Positioning of the α-subunit isoforms confers a functional signature to γ-aminobutyric acid type A receptors

摘要

Fast synaptic inhibitory transmission in the CNS is mediated by γ-aminobutyric acid type A (GABAA) receptors. They belong to the ligand-gated ion channel receptor superfamily, and are constituted of five subunits surrounding a chloride channel. Their clinical interest is highlighted by the number of therapeutic drugs that act on them. It is well established that the subunit composition of a receptor subtype determines its pharmacological properties. We have investigated positional effects of two different α-subunit isoforms, α1 and α6, in a single pentamer. For this purpose, we used concatenated subunit receptors in which subunit arrangement is predefined. The resulting receptors were expressed in Xenopus oocytes and analyzed by using the two-electrode voltage-clamp technique. Thus, we have characterized γ2β2α1β2α1, γ2β2α6β2α6, γ2β2α1β₂α₆, and γ₂β₂α₆β₂α₁ GABA_A receptors. We investigated their response to the agonist GABA, to the partial agonist piperidine-4-sulfonic acid, to the noncompetitive inhibitor furosemide and to the positive allosteric modulator diazepam. Each receptor isoform is characterized by a specific set of properties. In this case, subunit positioning provides a functional signature to the receptor. We furthermore show that a single α₆-subunit is sufficient to confer high furosemide sensitivity, and that the diazepam efficacy is determined exclusively by the α-subunit neighboring the γ₂-subunit. By using this diagnostic tool, it should become possible to determine the subunit arrangement of receptors expressed in vivo that contain α₁- and α₆-subunits. This method may also be applied to the study of other ion channels.