Abstract P076: Essential Role of Neuronal Nitric Oxide Synthase in Ca 2+ /Calmodulin-Dependent Protein Kinase II Activation in Cardiac Myocytes During β-Adrenergic Stimulation

Abstract

RATIONALE: Stimulation of the beta-adrenergic (beta-AR) pathway leads to positive inotropy, and is the major regulator of heart function. In addition to the traditional PKA pathway, activation of Ca 2+ /calmodulin-dependent protein kinase II (CaMKII) and neuronal nitric oxide synthase (NOS1) signaling also play important roles in the positive inotropy by modulating ryanodine receptor (RyR) activity. OBJECTIVE: The upstream activators of CaMKII during beta-AR stimulation are not well defined. The purpose of this study is to investigate if there is any cross-talk between the CaMKII and NOS1 signaling pathways. METHODS AND RESULTS: Myocytes were isolated from wildtype (WT, C57Bl/6) and NOS1 −/− mice. Ca 2+ transients (Fluo-4) and cell shortening (edge detection) were simultaneously measured. RyR activity was measured using the SR Ca 2+ leak/load relationship. CaMKII was acutely inhibited by KN93. In WT myocytes, KN93 decreased beta-AR stimulated contraction (Ca2+ transients (Fluo-4) and cell shortening). In NOS1 −/− myocytes, beta-AR stimulated contraction was blunted compared to WT, and KN93 had no further effect on contraction. Furthermore, beta-AR stimulated RyR activity was blunted in NOS1 −/− compared to WT myocytes. As with contraction, KN93 decreased beta-AR stimulated RyR activity in WT myocytes, but had no effect in NOS1 −/− myocytes. CONCLUSION: These data suggest that NOS1 is required for CaMKII-mediated RyR activation which contributes to positive inotropy during beta-AR stimulation. Further study of this pathway is warranted since CAMKII expression and activity are increased in cardiac hypertrophy and heart failure. A better understanding of the NOS1/CaMKII pathway during beta-AR stimulation has beneficial therapeutic potential for heart diseases.