NOS1‐dependent S‐nitrosylation of cardiac calcium‐handling proteins (RyR2, SERCA, and L‐type Ca2+ channel) modulates basal contractility and adrenergic inotropism

Abstract

We tested the hypothesis that NOS‐1 activity regulates basal and β‐adrenergic‐stimulated cardiac contractility by S‐nitrosylation of Ca2+‐handling proteins.We used the isolated rat heart preparation and biotin‐switch assay, to study the effect of selective NOS‐1 inhibitor (S‐methyl‐L‐thiocitrulline, SMTC) on the contractile response to isoproterenol (ISO) and S‐nitrosylation of sarcoplasmic Ca2+ channel (RyR2) and pump (SERCA2), L‐type Ca2+ channel (LTCC), and unspecific total cardiac proteins (TCP).SMTC (300nM, 20‐min) significantly decreased basal contractility (−15%) and the S‐nitrosylation of RYR2 (−70%), SERCA2 (−50%), LTCC (−80%), and TCP (−55%). ISO (10nM, 3‐min) increased S‐nitrosylation of RyR2 (+140%) and TCP (+30%), but did not change S‐nitrosylation of SERCA2 and LTCC. In the presence of SMTC, ISO caused proportionally similar changes in S‐nitrosylation of all these proteins, but the maximal inotropic response was reduced (−30%). Furthermore, in isolated mice cardiomyocytes, SMTC reduced ISO‐elicited maximal sarcomere shortening (−50%) and [Ca2+]i transient amplitude (−50%), and increased the time‐constant for [Ca2+]i decay (+30%).Thus, rapid and reversible NOS‐1 dependent S‐nitrosylation of specific Ca2+‐handling proteins is essential to keep basal cardiac contractility, and to accomplish a full inotropic response.Supported by Grants Fondecyt 1090757 & Anillos‐ACT71