Abstract

Picrotoxin is a plant alkaloid that is often used to block the activity of neuronal GABA and glycine receptors. However, the mechanism by which picrotoxin inhibits these receptors is still in debate. In this study, we investigated the picrotoxin inhibition on perch-ρ subunits expressed heterologously in Xenopus laevis oocytes, and on native GABA_C receptors of perch bipolar cells. Both competitive and noncompetitive mechanisms were observed for picrotoxin inhibition of the GABA_C receptor. In oocytes expressing the ρ1A subunit, terminating simultaneously the coapplication of GABA and picrotoxin induced a large rebound of membrane current. In addition, picrotoxin significantly accelerated the kinetics of GABA responses, particularly in the relaxation (offset) phase of GABA currents. Both current rebound and the large acceleration of GABA relaxation were unique to picrotoxin inhibition and were not observed with the competitive antagonist (1,2,5,6-tetrahydropyridin-4-yl)-methylphosphinic acid or the allosteric modulator zinc. The change in kinetics induced by picrotoxin was also observed on receptors formed by other GABA ρ subunits, as well as on the GABA_C receptors of retinal bipolar cells. Based on these observations, we proposed a model in which picrotoxin binds to the GABA_C receptor in both channel open and closed states. Overall, this model provides a remarkably good approximation of the experimental findings we observed for picrotoxin inhibition of GABA_C receptors. These results support an allosteric mechanism of picrotoxin inhibition of ligand-gated chloride channels.