QTL (quantitative trait loci) are chromosome sites containing alleles (genes) that influence a continuously distributed (quantitative) trait. Recently developed molecular and statistical methods utilizing PCR-based and RFLP marker loci now make it possible to detect and genetically map several QTL determining the intensity of withdrawal from ethanol. Using this approach, we have very recently gathered evidence strongly suggesting that there is a QTL affecting acute ethanol withdrawal intensity in the Pmv-7/D2Mit9 region of chromosome 2. An F2 cross between C57BL/6J and DBA/2J mice was used since these two parental strains differ markedly in ethanol withdrawal intensity after the same ethanol exposure. We propose to identify and map several additional QTL, each accounting for 20% or more of the genetic variance for both acute and chronic ethanol withdrawal intensity. Selective breeding from the QTL will be used to isolate the high and low predisposing alleles. From these, the most promising will be developed into congenic inbred strains through repeated backcrossing with one of the parental inbred strains. These strains will possess a small chromosome segment (1-2% of the genome) containing a known QTL from the C57BL/6J strain superimposed on a genetic background that is 98.99% from the DBA/2 strain, and vice-versa. Because of the near elimination of genetic """"""""noise"""""""" at irrelevant loci, these lines will be valuable in the identification of the precise gene reflected in a QTL (candidate gene), and its neurochemical mechanisms of gene expression, and in determining the influence of each QTL on withdrawal hyperexcitability associated with ethanol, pentobarbital, phenobarbital, diazepam, nitrous oxide and morphine. The selectively-bred Withdrawal Seizure Prone (WSP) and Resistant (WSR) lines will also be screened for the same QTL to further establish their influence in another genetic model. Given the high degree of genetic homology (conservation) between mice and humans, some of the mouse QTL for ethanol withdrawal may have direct counterparts in the human genome. The Alcohol Research Center at the University of Colorado has been funded to selectively-breed new lines for high and low ethanol withdrawal severity using the same methods and foundation population as the WSR/WSP project. We now have the opportunity to study the divergence of the new oppositely- selected lines in terms of changes in gene frequencies for markers closely linked to QTL identified in this project. In addition, the withdrawal intensities associated with ethanol, pentobarbital, phenobarbital, diazepam and morphine will be monitored at 3-generation intervals to determine if some of them diverge in parallel with ethanol withdrawal. If so, this would provide strong evidence that there are extensive common genetic influences between ethanol and the other dependence-producing drugs.
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