Mechanisms of potentiation of the mammalian GABA _A receptor by the marine cembranoid eupalmerin acetate

Background and purpose: Eupalmerin acetate (EPA) is a marine diterpene compound isolated from the gorgonian octocorals Eunicea succinea and Eunicea mammosa. The compound has been previously shown to modulate muscle-type and neuronal nicotinic acetylcholine receptors, which are inhibited in the presence of low micromolar concentrations of EPA. In this study, we examined the effect of EPA on another transmitter-gated ion channel, the GABA _A receptor. Experimental approach: Whole-cell and single-channel recordings were made from HEK 293 cells transiently expressing rat wild-type and mutant α1β2γ2L GABA _A receptors. Key results: Our findings demonstrate that, at micromolar concentrations, EPA potentiates the rat α1β2γ2L GABA _A receptor. The analysis of single-channel currents recorded in the presence of EPA showed that the kinetic mode of action of EPA is similar to that of neuroactive steroids. Mutations to residues α1Q241 and α1N407/Y410, previously shown to affect receptor modulation by neurosteroids, also diminished potentiation by EPA. Exposure to a steroid antagonist, (3α,5α)-17-phenylandrost-16-en-3-ol, reduced potentiation by EPA. Additionally, exposure to EPA led to potentiation of GABA _A receptors activated by very high concentrations (1-10 μM) of allopregnanolone. In tadpole behavioural assays, EPA caused loss of righting reflex and loss of swimming reflex. Conclusions and implications: We conclude that EPA either interacts with the putative neurosteroid binding site on the GABA _A receptor or shares with neurosteroids the key transduction elements involved in channel potentiation by steroids. The results indicate that cembranoids represent a novel class of GABA _A receptor modulators.