Regulation of cardiac CFTR Cl(-) channel activity by a Mg(2+)-dependent protein phosphatase.

Abstract

Dephosphorylation of the CFTR Cl(-) channel is known to be induced by both okadaic-acid- (OA-) sensitive and -insensitive protein phosphatases (PPs). In the present study, the effects of cytosolic free Mg(2+) on the cardiac CFTR Cl(-) current were examined in relation to the latter PP activity in guinea pig ventricular myocytes. Even when maintaining intracellular Mg-ATP at millimolar concentrations under whole-cell patch-clamp mode, cAMP-activated Cl(-) conductance was reversibly suppressed by cytosolic free Mg(2+), with an IC(50) of around 2.5 mmol/l. In contrast, changes in the cytosolic concentration of free Mg(2+) ([Mg(2+)](i)) had no effect on genistein-activated CFTR Cl(-) currents. The Mg(2+) effect on cAMP-activated CFTR Cl(-) conductance was completely reversed by application of anthracene-9-carboxylic acid (9-AC), which was previously shown to inhibit an OA-insensitive PP in cardiac myocytes. A 9-AC-sensitive fraction of endogenous PP activity in the extract of guinea pig ventricle was found to be activated by free Mg(2+) at millimolar concentrations but to be inactive at micromolar concentrations. The intracellular application of OA failed to activate basal Cl(-) conductance at millimolar [Mg(2+)](i). In the presence of OA, however, basal Cl(-) conductance became activated either by reducing [Mg(2+)](i) to micromolar concentrations or by applying 9-AC. Thus, we conclude that a Mg(2+)-dependent PP sensitive to 9-AC plays a key role in the cAMP-mediated regulation of cardiac CFTR Cl(-) channel at physiological [Mg(2+)](i)under both basal and cAMP-activated conditions. Also, it appears that the genistein-activated conformation of the cardiac CFTR channel is not sensitive to the Mg(2+)-dependent PP.