As a major cause of aldosterone producing adenomas, numerous gain-of-function mutations in the KCNJ5 gene (encoding the K(+) channel subunit GIRK4) have been identified. The human adrenocortical carcinoma cell line NCI-H295R is the most frequently used cellular model for in vitro studies related to regulation of aldosterone-synthesis. Because of the undefined role of KCNJ5 (GIRK4) in regulating synthesis of aldosterone, we aimed at identifying basal and G protein-activated GIRK4 currents in this paradigmatic cell line. The GIRK-specific blocker Tertiapin-Q did not affect basal current. Neither loading of the cells with GTP-γ-S via the patch-clamp pipette nor agonist stimulation of an infected A1-adenosine receptor resulted in activation of GIRK current. In cells co-infected with KCNJ5, robust activation of basal and adenosine-activated inward-rectifying current was observed. Although GIRK4 protein can be detected in Western blots of H295R homogenates, we suggest that GIRK4 in aldosterone-producing cells does not form functional G(βγ)-activated channels.
Keywords: Adrenocortical cell line; Electrophysiology; GIRK4; KCNJ5; Primary aldosteronism.
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