hyperactivity responses whereas larger doses decrease locomotion in adult Drosophila. Nicotine consumption during development increases heartrate, reduces survival to adulthood, and increases brain size. Like in mammals, nicotine induces dopamine release during both larval and adult stages, and nicotine-dependent changes in locomotion are dopamine dependent. However, there is a gap in our knowledge about whether nicotine is rewarding, and what the genetic and epigenetic mechanisms underlying this response might be. We developed a new conditioned odor preference assay to test the reward responses to nicotine in Drosophila, and found that flies reveal appetitive behavioral dynamics favoring an odor previously paired with doses of nicotine that induce acute locomotor hyperactivity. Furthermore, we conducted multiomic analysis of the Drosophila mushroom body, a structure required for memory formation, and determined complexity in expression of multiple nicotinic acetylcholine receptors, suggesting that nicotine has strong synergistic effects on cue-induced olfactory memory. Together, our data suggest Drosophila is a strong model to test the mechanisms underlying the addictive properties of nicotine.

¹Department of Neuroscience, Brown University;

²Department of Molecular Biology, Cell Biology and Biochemistry, Brown University,

³Department of Biology, University of Massachusetts Boston

Funding support: NIAAA R01AA024434, NIDA R01DA058947, NIAAA R01AA031702, Brown University

JD Wherry¹, MR Ross⁴, DV Gil⁴, GE Homanics^1,2,4, SP Farris^1,3,4

Alcohol Use Disorder (AUD) is a highly prevalent disease that causes deterioration of health and a steep socioeconomic cost. Individuals with AUD often report symptoms of chronic pain and many continue consuming alcohol to combat these symptoms, especially during withdrawal. Alcohol has deleterious effects on the neuroimmune system that cause aberrant inflammation and increased sensitivity to painful stimuli. However, the neurobiological connection between AUD and chronic pain remains ill-defined. Microglia are the primary immune cells in the central nervous system and mediate neuroinflammation associated with alcohol consumption and pain responses. We hypothesized that exposure to chronic intermittent ethanol vapor (CIEV) would lead to heightened spontaneous nociception (i.e. grimacing) and changes in microglial morphology indicative of a reactive phenotype. After five weeks of CIEV, PainFace software showed increased grimacing behavior in mice 24 hours into withdrawal compared to air controls, representative of chronic alcohol withdrawal-induced pain (CAWIP). Grimacing behavior was negatively correlated with territory occupied by microglia in the prefrontal cortex (PFC) indicative of increased microglial reactivity. As a positive control, a separate cohort of mice were injected with lipopolysaccharide (LPS; 1.0 mg/kg) to induce innate immune activation, and grimacing behavior was measured 24 hours post-injection. LPS treatment led to increased grimacing behavior and reductions in territory occupied by microglia compared to saline treatment; similar to mice undergoing withdrawal from CIEV. These data indicate an association between spontaneous nociception and microglial reactivity in mice undergoing withdrawal from chronic ethanol exposure and identify microglia as a possible therapeutic target for treating CAWIP.

¹Department of Anesthesiology & Perioperative Medicine, ²Department of Pharmacology & Chemical Biology, ³Department of Biomedical Informatics, ⁴Center for Neuroscience, University of Pittsburgh, Pittsburgh, PA.

Funding Support: Funding Support: NRSA T32 DA057922, NIAAA U01 AA020889, NIAAA R01 AA030257

AJ Brandner^1,2,3, C Guerra-Solano^1,2,3, AM Kogos^1,2,3, BK Taylor^1,2,3, SP Farris^1,2,3,4

