GABA(A) receptors, mediators of fast inhibitory neurotransmission, are heteropentameric assemblies from a large array of subunits. Differences in the sensitivity of receptor subtypes to endogenous GABA may permit subunit-dependent finely tuned responsiveness to the same GABAergic inputs. Using both radioligand binding and electrophysiology combined with mutagenesis, we identified a domain of four amino acids within the alpha subunits that mediates the distinct sensitivities to GABA allowing their selective switch between alphabeta3gamma2 combinations. Replacing this domain in alpha3 by the corresponding segments of alpha1-alpha5 resulted in mutant receptors displaying the GABA EC(50) values of the respective wild-type receptors. Vice versa, the alpha3 motif forced the low sensitivity to GABA of alpha3 upon alpha1beta3gamma2, alpha4beta3gamma2, and alpha5beta3gamma2. Binding of the GABA agonist [(3)H]muscimol was not affected by the exchange of the motif between alpha1 and alpha3 subunits. Thus, the equilibrium binding pocket is maintained upon replacement of the four amino acids. Taken together our data suggest that the identified motifs contribute to a structure involved in the transduction of the binding signal rather than to the binding itself.
