Nitric oxide increases cardiac IK1 by nitrosylation of cysteine 76 of Kir2.1 channels.
BACKGROUND
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.


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.
Gómez
Ricardo
R
Department of Pharmacology, School of Medicine, Universidad Complutense, 28040 Madrid, Spain.
Caballero
Ricardo
R
Barana
Adriana
A
Amorós
Irene
I
Calvo
Enrique
E
López
Juan Antonio
JA
Klein
Helene
H
Vaquero
Miguel
M
Osuna
Lourdes
L
Atienza
Felipe
F
Almendral
Jesús
J
Pinto
Angel
A
Tamargo
Juan
J
Delpón
Eva
E
D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D003545:Cysteine / Q000235:genetics / Q000378:metabolism; D045462:Endothelium-Dependent Relaxing Factors / Q000378:metabolism / Q000494:pharmacology*; D005260:Female; D006325:Heart Atria / Q000166:cytology / Q000378:metabolism; D006801:Humans; D051660:Intermediate-Conductance Calcium-Activated Potassium Channels / Q000235:genetics / Q000378:metabolism*; D008297:Male; D008564:Membrane Potentials / Q000187:drug effects* / Q000502:physiology; D016297:Mutagenesis, Site-Directed; D032383:Myocytes, Cardiac / Q000166:cytology / Q000378:metabolism*; D009569:Nitric Oxide / Q000378:metabolism / Q000494:pharmacology*; D010084:Oxidation-Reduction / Q000187:drug effects; D024661:Potassium Channels, Inwardly Rectifying / Q000235:genetics / Q000378:metabolism*
D045462:Endothelium-Dependent Relaxing Factors; D051660:Intermediate-Conductance Calcium-Activated Potassium Channels; C489457:KCNJ2 protein, human; C496923:KCNN4 protein, human; D024661:Potassium Channels, Inwardly Rectifying; D009569:Nitric Oxide; D003545:Cysteine