STABLE EXPRESSION OF A FUNCTIONAL HOMOOLIGOMERIC DROSOPHILA GABA RECEPTOR IN A DROSOPHILA CELL-LINE

A cloned Drosophila gamma-aminobutyric acid (GABA) receptor subunit (Rdl) has been stably expressed as a functional homo-oligomeric ion channel in a Drosophila cell line. Stably-transfected clonal cell lines which expressed high levels of GABA receptor were identified by specific [H-3]-muscimol binding. Expression of functional GABA-gated ion channels in these cell lines was demonstrated by electrophysiological recording. Rapid and pronounced rundown of responses to GABA during whole-cell patch clamp recordings was overcome by the inclusion of EGTA in the pipette solution, indicating a possible role for calcium-dependent processes in the functional regulation of this GABA receptor. Relative agonist potencies of the expressed receptor were found to be in the order GABA = TACA > CACA. We have observed a reversible block of the receptor by the convulsant antagonists, picrotoxinin and EBOB, and by the insecticide fipronil. Potentiation of GABA responses was seen with the anaesthetic steroid 5 alpha-pregnan-3 alpha-ol-20-one. No significant effects (either agonist, antagonist or modulatory) were observed with bicuculline (a vertebrate GABA(A)R antagonist), benzodiazepines or barbiturates (vertebrate GABA(A)R modulators), or with glycine (agonist of the closely related vertebrate glycine receptors). The suitability of this Drosophila stable expression system for the characterization of receptors and ion channels is discussed.