The R100Q mutation of the GABA(A) alpha(6) receptor subunit may contributeto voluntary aversion to ethanol in the sNP rat line

We have investigated the GABA, a, subunit molecular composition in two rat lines selectively bred for high or low ethanol preference and consumption, namely Sardinian alcohol-preferring (sP) and Sardinian non-alcohol-preferri ng (sNP) rats, which have been bred at the University of Cagliari, Italy, s ince 1981. A total of 27 sP, 22 sNP and 25 control rats belonging to five o ther different strains, were studied by direct sequencing and amplification refractory mutation system analysis. Among the sNPs, only one was found to be normal, 11 heterozygotes, and 10 homozygotes for the G-->A substitution in codon 100, the same R100Q point mutation previously described in Alcoho l Non Tolerant rats, while no other animal showed any mutated allele. Pharm acological studies have extensively demonstrated that this substitution in the mature peptide changes the benzodiazepine-insensitive receptor to a sen sitive one. In order to test the functional significance of this mutation i n native cerebellar GABA, receptors, selective breeding from Q/R rats was e mployed to obtain a sufficient number of R/R homozygotes. Xenopus laevis oo cytes were then injected with cerebellar synaptosomes extracted from Q/Q, R /Q and R/R sNP rats. Consistently, utilizing the two-electrode voltage-clam p technique, GABA-evoked currents mediated by GABA, receptors containing th e mutated a, subunit were potentiated by diazepam with about a two-fold inc reased potency, as compared to receptors containing the wild-type, benzodia zepine-insensitive a, subunit. Our data show for the first time that a muta ted GABA, a, receptor subunit segregates in a rat line which voluntarily av oids alcohol consumption, and further support a possible involvement of the GABA, receptor containing a mutated or, subunit in the genetic predisposit ion to alcohol preference. (C) 2001 Elsevier Science B.V. All rights reserv ed.