Reduced phospholamban phosphorylation is associated with impaired relaxation in left ventricular myocytes from neuronal NO synthase-deficient mice.
Stimulation of nitric oxide (NO) release from the coronary endothelium facilitates myocardial relaxation via a cGMP-dependent reduction in myofilament Ca2+ sensitivity. Recent evidence suggests that NO released by a neuronal NO synthase (nNOS) in the myocardium can also hasten left ventricular relaxation; however, the mechanism underlying these findings is uncertain. Here we show that both relaxation (TR50) and the rate of [Ca2+]i transient decay (tau) are significantly prolonged in field-stimulated or voltage-clamped left ventricular myocytes from nNOS-/- mice and in wild-type myocytes (nNOS+/+) after acute nNOS inhibition. Disabling the sarcoplasmic reticulum abolished the differences in TR50 and tau, suggesting that impaired sarcoplasmic reticulum Ca2+ reuptake may account for the slower relaxation in nNOS-/- mice. In line with these findings, disruption of nNOS (but not of endothelial NOS) decreased phospholamban phosphorylation (P-Ser16 PLN), whereas nNOS inhibition had no effect on TR50 or tau in PLN-/- myocytes. Inhibition of cGMP signaling had no effect on relaxation in either group whereas protein kinase A inhibition abolished the difference in relaxation and PLN phosphorylation by decreasing P-Ser16 PLN and prolonging TR50 in nNOS+/+ myocytes. Conversely, inhibition of type 1 or 2A protein phosphatases shortened TR50 and increased P-Ser16 PLN in nNOS-/- but not in nNOS+/+ myocytes, in agreement with data showing increased protein phosphatase activity in nNOS-/- hearts. Taken together, our findings identify a novel mechanism by which myocardial nNOS promotes left ventricular relaxation by regulating the protein kinase A-mediated phosphorylation of PLN and the rate of sarcoplasmic reticulum Ca2+ reuptake via a cGMP-independent effect on protein phosphatase activity.
Zhang
Yin Hua
YH
Department of Cardiovascular Medicine, University of Oxford, John Radcliffe Hospital, United Kingdom.
Zhang
Mei Hua
MH
Sears
Claire E
CE
Emanuel
Krzysztof
K
Redwood
Charles
C
El-Armouche
Ali
A
Kranias
Evangelia G
EG
Casadei
Barbara
B
D000818:Animals; D002135:Calcium-Binding Proteins / Q000378:metabolism*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D006152:Cyclic GMP; D006352:Heart Ventricles / Q000473:pathology*; D051379:Mice; D018345:Mice, Knockout; D032342:Muscle Cells / Q000378:metabolism / Q000473:pathology*; D009200:Myocardial Contraction; D052248:Nitric Oxide Synthase Type I / Q000172:deficiency / Q000502:physiology*; D010766:Phosphorylation; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases / Q000378:metabolism; D000099211:Phospholamban
D002135:Calcium-Binding Proteins; D017868:Cyclic AMP-Dependent Protein Kinases; D006152:Cyclic GMP; D052248:Nitric Oxide Synthase Type I; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases; D000099211:Phospholamban