Background: This study aims to identify genetic contributions to voluntary ethanol consumption variability in mice. Intermittent Ethanol Access (alternating periods of free access to ethanol, and forced abstinence) has been shown to produce progressively increasing ethanol consumption in mice, and models the repeated, intermittent, and increasing alcohol consumption that often leads to dependence in humans. In order to identify Quantitative Trait Loci (QTL) at higher mapping resolution, allelic variation, and trait variability than previous studies, we used Diversity Outbred (DO) mice for this study.
Methods: DO mice (n=539, post-QC) were exposed to 3 24hr periods of three-bottle choice (15% and 30% ethanol, and water) per week, for 4 weeks, without any prior ethanol exposure. Genotyping was performed on gigaMUGA arrays, and imputation and analysis were performed using the qtl2 R package. We conducted QTL analysis for first and last week mean 30% ethanol choice (mL30%EtOH/mLTotalEtOH) and total ethanol consumption (gEtOH/kgmouse) and preference (mLEtOH/mLTotalFluid).
Results: On average, mice consumed 6.07g/kg and 7.66g/kg ethanol, had preference levels of 23.83% and 27.48%, and 30% EtOH choice levels of 4.27% and 4.09%, during the first and last weeks of ethanol access, respectively. Paired t-tests revealed significant differences between first- and last-week phenotypes (t=6.34, p=4.631*10-10 for consumption; t=4.12, p=4.400*10-5 for preference; t=-2.01, p=0.045 for 30% EtOH choice). QTL analysis revealed 3 genome-wide significant loci, and several suggestive loci (LOD>6). The significant locus for last-week mean ethanol consumption (LOD=8.22) was found at 8.41Mb Chr4, with a 95% Bayesian support interval of 1.05Mb. This region contained genes Car8, Rab2a, Chd7, and Clvs1, all of which are expressed most highly in mouse cerebellum. The significant locus for first week mean total ethanol preference (LOD=7.52) was found at 79.35Mb on Chr12, with a support interval of 1.82Mb. This region contains several genes, with the peak signal lying just upstream of Rad51b, which is expressed most highly in mouse central nervous system and liver and is involved in DNA repair. Finally, the significant locus for last week mean 30% EtOH choice (LOD=8.63) was found at 108.23Mb on Chr3, with a support interval of 3.77Mb, across a gene-rich region. Support intervals for significant and suggestive signals did not overlap between first- and last-week phenotypes. Ongoing analyses involve narrowing candidate gene lists within the support intervals of significant regions, by finding loci with genotypic patterns across founders that follow the pattern of founder haplotype effects at each QTL.
Conclusion: This study identified several novel genetic loci, with unprecedentedly narrow support intervals, for voluntary ethanol consumption and preference, and choice for higher-concentration ethanol, by examining a genetically and phenotypically highly diverse mouse population. Results suggest that different genetic mechanisms drive these behaviors during the initial period of ethanol exposure versus post-long-term exposure, also evidenced by the mean differences between first- and last-week phenotypes. Preliminary assessment of candidate genes indicate that the liver and CNS may both be involved during ethanol access initiation, whereas specifically the cerebellum may be more involved after long-term consumption.
Animal models , Substance use: Alcohol, Nicotine, Drugs
