Decreased agonist sensitivity of human GABA A receptors by an amino acid variant, isoleucine to valine, in the α1 subunit

Abstract

Recombinant human GABA A receptors were investigated in vitro by coexpression of cDNAs coding for α 1, β 2, and γ 2 subunits in the baculovirus/Sf-9 insect cell system. We report that a single amino acid exchange (isoleucine 121 to valine 121) in the N-terminal, extracellular part of the α 1 subunit induces a marked decrease in agonist GABA A receptor ligand sensitivity. The potency of muscimol and GABA to inhibit the binding of the GABA A receptor antagonist [ 3H]SR 95531 (2-(3-carboxypropyl)-3-amino-6-(4-methoxyphenyl)pyridazinium bromide) was higher in receptor complexes of α 1(ile 121) β 2 γ 2 than in those of α 1(val 121) β 2 γ 2 (IC 50 values were 32-fold and 26-fold lower for muscimol and GABA, respectively). The apparent affinity of the GABA A receptor antagonist bicuculline methiodide to inhibit the binding of [ 3H]SR 95531 did not differ between the two receptor complex variants. Electrophysiological measurements of GABA induced whole-cell Cl − currents showed a ten-fold decrease in the GABA A receptor sensitivity of α 1(val 121) β 2 γ 2 as compared to α 1(ile 121) β 2 γ 2 receptor complexes. Thus, a relatively small change in the primary structure of the α 1 subunit leads to a decrease selective for GABA A receptor sensitivity to agonist ligands, since no changes were observed in a GABA A receptor antagonist affinity and benzodiazepine receptor binding.