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

Abstract

Recombinant human GABAA receptors were investigated in vitro by coexpression of cDNAs coding for α1, β2, and γ2 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 α1 subunit induces a marked decrease in agonist GABAA receptor ligand sensitivity. The potency of muscimol and GABA to inhibit the binding of the GABAA receptor antagonist [3H]SR 95531 (2-(3-carboxypropyl)-3-amino-6-(4-methoxyphenyl)pyridazinium bromide) was higher in receptor complexes of α1(ile 121)β2γ2 than in those of α1(val 121)β2γ2 (IC50 values were 32-fold and 26-fold lower for muscimol and GABA, respectively). The apparent affinity of the GABAA 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 GABAA receptor sensitivity of α1(val 121)β2γ2 as compared to α1(ile 121)β2γ2 receptor complexes. Thus, a relatively small change in the primary structure of the α1 subunit leads to a decrease selective for GABAA receptor sensitivity to agonist ligands, since no changes were observed in a GABAA receptor antagonist affinity and benzodiazepine receptor binding.