Alcohol Use Disorder (AUD) is characterized as a chronic medical condition known to be comorbid with pain. Mice experience mechanical and spontaneous pain during alcohol withdrawal, known as chronic alcohol withdrawal-induced pain (CAWIP). We now wanted to characterize a central, brain mechanism that underlies alcohol-induced hypersensitivity during alcohol withdrawal. We hypothesized that the parabrachial nucleus (PBN), a hub for pain and aversion, mediated pain-like responses during alcohol withdrawal. B6J mice were exposed to four weeks of chronic intermittent ethanol vapor (CIEV) or Air (16 hrs ON/8 hrs OFF 4 days/week) to induce pain. During withdrawal, mice underwent 1) immediate early gene (Fos) activation or 2) in vivo fiber photometry experiments to measure PBN neuron activity. To induce Fos expression, mice were anesthetized and their left hind paw was stimulated with either a light brush or a noxious pinch. After 90 minutes, brain tissue was taken for histology. For fiber photometry, stimuli, ranging from innocuous to noxious, were applied to the left hind paw of the mice while calcium activity was recorded in real-time. Both PBN Fos expression and PBN calcium activity were increased by non-painful and painful stimulation in mice treated with CIEV when compared to Air controls. Our studies indicate that PBN neurons are more excitable by innocuous (allodynia) and painful (hyperalgesia) stimuli during alcohol withdrawal. Alcohol-induced sensitization of PBN neurons may underlie the hypersensitivity phenotype we see in our laboratory mice during withdrawal from CIEV. Future studies will focus on PBN neuron inhibition to reduce pain.

Affiliations: Center for Neuroscience¹, Pittsburgh Center for Pain Research², Department of Anesthesiology and Perioperative Medicine³, Department of Biomedical Informatics⁴, Pittsburgh, PA, 15261 USA

Funding Support: NIAAA, NIDA, and NINDS grants AA024836 (SPF), AA020889 (SPF), AA030257 (SPF), DA037621 (BKT), NS045954 (BKT), NS112632 (BKT), AA031431 (AJB), and NS073548 (AJB). We also acknowledge support from Bridging Connections in Addiction Research (BCAR) at the University of Pittsburgh and the Pittsburgh Foundation.

André Lucas Silva Borges, Rebeka Sultana, Amanda M Barkley-Levenson

Alcohol use disorders are known to be major health issues that result in a high number of direct and indirect deaths every year. The gene solute carrier 39a8 (Slc39a8), which encodes a transporter of zinc and other metals, has been found in multiple genome-wide association studies (GWAS) to be associated with alcohol consumption and problematic alcohol use. Despite this association, very few studies have characterized the relationship between Slc39a8 and ethanol-related traits. Thus, we employed ethanol conditioned place preference (CPP) and aversion (CPA), and open field tests to further understand the role of this gene in sensitivity to ethanol’s motivational, locomotor, and anxiolytic effects. Adult naïve Slc39a8 heterozygous knockout mice (HET) and their wild-type (WT) littermates of both sexes were used. For both the CPP (injections immediately before conditioning trials) and CPA (injections immediately after conditioning trials) studies, 2 g/kg injections of ethanol were paired with tactile floor cues during conditioning sessions, and floor preference/avoidance was then assessed during drug-free tests. We found no effect of genotype on aversive sensitivity to ethanol. Interestingly, HET mice showed significantly higher ethanol CPP expression than WT mice after two weeks of conditioning. This suggests Slc39a8-deficiency increases ethanol reward sensitivity without altering aversive sensitivity. In ongoing studies in our lab, we are testing the effects of increasing doses of ethanol on locomotor and anxiety-like behavior in an open field apparatus, which will add important insight on how changes in Slc39a8 expression impact sensitivity to other effects of ethanol

University of New Mexico

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Ivanna Ihnatovych¹, Ryu P. Dorn¹, Eduardo Cortes Gomez², Harneet Sandhu¹, David A. Bennett³, Jianmin Wang², Kinga Szigeti¹

