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(Circulation Research. 2009;105:383.)
© 2009 American Heart Association, Inc.

Clinical Research

Nitric Oxide Increases Cardiac I_K1 by Nitrosylation of Cysteine 76 of Kir2.1 Channels

Ricardo Gómez^*, Ricardo Caballero^*, Adriana Barana, Irene Amorós, Enrique Calvo, Juan Antonio López, Helene Klein, Miguel Vaquero, Lourdes Osuna, Felipe Atienza, Jesús Almendral, Ángel Pinto, Juan Tamargo, Eva Delpón

From the Department of Pharmacology (R.G., R.C., A.B., I.A., M.V., L.O., J.T., E.D.), School of Medicine, Universidad Complutense, Madrid, Spain; Centro Nacional de Investigaciones Cardiovasculares (E.C., J.A.L.), Madrid, Spain; Department of Physiology, Montreal University (H.K.), Canada; and Hospital General Universitario Gregorio Marañón (F.A., J.A., A.P.), Madrid, Spain.

Correspondence to Ricardo Caballero, Department of Pharmacology, School of Medicine, Universidad Complutense, 28040 Madrid, Spain. E-mail rcaballero{at}med.ucm.es

Rationale: The cardiac inwardly rectifying K⁺ current (I_K1) plays a critical role in modulating excitability by setting the resting membrane potential and shaping phase 3 of the cardiac action potential.

Objective: This study aims to analyze the effects of nitric oxide (NO) on human atrial I_K1 and on Kir2.1 channels, the major isoform of inwardly rectifying channels present in the human heart.

Methods and Results: Currents were recorded in enzymatically isolated myocytes and in transiently transfected CHO cells, respectively. NO at myocardial physiological concentrations (25 to 500 nmol/L) increased inward and outward I_K1 and I_Kir2.1. These effects were accompanied by hyperpolarization of the resting membrane potential and a shortening of the duration of phase 3 of the human atrial action potential. The I_Kir2.1 increase was attributable to an increase in the open probability of the channel. Site-directed mutagenesis analysis demonstrated that NO effects were mediated by the selective S-nitrosylation of Kir2.1 Cys76 residue. Single ion monitoring experiments performed by liquid chromatography/tandem mass spectrometry suggested that the primary sequence that surrounds Cys76 determines its selective S-nitrosylation. Chronic atrial fibrillation, which produces a decrease in NO bioavailability, decreased the S-nitrosylation of Kir2.1 channels in human atrial samples as demonstrated by a biotin-switch assay, followed by Western blot.

Conclusions: The results demonstrated that, under physiological conditions, NO regulates human cardiac I_K1 through a redox-related process.

Key Words: nitric oxide • Kir2.1 channels • resting membrane potential • cardiac myocytes • I_K1