Mechanisms of potentiation of the mammalian GABAA receptor by the marine cembranoid eupalmerin acetate.
OBJECTIVE
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.


METHODS
Whole-cell and single-channel recordings were made from HEK 293 cells transiently expressing rat wild-type and mutant alpha1beta2gamma2L GABA(A) receptors.


RESULTS
Our findings demonstrate that, at micromolar concentrations, EPA potentiates the rat alpha1beta2gamma2L 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 alpha1Q241 and alpha1N407/Y410, previously shown to affect receptor modulation by neurosteroids, also diminished potentiation by EPA. Exposure to a steroid antagonist, (3alpha,5alpha)-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 microM) of allopregnanolone. In tadpole behavioural assays, EPA caused loss of righting reflex and loss of swimming reflex.


CONCLUSIONS
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.
Li
P
P
Department of Anesthesiology, Washington University School of Medicine, St Louis, MO 63110, USA.
Reichert
D E
DE
Rodríguez
A D
AD
Manion
B D
BD
Evers
A S
AS
Eterović
V A
VA
Steinbach
J H
JH
Akk
G
G
D000737:Androstenols / Q000494:pharmacology; D000818:Animals; D001522:Behavior, Animal / Q000187:drug effects; D001665:Binding Sites; D002460:Cell Line; D003063:Cnidaria / Q000737:chemistry*; D004224:Diterpenes / Q000008:administration & dosage / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D004594:Electrophysiology; D006801:Humans; D007814:Larva; D009154:Mutation; D011280:Pregnanolone / Q000008:administration & dosage / Q000494:pharmacology; D021122:Protein Subunits; D051381:Rats; D011963:Receptors, GABA-A / Q000187:drug effects* / Q000378:metabolism; D014982:Xenopus laevis
C485374:17-phenylandrost-16-en-3-ol; D000737:Androstenols; D004224:Diterpenes; D021122:Protein Subunits; D011963:Receptors, GABA-A; C082581:eupalmerin acetate; D011280:Pregnanolone