Human specific gene CHRFAM7A, a fusion product between ULK4/FAM7A and CHRNA7, is present in 99% of human population in different copy number (CN) and orientation. Both the direct and inverted alleles are transcribed with similar frequency, suggesting they underwent similar selective pressure and that both are functional. The direct allele is translated, and the protein incorporates into a7 nAChR modifying its function; the mechanism of the inverted allele is unclear. As the inverted allele (CHRFAM7A_Δ2bp) is not translated and its expression does not affect CHRNA7, we hypothesized that the CHRFAM7A_Δ2bp mechanism is RNA mediated regulation of ULK4. Human brain RNAseq data, brain tissue, primary macrophages (MΦ), and the iPSC model were used to study ULK4 expression and regulation. Analyzing RNAseq data from ROSMAP, we detected the long and short ULK4 transcripts and demonstrated that the long isoform expression correlates with CHRFAM7A_Δ2bp CN. qPCR with isoform-specific primers further confirmed that with increasing CHRFAM7A_Δ2bp CN, correlation between the isoforms decreases in human brain and MΦ. In the iPSC model, the increased long to short isoform ratio - both at the mRNA and protein level - was detected in neuronal progenitors and MΦ containing CHRFAM7A_Δ2bp compared to CHRFAM7A null cells. In the inverted line, higher ULK4 Long/ ULK4 Short ratio correlated with higher level of α-tubulin acetylation and polymerization. These findings suggest that the CHRFAM7A_Δ2bp exerts genetic epistasis on ULK4 resulting in increased long to short ULK4 isoform ratio. Our iPSC model presents an opportunity to elucidate the molecular mechanism of ULK4 regulation.

¹ Department of Neurology, State University of New York at Buffalo, 875 Ellicott St., Buffalo, NY, 14203, USA, ² Roswell Park Comprehensive Cancer Center, 665 Elm St, Buffalo, NY 14203, USA, ³ Rush Alzheimer’s Disease Center, Rush University Medical Center, Chicago, IL, USA

Funding Support: Community Foundation for Greater Buffalo (Kinga Szigeti). ROSMAP is supported by NIA grants P30AG10161, P30AG72975, R01AG15819, R01AG17917, U01AG46152, and U01AG61356.

Jingjing Yan*¹, Sydney Christensen*^1,2, Kelsey Mainard*¹, Fumihiro Ito¹, Phil Zhang¹, Yangyuan Li¹, Wanhe Li^1,2

Histone monoaminylations represent a novel class of histone post-translational modifications (PTMs), in which monoamine neurotransmitters—such as dopamine, serotonin, and histamine—are covalently attached to the tail region of histone H3 via transamidation reactions. In Drosophila, monoaminergic neurotransmission and neural circuits play key roles in regulating sleep and wakefulness. We therefore hypothesized that this novel form of histone modification might play a direct role in controlling sleep in Drosophila.

Using a set of Drosophila neurogenetics tools, we found that perturbing histone monoaminylation caused nighttime sleep loss, specifically within a defined circadian window, indicating a defect in sleep maintenance. We conducted a large-scale, unbiased, circuit-based screen and identified the cell types that supported this histone monoaminylation-dependent sleep phenotype. Unexpectedly, we discovered that the inhibitory neurotransmitter γ-aminobutyric acid (GABA), which also contains a primary amine group, could similarly modify histone H3 via a transamidation reaction. This novel histone mark, termed histone H3-carboxypropylaminylation, along with other histone monoaminylation marks, exhibited circadian features and regulated sleep/wake behavior in a time-of-day-dependent manner. We employed a comprehensive set of biochemistry, genetic, and genomic approaches to further characterize the gene regulatory network underlying histone monoaminylation-dependent sleep regulation. Because monoamine biochemistry and histone proteins are remarkably conserved between humans and flies, this work may reveal epigenetic mechanisms of sleep regulation that are evolutionarily conserved.

*authors contributed equally.

1.Center for Biological Clocks Research, Department of Biology, Texas A&M University, College Station, TX 77843

2.Interdisciplinary Graduate Program in Genetics and Genomics, Texas A&M University, College Station, TX 77843

Funding Support: This work was supported by the Cancer Prevention and Research Institute of Texas (RR220021 to W.L.) and the National Institute of General Medical Sciences (GM150832 to W.L.).

