Background: Acute functional tolerance (AFT) develops shortly after ethanol administration, and is determined as the change in brain or blood ethanol concentration (BEC) measured at 2 behavioral or physiological endpoints. Acute functional tolerance studies in some rodent strains support a long-held hypothesis that more sensitive strains develop more within-session tolerance. We used the new, 74-strain L x S recombinant inbred (RI) panel, developed from inbred long-sleep (ILS) and inbred short-sleep (ISS) strains, to revisit this hypothesis and to map quantitative trait loci (QTLs) for AFT. We report replication of QTL regions reported by earlier studies of AFT and preliminary application of a coarse single nucleotide polymorphism map analysis to limit QTL intervals for subsequent candidate gene hypotheses.
Methods: Acute functional tolerance was assayed using a test of ataxia: loss and regain of balance on a stationary wooden dowel. Following an initial dose of 1.75 g/kg, BEC was measured at initial loss (BEC(0)) and regain of balance (BEC(1)). A second injection (2.0 g/kg) was administered and blood taken at the second regain of balance (BEC(2)). Acute functional tolerance was calculated as a difference score in 2 ways: (1) between BEC at the 2 successive regains of balance (AFT(1)), or (2) as the difference in BEC at final regain and at initial loss of balance (AFT(2)). We mapped QTLs for BEC(0), a measure of initial sensitivity, and both AFT scores.
Results: All 4 parental strains (LS, SS, ILS, and ISS) developed tolerance, replicating previous published reports. There were significant sex effects for 3 of these strains. The L x S panel showed a 128-fold range in tolerance, with a few strains showing negative tolerance (sensitization). The ISS surpassed the next highest RI strain by 55% and was more than 4 times greater than SS. Heritability estimates for both AFT measures were close to 0.25 for both sexes. One significant QTL accounting for approximately 18% of phenotypic variance (V(P)), on chromosome 12 (AFT(1)), and 1 suggestive QTL (16% V(P)), on chromosome 16 (AFT(2)), were identified. These QTLs replicated regions reported in other studies. A multiple QTL model incorporating the effects of all significant interacting QTLs was developed, explaining almost 60% of V(P). The chromosome 12 region was further investigated by haplotype analysis, which identified many nonpolymorphic regions within the confidence interval, and possible candidate genes in the polymorphic regions.
Conclusions: Both SS and ISS developed greater AFT, assessed by both methods, than LS and ILS; this difference was significant in virtually all sex by strain comparisons. In the L x S RI, there was no correlation between initial sensitivity, measured by BEC at initial loss of balance, and either measure of AFT, on a stationary dowel. These results indicate that in this model system, initial sensitivity does not predict tolerance. Several QTLs for tolerance were identified; candidates in the narrowed chromosome 12 region, which has been reported in 2 other mapping studies, merit additional study.