Structural domains of the human GABA sub(A) receptor beta 3 subunit involved in the actions of pentobarbital

* 1 This study was conducted to search for the residues of the beta 3 subunit which affect pentobarbital action on the gamma -aminobutyric acid type A (GABA sub(A)) receptor. Three chimeras were constructed by joining the GABA sub(A) receptor beta 3 subunit to the rho 1 subunit. For each chimera, the N-terminal sequence was derived from the beta 3 subunit and the C-terminal sequence from the rho 1 subunit, with junctions located between the membrane-spanning regions M2 and M3, in the middle of M2, or in M1, respectively. * 2 In receptors obtained by the coexpression of alpha 1 with the chimeric subunits, in contrast with those obtained by the coexpression of alpha 1 and beta 3, pentobarbital exhibited lower potentiation of GABA-evoked responses, and in the direct gating of Cl super(-) currents, an increase in the EC sub(50) together with a marked decrease in the relative maximal efficacy compared with that of GABA. * 3 Estimates of the channel opening probability through variance analysis and single-channel recordings of one chimeric subunit showed that the reduced relative efficacy for gating largely resulted from an increase in gating by GABA, with little change in efficacy of pentobarbital. * 4 A fit of the time course of the response by the predictions of a class of reaction schemes is consistent with the conclusion that the change in the concentration dependence of activation by pentobarbital is due to a change in pentobarbital affinity for the receptor. Therefore, the data suggest that residues of the beta 3 subunit involved in pentobarbital binding to GABA sub(A) receptors are located downstream from the middle of the M2 region.