K. T. Reyes¹, N.K. Amissah¹, S. Sawant², E. Olusanya¹, P.J. Meyer¹

Genome-wide association studies have identified teneurin-4 (Tenm4) as a potential candidate gene underlying individual variation in drug-related traits, including cue-reactivity. Teneurin-4 produces the peptide Teneurin C-terminal Associated Peptide 4 (TCAP-4), which we hypothesized would alter reward cue-reactivity as tested by Pavlovian conditioned approach (PavCA). Based on prior evidence that another TCAP (TCAP-1) crosses the blood-brain barrier, we chose intravenous administration instead of ICV. Fifty-six female Sprague Dawley rats were implanted with intravenous jugular catheters to administer TCAP-4. Rats were randomly assigned to one of four groups: 0 pmol (n=14), 300 pmol (n=14), 1000 pmol (n=14) or 3000 pmol (n=14). Rats underwent eight daily PavCA sessions. Rats received IV TCAP-4 (300, 1000, or 3000 pmol) or vehicle (0 pmol) 30 minutes before each of five PavCA sessions, followed by three sessions in which TCAP-4 was not given. The main measures in PavCA testing included lever contacts, food cup entries, lever and food cup latency, and PavCA index. Following the last PavCA session, two days of 40-minute conditioned reinforcement tests were conducted to evaluate how well the lever reinforces an instrumental response. TCAP-4 did not produce the expected reduction in cue approach (sign-tracking), although food cup entries (goal-tracking) increased significantly (p < 0.05), with the largest increases in early sessions. This suggests a shift away from lever-directed incentive salience rather than a global reduction in learning. The conditioned reinforcement test suggested TCAP-4 reduced reinforcing value of the cue. These results identify Tenm4 as a potential regulator of individual differences in cue-guided motivation.

¹Department of Psychology, University at Buffalo, Buffalo NY, ²UC James L. Winkle College of Pharmacy, Cincinnati, Ohio

Funding Support: DA060669

EB Palmieri 1, B Bryd 1, L Sutton 1

Alcohol use disorder (AUD) affects 1 in 10 people in the United States alone, causing significantly reduced quality of life in affected individuals. Despite the prevalence and social, economic, and medical burdens associated with AUD, understanding of genetic contributors remains limited. One of the major molecular mechanisms underlying alcohol seeking and reward is mammalian target of rapamycin complex 1 (mTORC1) signaling. mTORC1 is the master cell growth regulator; it is ubiquitously expressed, and it integrates nutrient signals in order to direct cell growth through a general translation mechanism. In the brain mTORC1 is involved in neuronal growth, in part directing formation of learning and memory. Through this process, mTORC1 is thought to encode the rewarding effects of addictive stimuli. In fact, research has found that abnormal mTORC1 activity alters drug seeking, including ethanol seeking. However, the exact mechanism through which mTORC1 affects drug seeking is poorly understood. Here, we investigate the role of a novel modulator of mTORC1 signaling with specific neuronal expression, GPR155, on ethanol-related behavior. We investigated recovery from ethanol-induced sedation as well as ethanol drinking paradigms in a GPR155 knockout mouse model. While we find that motor recovery from ethanol-induced sedation and blood ethanol clearance are unaffected by GPR155 knockout status, ethanol preference is decreased in socially-housed female GPR155 knockout mice. Findings are not only the first to identify the behavioral role of GPR155, but they are also the first to identify its role in ethanol drinking behaviors, identifying it as a novel target of alcohol research.

1Biology Department University of Maryland Baltimore County, Baltimore, MD, USA

Doris I. Olekanma ¹, Sonia A. Cavigelli ^1,2, Helen M. Kamens ^1,2
1 Huck Institute of Life Sciences, Neuroscience, Pennsylvania State University
² Department of Biobehavioral Health, Pennsylvania State University

Opioid Use Disorder remains an unrelenting public health concern in the United States, with hundreds of overdose deaths occurring daily. Clinical data suggest that women may be more susceptible to the addictive properties of opioids than men. Although adolescent stress is a risk factor for later use, it remains unclear whether biological sex moderates vulnerability to stress-induced increases in opioid use. To address this gap, we investigated how sex and genetic background influence the effects of adolescent stress on opioid intake. Male and female C57BL/6J and BALB/cJ mice were exposed to repeated restraint stress (2 h/day for 14 days; PND 37 – 50) or control conditions. Body weight was measured daily, and blood corticosterone was obtained at the end of the final restraint session. In adulthood (PND 60), morphine consumption was examined using a two-bottle choice protocol. Repeated daily restraint in adolescence was an effective stressor, as evidenced by delayed body weight gain and elevated corticosterone levels, even after 14 exposures, in both strains and sexes. Adolescent stress selectively increased morphine consumption in female C57BL/6J mice compared to controls, with no significant effects observed in male C57BL/6J or BALB/cJ mice. Our findings highlight how adolescent stress can alter future intake of opioids in a strain and sex specific manner, supporting gene X environment contributions to Opioid Use Disorder.

