Mechanisms for picrotoxinin and picrotin blocks of alpha2 homomeric glycine receptors.
Résumé
Contrary to its effect on the -aminobutyric acid type A and C receptors, picrotoxin antagonism of the a1 homomeric glycine receptors (GlyRs) has been shown to be non-use-dependent and non-selective between the picrotoxin components picrotoxinin and picrotin. Picrotoxin antagonism of the embryonic a2 homomeric GlyR is known to be use-dependent, reflects a channel-blocking mechanism, but the selectivity of picrotoxin antagonism of the embryonic a2 homomeric GlyRs between picrotoxinin and picrotin is unknown. Hence, we used the patch-clamp recording technique in the outside-out configuration to investigate, at the single channel level, the mechanism of picrotin- and picrotoxinin-induced inhibition of currents, which were evoked by the activation of a2 homomeric GlyRs stably transfected into Chinese hamster ovary cells. Although both picrotoxinin and picrotin inhibited glycine-evoked outside-out currents, picrotin had a 30 times higher IC50 than picrotoxinin.. Picrotin-evoked inhibition displayed voltage-dependency, whereas picrotoxinin did not. Picrotoxinin and picrotin decreased the mean open time of the channel in a concentration-dependent manner, indicating that these picrotoxin components can bind to the receptor in its open state. When picrotin and glycine were co-applied, a large rebound current was observed at the end of the application. This rebound current was considerably smaller when picrotoxinin and glycine were co-applied. Both picrotin and picrotoxinin were unable to bind to the unbound conformation of the receptor, but could both be trapped at their binding site when the channel closed during glycine dissociation. Our data indicate that picrotoxinin and picrotin are not equivalent in blocking a2 homomeric GlyR.