title: NCI-H295R cell line as in vitro model of hyperaldosteronism lacks functional KCNJ5 (GIRK4; Kir3.4) channels.
abstract: 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.
authors.affiliation: Institute of Physiology, Ruhr-University Bochum, D-44780 Bochum, Germany. Electronic address: cecile.kienitz@rub.de.
authors.forename: Marie-Cécile
authors.initials: MC
authors.lastname: Kienitz
authors.affiliation: Department of Pharmacology and Toxicology, Ruhr-University Bochum, D-44780 Bochum, Germany.
authors.forename: Evanthia
authors.initials: E
authors.lastname: Mergia
authors.affiliation: Institute of Physiology, Ruhr-University Bochum, D-44780 Bochum, Germany.
authors.forename: Lutz
authors.initials: L
authors.lastname: Pott
mesh_terms: D045744:Cell Line, Tumor; D002462:Cell Membrane / Q000378:metabolism; D051676:G Protein-Coupled Inwardly-Rectifying Potassium Channels / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D006929:Hyperaldosteronism / Q000235:genetics / Q000378:metabolism*; D008564:Membrane Potentials; D021122:Protein Subunits / Q000235:genetics / Q000378:metabolism; D021381:Protein Transport
chemical_list: D051676:G Protein-Coupled Inwardly-Rectifying Potassium Channels; C484274:KCNJ5 protein, human; D021122:Protein Subunits
keywords: Adrenocortical cell line; Electrophysiology; GIRK4; KCNJ5; Primary aldosteronism