Hariom Sharma1, Noel McGrory1, Giovannina Kirby1, Harold A Burgess1

Abstract: Disruptions in sensory gating contribute to several neuropsychiatric disorders, including schizophrenia, yet the neural mechanisms governing short-interval prepulse inhibition (PPI) remain incompletely understood. PPI is a conserved sensorimotor gating mechanism that relies on precise temporal processing of sensory inputs. Here, we identify a hindbrain circuit that selectively regulates PPI at short inter-stimulus intervals (ISIs) using the Gal4 driver lines y242 and y467. Both lines share overlapping expression in rhombomere 3, revealing this region as a critical locus for short-ISI PPI modulation. Functional analyses demonstrate that neurons within rhombomere 3 specifically regulate PPI at brief ISIs without affecting long-ISI PPI. Within this population, a medially located subpopulation of GABAergic neurons was identified, implicating inhibitory control as a key mechanism. Anatomical tracing shows that rhombomere 3 neurons project directly to the Mauthner (M) cell, a command neuron essential for startle responses. Consistently, the M cell expresses GABA receptor 5, and disruption of this receptor impairs PPI. Together, these findings define a rhombomere 3, Mauthner cell inhibitory pathway that gates sensorimotor responses at short ISIs, providing novel insight into the neural substrates of temporal processing and potential mechanisms underlying sensory gating deficits in neuropsychiatric disorders.

1National Institute of Child Health and Human Development, National Institute of Health, Bethesda, USA

In humans, neuroligin 3 (NLGN3) is a gene associated with autism, yet its role in behaviour and lifespan remains unclear. Loss of function of nlg3, the Drosophila melanogaster ortholog of human NLGN3, has been shown to disrupt social behaviours, suggesting its involvement in circuits underlying social interaction. To test which genetic pathways were influenced by nlg3, we analyzed RNA-seq data from Drosophila nlg3 loss of function mutants. We found that genes linked to longevity were downregulated, with gene ontology analysis indicating enrichment in aging-related pathways, including key regulators such as Drosophila FOXO (dFOXO) and heat shock proteins HSPs (stress-responsive chaperones linked to proteostasis). We manipulated nlg3 expression using deficiency lines, downregulation, and overexpression using the Gal4-UAS system. RT-qPCR revealed that overexpression of nlg3 significantly suppressed the expression of genes encoding HSP68 and HSP70; while the expression of dFOXO remained unchanged. In contrast, nlg3 reduction showed downregulation of the genes encoding dFOXO, HSP68, and HSP70. Behavioural assays showed that nlg3 overexpression increased social spacing and reduced lifespan, while partial loss of nlg3 in heterozygous deficiency flies extended lifespan without affecting social behaviour. Furthermore, RNAi-mediated knockdown led to an even stronger lifespan extension, but in that case, with increased social spacing. This supports a dosage-sensitive role for nlg3 in regulating social behaviour and lifespan in Drosophila, with implications for aging and neurodevelopmental disorders in more complex organisms.

