A pharmacological characterization of GABA, THIP and DS2 at binary alpha 4 beta 3 and beta 3 delta receptors: GABA activates beta 3 delta receptors via the beta 3(+) delta (-) interface

There is growing evidence that GABA ( gamma -aminobutyric acid) can activate GABAA receptors (GABAARs) in the absence of an alpha subunit. In this study, we compared the pharmacology of homomeric and binary alpha 4, beta 3 or delta subunits with ternary alpha 4 beta 3 delta to identify subunit interfaces that contribute to the pharmacology of GABA, THIP, and DS2, and the antagonists, Zn2+, gabazine and bicuculline. beta 3 delta receptors form functional GABA-gated channels when expressed in Xenopus oocytes with a pharmacology that differs to homomeric beta 3, binary alpha 4 beta 3 and ternary alpha 4 beta 3 delta receptors. GABA had similar potency at alpha 4 beta 3 and beta 3 delta receptors (25 mu M and 26 mu M, respectively) but differed at alpha 4 beta 3 delta receptors where GABA exhibited a biphasic concentration-response (EC50 (1)=12.6nM; EC50 (2)=6.3 mu M). THIP activated beta 3 delta receptors (EC50=456 mu M) but was a more potent activator of alpha 4 beta 3 (EC50=27 mu M) and alpha 4 beta 3 delta receptors (EC50 (1)=27.5nM; EC50 (2)=29.5 mu mu ), indicating that the alpha 4 subunit significantly contribute to its potency. The delta -preferring modulator, DS2 had marginal or no effect at beta 3 delta and alpha 4 beta 3 receptors, indicating a role for both the alpha 4 and delta subunits for its potency. Gabazine inhibited GABA-elicited currents at beta 3 delta receptors whereas bicuculline activated these receptors. Mutational analysis verified that GABA binds to the beta 3(+) delta (-) interface formed by the beta 3 and delta subunits. In conclusion, evaluating agents against binary GABAARs such as beta 3 delta and alpha 4 beta 3 receptors enables identification of interfaces that may contribute to the pharmacology of the more complex ternary alpha 4 beta 3 delta receptors.