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PLoS One. 2014 Feb 3;9(2):e87495. doi: 10.1371/journal.pone.0087495. eCollection 2014.

Nitric oxide-dependent activation of CaMKII increases diastolic sarcoplasmic reticulum calcium release in cardiac myocytes in response to adrenergic stimulation.

Author information

1
Davis Heart and Lung Research Institute, Department of Physiology and Cell Biology, The Ohio State University, Columbus, Ohio, United States of America.
2
Department of Molecular Biophysics and Physiology, Rush University, Chicago, Illinois, United States of America.
3
Department of Pharmacology, University of California Davis, Davis, California, United States of America.

Abstract

Spontaneous calcium waves in cardiac myocytes are caused by diastolic sarcoplasmic reticulum release (SR Ca(2+) leak) through ryanodine receptors. Beta-adrenergic (β-AR) tone is known to increase this leak through the activation of Ca-calmodulin-dependent protein kinase (CaMKII) and the subsequent phosphorylation of the ryanodine receptor. When β-AR drive is chronic, as observed in heart failure, this CaMKII-dependent effect is exaggerated and becomes potentially arrhythmogenic. Recent evidence has indicated that CaMKII activation can be regulated by cellular oxidizing agents, such as reactive oxygen species. Here, we investigate how the cellular second messenger, nitric oxide, mediates CaMKII activity downstream of the adrenergic signaling cascade and promotes the generation of arrhythmogenic spontaneous Ca(2+) waves in intact cardiomyocytes. Both SCaWs and SR Ca(2+) leak were measured in intact rabbit and mouse ventricular myocytes loaded with the Ca-dependent fluorescent dye, fluo-4. CaMKII activity in vitro and immunoblotting for phosphorylated residues on CaMKII, nitric oxide synthase, and Akt were measured to confirm activity of these enzymes as part of the adrenergic cascade. We demonstrate that stimulation of the β-AR pathway by isoproterenol increased the CaMKII-dependent SR Ca(2+) leak. This increased leak was prevented by inhibition of nitric oxide synthase 1 but not nitric oxide synthase 3. In ventricular myocytes isolated from wild-type mice, isoproterenol stimulation also increased the CaMKII-dependent leak. Critically, in myocytes isolated from nitric oxide synthase 1 knock-out mice this effect is ablated. We show that isoproterenol stimulation leads to an increase in nitric oxide production, and nitric oxide alone is sufficient to activate CaMKII and increase SR Ca(2+) leak. Mechanistically, our data links Akt to nitric oxide synthase 1 activation downstream of β-AR stimulation. Collectively, this evidence supports the hypothesis that CaMKII is regulated by nitric oxide as part of the adrenergic cascade leading to arrhythmogenesis.

PMID:
24498331
PMCID:
PMC3911966
DOI:
10.1371/journal.pone.0087495
[Indexed for MEDLINE]
Free PMC Article
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