BETA3‐ADRENERGIC RECEPTOR REGULATION OF nNOS SIGNALING IN CARDIOMYOCYTES

Abstract

β3‐AR plays a pivotal role in modulating cardiac function. We recently showed that β3‐mediated cardioprotection is lost in nNOS−/− animals under pressure overload as compared with WT. However, the manner in which β3‐AR regulates nNOS signaling at the cardiomyocyte level is unknown. Here, we used specific β3‐ AR agonist BRL37344 (BRL) to assess the role of β3‐AR regulation of nNOS in hypertrophied isolated neonatal rat ventricular cardiomyocytes. Hypertrophy was induced by 72‐hour norepinephrine pretreatment, which increased cell size and β3‐AR mRNA as compared with non‐hypertrophied cells. In hypertrophied myocytes, BRL increased NOS activity, nNOS phosphorylation at stimulatory site Ser1416, and dephosphorylation of deactivation site Ser847. NOS activity was attenuated by nNOS inhibitor LVNIO and phosphomemetic mutants Ser1416A and Ser847D. In addition, Gi/o and Akt inhibition suppressed BRL‐induced nNOS‐Ser1416 phosphorylation and NOS activity. Our data suggest that BRL regulates myocardial NOS activity via nNOS phosphorylation. This is first study to demonstrate a role for nNOS phosphorylation as a key factor in β3‐AR signaling. These results contribute significantly to our understanding of the negative inotropic properties of cardiac β3‐AR signaling during sympathetic overstimulation, and will ultimately aid in drug discoveries that target molecular mechanisms associated heart failure.