Optimizing the expression of recombinant αβγ GABA A receptors in HEK293 cells for high-throughput screening

Abstract

Despite being important clinical targets, it is not straightforward to reliably express recombinant trimeric αβγ GABA-A receptors (GABAARs) for high-throughput screening. This study therefore sought to devise a simple and reliable means of transiently expressing α1β1γ1 and α1β1γ2 GABAARs in HEK293 cells. Expression efficiencies resulting from 5 different transfection strategies were assessed by flow cytometry and pharmacological analysis using an anion-sensitive yellow fluorescent protein-based assay. PolyFect™ and Effectene™, employed according to the manufacturers' instructions, conferred the strongest and most reliable expression of trimeric αβγGABAARs. Functional analysis via the yellow fluorescent protein assay revealed dramatic differences in the pharmacological properties of γ1- and γ2-containing receptors, consistent with previous electrophysiological characterizations. The authors conclude that this method of expressing and screening recombinant GABAARs provides an effective means of discovering novel GABAAR modulators for use as therapeutic lead compounds and pharmacological probes. © 2009 Society for Biomolecular Sciences.