Abstract 253: S-Nitrosylation-Based Signaling Regulates Myocardial Contractility
Introduction: The downstream NO signaling pathways regulating myocardial contractility remain controversial. S-nitrosoglutathione reductase (GSNOR) serves a newly appreciated function in metabolizing S-nitrosothiols (SNO). Here we tested the hypothesis that SNO signaling plays a fundamental role in myocardial contractility by assessing the inotropic response in isolated myocytes from a GSNOR-null mouse.
Methods and results: Western-blot and confocal microscopy revealed the presence of GSNOR in the hearts of wild type but not GSNOR−/ − mice. Sarcomere shortening (SL) and Ca2+ transient ([Ca2+]i) was measured across a range of isoproterenol concentrations (10−10 -10−6 M). In GSNOR−/ − myocytes, the SL and [Ca2+]i responses were lower than in WT (P<0.05, n=10). Inhibition of NO production with L-NMMA (NO synthase inhibitor) restored both SL and [Ca2+]i in GSNOR−/ − myocytes, whereas inhibition of the cGMP-PKG pathway with KT5823 had no effect (P<0.05). Furthermore, elimination of NO production by NOS3 restored contractile responses in a GSNOR-null background (NOS3−/ −/GSNOR−/ −). Protein S-nitrosylation in whole heart homogenates, as assessed by biotin-switch, was elevated in GSNOR−/ − vs. WT (P<0.05, n=3) and a number of calcium-cycling SNO-proteins were identified, including, ryanodine receptor, L-type Ca2+ channel and SERCA2 (Na+/Ca2+ -exchanger was not among the SNO substrates)
Conclusion: Our findings support the notion that SNO signaling is a major participant in myocardial contractility through effects on calcium cycling. The absence of GSNOR, which would normally curtail SNO signaling, leads to increased levels of SNO proteins, many of which are cardiac ion channels, with consequent effect upon E-C coupling. These data indicate a greater role for SNOs in cardiac contractility than has been previously appreciated and highlight a key role for the GSNOR in cardiac physiology.