Rachel C. Rice, Remy R. Frawley, Daniela V. Gil, Annalisa M. Baratta, Shirley Y. Hill, Gregg E. Homanics, Sean P. Farris

Alcohol Use Disorder (AUD) is a serious neuropsychiatric disorder caused by genetic and environmental factors impacting widespread changes in molecular pathways that may be inherited across generations. Our current study investigated unbiased system-wide changes in gene expression in the blood and brain (i.e., medial prefrontal cortex and central nucleus of amygdala) of alcohol naïve male and female F1 C57BL/6J offspring from paternal preconception ethanol exposure (PPE), maternal preconception ethanol exposure (MPE), and biparental preconception alcohol exposure (BPE). In parallel, we conducted an unbiased RNA-Seq analysis of human blood lymphocytes from a 30+ year longitudinal multigenerational study of families with and without a history of AUD to identify evolutionary conserved changes in gene expression related to disease. Differential gene expression analyses revealed sex- and alcohol-dependent changes in gene expression that were evolutionary conserved across species. Only approximately 40% of differentially expressed genes were protein-coding, suggesting a potential epigenetic role for non-coding RNAs in the risk of developing AUD. A systematic meta-analysis identified shared enriched biological pathways for protein-coding genes across PPE, MPE, and BPE groups related to proinflammatory-, metabolic-, growth-, and cancer-related processes, with differing directionalities pathways depending on sex of the offspring. Overall, this work encompasses the first comprehensive cross-generational transcriptomic study on preconception ethanol exposure and may identify potential biomarkers of cross-generational alcohol-related disease risk. We gratefully acknowledge the support of NIH/NIAAA grants AA020889, AA030257, and AA031168 as well as internal support from Bridging Connections in Addiction Research at the University of Pittsburgh.

University Of Pittsburgh Center For Neuroscience, Pittsburgh, Pa 15261; University Of Pittsburgh Department Of Anesthesiogy And Perioperative Medicince, Pittsburgh, Pa 152161; University Of Pittsburgh Department Of Psychiatry, Pittsburgh, Pa 15261; University Of Pittsburgh Department Of Biomedical Informatics, Pittsburgh, Pa 15261

A Takahashi1, N Mimura1, B Hu1, K Okamoto1, K Ohashi2, T Kawase2, A Toyoda3, T Tsukahara2, K Mitsui1

There are a large individual differences in aggression within a species, influenced by genetic and environmental factors, including gut microbiota. In this study, we found significant differences in aggression between ICR mouse strains obtained from CLEA Japan (ICR^CLEA) and Charles River Laboratories Japan (ICR^CR; currently the Jackson Laboratory Japan). In a 3-day resident-intruder test, 90% of ICR^CR males showed aggressive behavior, compared to only 37% of ICR^CLEA males. Analysis of fecal samples using 16S rRNA sequencing showed distinct differences in gut microbiota composition between breeders. Fecal microbiota transplantation (FMT) between ICR^CLEA and ICR^CR males demonstrated bidirectional changes in aggression: FMT from ICR^CLEA donors to ICR^CR males significantly reduced aggressive behavior, whereas FMT from ICR^CR donor to ICR^CLEA males increased aggression. These results indicate that the gut microbiota composition influences aggressive behavior. Further, 16S rRNA sequencing of fecal samples identified five bacterial genera significantly enriched in high-aggression groups related to ICR^CR and one bacterial genus characteristic of low-aggression groups related to ICR^CLEA. When low-aggressive ICR^CLEA was subjected to post-weaning social isolation stress, which is known to escalate aggressive behavior of male rodents, the bacterial genus characteristic of ICR^CLEA–associated low aggression showed significant reduction in the socially isolation group. These results suggest that gut microbiome associated with aggression are sensitive to social isolation stress.

1 Laboratory of Behavioral Neurobiology, University of Tsukuba, Tsukuba, Ibaraki, Japan, 2 Kyoto Institute of Nutrition & Patholohy, Tsuzuki, Kyoto, Japan, 3 College of Agriculture, Ibaraki University, Ami, Ibaraki, Japan

Funding Support: JST FOREST Program JPMJFR214A, JSPS KAKENHI 22K19744, KAKENHI 22H02660, Astellas Foundation for Research on Metabolic Disorders, and Project for University-Industry Cooperation Strengthening in Tsukuba (Ibaraki Prefecture), JAPAN

Tatsuhiko Goto¹, Prudence Nyirimana¹, Riku Sasaki¹, Dipson Gyawali¹, Atsuhi J. Nagano², Akira Ishikawa³

Animal tameness is one of key behavioral characteristics in domestic animals. Several experimental models including foxes, rats, and mice have been investigated in behavior genetics of tameness. In the livestock industry, tame behavior is desirable for animal production. Given that the genetic basis of tame behavior is revealed using Japanese indigenous chicken resources, tamed chickens will be created efficiently. In this study, we aim to find quantitative trait loci (QTLs) for tame behavior in chickens. From 491 segregating individuals based on a cross between Darumachabo and Tosa-jidori breeds, phenotypes and genotypes were collected. Tame behaviors (number of “steps”, “avoiding”, and “heading”) at 30 weeks of age were evaluated by a 1-min handling test. RAD-seq was used to collect genotypic data. Fully informative 425 SNPs on Chrs. 1-27 were used for QTL analysis of tame behavior using R/qtl. Genome-wide thresholds were calculated by permutation tests. This study revealed two significant QTLs on Chr. 2 (LOD = 4.2) and Chr. 5 (LOD = 3.8) in activity (“step”) during handling test. In addition, two suggestive QTLs were found for “heading” on Chr. 2 (LOD = 3.4) and “avoiding” on Chr. 3 (LOD = 3.3). In these four QTLs, homozygotes for the Tosa-jidori allele indicated high avoidance activity. These QTLs explained a small part (around 8%) of phenotypic variance in tame behavior. Therefore, further genetic mapping is needed for a better understanding of the genetic basis underlying animal tameness.

¹Obihiro University of Agriculture and Veterinary Medicine, Obihiro, Japan, ²Ryukoku University, Otsu, Japan, ³Nagoya University, Nagoya, Japan