Amiloride, a diuretic used in the treatment of hypertension and congestive heart failure, and 2-guanidine-4-methylquinazoline (GMQ) are guanidine compounds that modulate acid-sensing ion channels. Both compounds have demonstrated affinity for a variety of membrane proteins, including members of the Cysloop family of ligand-gated ion channels, such as the heteromeric GABA-A abg receptors. The actions of these guanidine compounds on the homomeric GABA-A r1 receptor remains unclear, especially in light of how many GABA-A abg receptor modulators have different effects in the GABA-A r1 receptors. We sought to characterize the influence of amiloride and GMQ on the human GABA-A r1 receptors using whole-cell patch-clamp electrophysiology. The diuretic amiloride potentiated the human GABA-A r1 GABA-mediated current, whereas GMQ antagonized the receptor. Furthermore, a GABA-A second transmembrane domain site, the intersubunit site, responsible for allosteric modulation in the heteromeric GABA-A receptors mediated amiloride’s positive allosteric actions. In contrast, the mutation did not remove GMQ antagonism but only changed the guanidine compound’s potency within the human GABA-A r1 receptor. Through modeling and introduction of point mutations, we propose that the GABA-A r1 intersubunit site plays a role in mediating the allosteric effects of amiloride and GMQ.
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