1 Department of Biology, Faculty of Science, University of Western Ontario, London

Smelling diacetyl, a microbial‐fermentation byproduct, is a key food cue for Caenorhabditis elegans. Chemotaxis assays serve as a traditional model for olfactory behavior. However, the N2 reference strain may not reflect species-wide sensitivity. We first sought to establish straightforward, replicable conditions to enhance the reproducibility of the volatile chemotaxis response, so that we could use the assay to compare diacetyl responses across wild strains. After systematically optimizing hermaphrodite age, lay duration, brood size, and incubation temperature, we found that four gravid day-1 adults and a 16-hour lay at 20 °C produced abundant, synchronized progeny and highly reproducible chemotaxis responses. We established a robust dose–response curve for diacetyl and identified 3 × 10⁻⁴ as the dilution that elicited an N2 chemotaxis index (CI) of ~0.6 so that both diminished and elevated CIs relative to N2 could be observed. Applying this protocol to 48 wild strains (the 12 divergent + 36 mapping strains) together with N2 showed that across the 49-strain panel, CIs ranged from approximately 0.20–0.85. Several strains exceeded N2 while others underperformed, placing N2 in the upper-middle rather than at the maximal end of the species’ olfactory spectrum. In sum, our streamlined conditions enable sensitive, comparable measurements across strains and set the stage for genome-wide association (GWA) studies to explore how natural genetic variation between wild strains contributes to differences in olfactory behavior.

¹Department of Biological Sciences, University at Buffalo, The State University of New York, Buffalo, NY, USA

Funding Support: This work was supported by the National Science Foundation (IOS-2334104 to DMF 3 and DEB-2049947 to OG), the National Institutes of Health (R35GM156519 to OG), and 4 a Human Frontiers Science Foundation grant (RGP0001/2019) to ECA.

Maulik Masaliya1 , Dielin Wang2 , Yashita Arora3 , Shuai Wang4 , Giulia Demo5 , Jamal B. Williams6

Background. ADHD is highly heritable, but case-control GWAS explain only a small fraction of that heritability. Genetic variants interacting with continuous behavioral traits may account for some missing heritability. We tested this using interaction-based GWAS across six ADHD phenotypes and structural MRI in a pediatric cohort. Methods. We used the ABCD Study (n up to 8,302 children, ages 9–11), stratified into five ancestry groups. Six phenotypes: hyperactivity, inattention, impulsivity, emotional dysregulation, CBCL Attention Problems T-scores, and Flanker inhibitory control. SNP × behavior interactions were modeled on 34 sMRI measures from the Desikan–Killiany atlas in PLINK and meta-analyzed across ancestries with METAL. Significant loci (p < 5×10−8 ) collapsed into 1 Mb windows. Results. We identified 589 genome-wide significant interaction loci — substantially exceeding prior ADHD GWAS of comparable sample size. rs56817477 (Chr 2) reached Z = −7.76 (p = 8.7×10−15) for 3rd ventricle volume independently in hyperactivity, emotional dysregulation, and inattention, with identical direction across all three analyses — within-study SNP-level replication not previously reported for a structural brain locus in ADHD. Precentral gyrus volume was significant across all five symptom composites, identifying primary motor cortex as a transdiagnostic neuroanatomical hub. Impulsivity yielded 18 independent white-matter hyperintensity loci across 13 chromosomes. Sixty-three ventricular clusters showed convergence across three or more phenotypes. Interaction effect directions reversed across phenotypes, revealing phenotype-specific genetic architectures. Conclusions. SNP-by-behavior interactions expose ADHD neurobiology invisible to case-control designs. Phenotype-specific pathways converge on ventricular, sensorimotor, and fronto-parietal systems. The chromosome 2 locus is the most robustly replicated structural brain locus in pediatric ADHD genetics to date.

Keywords: ADHD · interaction GWAS · structural MRI · SNP×behavior · Desikan–Killiany · ABCD Study · ventricular volume · precentral gyrus · white matter

1 Jacobs School of Medicine and Biomedical Sciences, State University of New York at Buffalo, Buffalo, NY 2Department of Psychiatry, Jacobs School of Medicine and Biomedical Sciences, State University of New York at Buffalo, Buffalo, NY 3Department of Pediatrics, University at Buffalo, Buffalo, NY 4Department of Psychiatry, Jacobs School of Medicine and Biomedical Sciences, State University of New York at Buffalo, Buffalo, NY 5Department of Psychiatry, Jacobs School of Medicine and Biomedical Sciences, State University of New York at Buffalo, Buffalo, NY 6Department of Psychiatry, Jacobs School of Medicine and Biomedical Sciences, State University of New York at Buffalo, Buffalo, NY