Abstract 488: Neuronal Nitric Oxide Synthase Enhances Cardiac Contractility by Modulation of Phospholamban Phosphorylation
Neuronal nitric oxide synthase (nNOS, NOS1) is localized to the sarcoplasmic reticulum (SR). However, its effects on contraction are controversial. Studies have shown that NOS1 knockout (NOS1−/ −) mice have decreased basal contractility and β-adrenergic responsiveness. However, other studies have demonstrated the opposite effects. Nevertheless, studies consistently have shown a slowed rate of relaxation in NOS1−/ − mice. In addition, limited studies have investigated the mechanisms of NOS1 regulation of contractility. The purpose of this study is to determine the involvement of phospholamban (PLB) in the functional effects of NOS1. We hypothesize that PLB is a major end target of NOS1 signaling. We simultaneously measured Ca2+ transients (Fluo-4) and myocyte shortening (video edge detection) in myocytes isolated from wildtype (WT), NOS1−/ − and PLB knockout (PLB−/ −) mice. Basal contractility was decreased with NOS1 knockout or specific NOS1 inhibition. That is, NOS1−/ − myocytes or specific NOS1 inhibition with S-Methyl-L-Thiocitrulline (SMLT) in WT myocytes decreased Ca2+ transient amplitude (47% and 67% of WT, respectively; P<0.05), shortening amplitude (46% and 51% of WT; P<0.05), slowed the decline of [Ca]i (162% and 148% of WT, P<0.05) and decreased SR Ca2+ load (71% and 71% of WT, P<0.05). SMLT had no effect in NOS1−/ − myocytes. Interestingly, NOS1 inhibition with SMLT in PLB−/ − myocytes had no effect on Ca2+ transient amplitude (101% of PLB−/ − control), shortening amplitude (87% of PLB−/ − control) or rate of [Ca]i decline (112% of PLB−/ − control). In addition, NOS1−/ − hearts had a decrease in basal PLB phosphorylation at Ser16 (47% of WT, P<0.05). Our results suggest that NOS1 signaling leads to a positive inotropic and lusitropic effect. NOS1 signaling increases PLB phosphorylation leading to greater SR Ca2+ load, which ultimately increases contractility. Thus, PLB is the major end target for NOS1 signaling.