Hypercontractile Female Hearts Exhibit Increased S -Nitrosylation of the L-Type Ca 2+ Channel α1 Subunit and Reduced Ischemia/Reperfusion Injury

Abstract

Mechanisms underlying gender differences in cardiovascular disease are poorly understood. We found previously that, under hypercontractile conditions, female hearts exhibit significantly less ischemia/reperfusion injury than males. Here we show that male wild-type (WT) mouse hearts pretreated with 10 nmol/L isoproterenol before ischemia exhibited increased injury versus female hearts, but this relative protection in females was absent in eNOS −/− and nNOS −/− hearts. In isoproterenol-treated female versus male hearts, there was also more endothelial NO synthase (eNOS) associated with cardiomyocyte caveolin-3, and more neuronal NOS (nNOS) translocation to caveolin-3 during ischemia/reperfusion. S -nitrosothiol (SNO) formation was increased in isoproterenol-treated ischemic/reperfused hearts in all mouse genotypes, but only in WT mice was SNO content significantly higher in females than males. Using the biotin switch method, we identified the L-type Ca 2+ channel α1 subunit as the predominant S -nitrosylated protein in membrane fractions, and following isoproterenol and ischemia/reperfusion male/female differences in SNO were seen only in WT hearts, but not in constitutive NOS −/− genotypes. The isoproterenol-induced increase in L-type Ca 2+ current ( I Ca ) was smaller in females versus in males, but NOS blockade increased I Ca in females. This gender difference in I Ca in isoproterenol-treated myocytes (and abolition on NOS inhibition) was mirrored exactly in Ca 2+ transients and SR Ca 2+ contents. In conclusion, these data suggest that eNOS and nNOS both play roles in the gender differences observed in ischemia/reperfusion injury under adrenergic stimulation, and also demonstrate increased S -nitrosylation of the L-type Ca 2+ channels in female cardiomyocytes.