SERCA Cys674 sulphonylation and inhibition of L-type Ca2+ influx contribute to cardiac dysfunction in endotoxemic mice, independent of cGMP synthesis

The goal of this study was to identify the cellular mechanisms responsible for cardiac dysfunction in endotoxemic mice. We aimed to differentiate the roles of cGMP [produced by soluble guanylyl cyclase (sGC)] versus oxidative posttranslational modifications of Ca2+ transporters. C57BL/6 mice [wild-type (WT) mice] were administered lipopolysaccharide (LPS; 25 mu g/g ip) and euthanized 12 h later. Cardiomyocyte sarcomere shortening and Ca2+ transients (Delta Ca-i) were depressed in LPS-challenged mice versus baseline. The time constant of Ca2+ decay (tau(Ca)) was prolonged, and sarcoplasmic reticulum Ca2+ load (Ca-SR) was depressed in LPS-challenged mice (vs. baseline), indicating decreased activity of sarco(endo)plasmic Ca2+-ATPase (SERCA). L-type Ca2+ channel current (I-Ca,I-L) was also decreased after LPS challenge, whereas Na+/Ca2+ exchange activity, ryanodine receptors leak flux, or myofilament sensitivity for Ca2+ were unchanged. All Ca2+-handling abnormalities induced by LPS (the decrease in sarcomere shortening, Delta Cai, CaSR, ICa, L, and tau(Ca) prolongation) were more pronounced in mice deficient in the sGC main isoform (sGC alpha(-/-)(1) mice) versus WT mice. LPS did not alter the protein expression of SERCA and phospholamban in either genotype. After LPS, phospholamban phosphorylation at Ser(16) and Thr(17) was unchanged in WT mice and was increased in sGC alpha(-/-)(1) mice. LPS caused sulphonylation of SERCA Cys(674) (as measured immunohistochemically and supported by iodoacetamide labeling), which was greater in sGC alpha(-/-)(1) versus WT mice. Taken together, these results suggest that cardiac Ca2+ dysregulation in endotoxemic mice is mediated by a decrease in L-type Ca2+ channel function and oxidative posttranslational modifications of SERCA Cys(674), with the latter (at least) being opposed by sGC-released cGMP.