The cerebellar GABAAR α6-R100Q polymorphism alters ligand binding in outbred Sprague–Dawley rats in a similar manner as in selectively bred AT and ANT rats

Abstract The alcohol-tolerant AT and alcohol-nontolerant ANT rat lines have been selectively bred for innate sensitivity to ethanol-induced motor impairment. The cerebellar GABA A receptor (GABA A R) α6 subunit alleles α6-100R and α6-100Q are segregated in the AT and ANT rats, respectively. This α6 polymorphism might explain various differences in pharmacological properties and density of GABA A Rs between the rat lines. In the present study, we have used nonselected outbred Sprague–Dawley rats homozygous for the α6-100RR (RR) and α6-100QQ (QQ) genotypes to show that these RR and QQ rats display similar differences between genotypes as AT and ANT rat lines. The genotypes differed in their affinity for [ 3 H]Ro 15-4513 and classic benzodiazepines (BZs) to cerebellar “diazepam-insensitive” (DZ-IS) binding sites, in density of cerebellar [ 3 H]muscimol binding and in the antagonizing effect of furosemide on GABA-induced inhibition of [ 3 H]EBOB binding. The results suggest the involvement of α6-R100Q polymorphism in these line differences and in the differences previously found between AT and ANT rats. In addition, the α6-R100Q polymorphism induces striking differences in [ 3 H]Ro 15-4513 binding kinetics to recombinant α6β3γ2s receptors and cerebellar DZ-IS sites. Association of [ 3 H]Ro 15-4513 binding was ∼10-fold faster and dissociation was ∼3–4-fold faster in DZ-IS α6βγ2 receptors containing the α6-100Q allele, with a resulting change of ∼2.5-fold in equilibrium dissociation constant ( K D ). The results indicate that in addition to the central role of the homologous α6-100R/Q (α1-101H) residue in BZ binding and efficacy, this critical BZ binding site residue has a major impact on BZ binding kinetics.