Work completed during the previous project period has isolated several quantitative trait loci (QTLs) for traits that index positive and negative motivation effects of alcohol such as preference drinking and conditioned taste aversion (CTA). Because these and several other drug-related traits have been provisionally linked to chromosomes (Chr) 1 and 9, this PARC component will focus on identifying ethanol-relevant genes on those two chromosomes. The main goal is finer mapping of traits on Chr 1 and 9 toward the ultimate end-point of gene identification. We will also examine the interplay among QTLs and other loci (modifiers) that influence or control QTL expression by conducting a genome-wide search for epistastically interacting loci. In S.A.1, we will narrow the Chr1 and 9 QTL regions by testing existing Chr 1 and 9 congenic strains for EtOH phenotypes mapped to these sites. In S.A.2., we propose to develop interval specific congenic strains (ISCS) to further narrow the region(s) to further narrow the region(s) of focus for each phenotype. In S.A.3, we will develop small donor segment (SDS) congenics, and pursue gene identification by sequencing, beginning with candidate genes for EtOH traits associated with regions of approximately 1 cM. In S.A.4, we will carry out a genome-wide search for modifier loci (genes) of known QTLs (epistasis). A secondary goal of this component is to evaluate the dopamine D2 receptor gene (Drd2) and the serotonin 1B receptor gene (Htr1b) as candidate genes for traits mapped in their vicinity. The Chr9 voluntary ethanol consumption QTL residues near both of these genes. We have found that dopamine D2 receptor knockout mice consume less EtOH than do their wildtype (WT) littermates (MS#6-4). Although the important finding that 5-HT/1B gene knockout mice consumed more EtOH than did wildtype controls has not been replicated in more recent studies, we have not discounted Htr1b as a candidate for the Chr9 QTL. One possible explanation for this change is that a modifier gene in the current population is masking the differential phenotype. In S.A.5, we will assess D2 and 5-HT/1B receptor distribution/density and gene expression in specific brain areas of D2 and B6 mice given free choice of EtOH vs. water. In S.A.6, we will test 5-HT/1B knockouts and WT mice on a C57/BL/6J background and D2 knockouts and WT on a 129/J background for voluntary EtOH consumption. In S.A.78, we will backcross the current mixed 129/SvPas x 129/SvEvTac background 5- HT1B mice to the original 129/SvTer x 129/SvPas strain and test offspring for voluntary EtOH consumption. We predict that relatively enhanced consumption will be restored in the backcrossed mutant if a modifier from 129/SvEvTac is currently masking the enhanced drinking phenotype. In addition, our epistasis search could provide of where a modifier of Htr1b effects be located.
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