GABA antagonists differentiate between recombinant GABA(A)/benzodiazepine receptor subtypes

Abstract

Seventeen rat GABA(A) receptor subtypes were transiently expressed in the human embryonic kidney 293 cell line from α1, α2, α3, α5, or α6 variants with any of the three subunits and γ2S or γ3. We obtained fingerprints in the form of subtype characteristic concentration-response curves of 35S- TBPS binding to GABA and the GABA(A) antagonists SR 95531 and bicuculline. α3β3γ2S/3 and α5β3γ2S/3 containing receptors effectively recognized 35S-TBPS but not when β3 was replaced by the β1 or β2 subunit. This indicates a specific interaction of α and β variants to form high-affinity 35S-TBPS binding sites. At low levels GABA allosterically increased 35S- TBPS binding to all receptors with the concentration and magnitude depending on the subunit combination. Exchange of the β variant did not alter the concentration-response curves for α1 and α6 containing receptors but did so for α2 containing receptors. α2β1γ3 receptors displayed strong GABA- induced stimulation of 35S-TBPS binding, whereas binding to α2β3γ3 receptors was marginally increased. SR 95531 and bicuculline decreased 35S- TBPS binding to all γ3 containing receptors. In addition, bicuculline was effective on α1βxγ2 receptors. SR 95531 was threefold more potent than bicuculline in reversing GABA-induced modulation of 35S-TBPS binding in most receptor types, but was 30-fold more potent on α2β1γ3 and α6β1γ2S receptors. We conclude that the recognition site for GABA analogs on GABA(A) receptors is jointly affected by all three subunit classes present in a receptor.