Mechanismen der Urocortin-II-induzierten Stimulation der NO-Produktion in isolierten Kaninchen-Ventrikelmyozyten

Abstract

Urocortin II (UcnII), a peptide from corticotropin-releasing factor (CRF) family, is exhibiting beneficial effects in heart failure. In cardiomyocytes, it elicits a cAMP- and Ca2+-dependent positive inotropic and lusitropic effect. Our aim was to characterize the UcnII activities on cardiac nitric oxide (NO) signalling and to evaluate the underlying signalling pathways. UcnII (100nM) caused a time-dependent increase in phosphorylation of Akt at Ser473 and Thr308 and endothelial NO-synthase (eNOS) at Ser1177, which were blocked by wortmannin (0.3 µM) and LY294002 (10 µM), two inhibitors of phosphatidylinositol 3-kinase (PI3K). Confocal imaging of cellular NO-production and fractional shortening revealed an increase of [NO]i and fractional shortening with a similar time course, that was diminished by inhibitors of eNOS, L-NAME (1mM) and L-NIO (10µM). The inhibitor of protein kinase A (PKA), H89 (5µM), significantly abolished the UcnII-induced phosphorylation of eNOS, but did not affect the phosphorylation of Akt. When both, PI3K/Akt and cAMP/PKA signaling, were inhibited the UcnII-induced synthesis of [NO]i was reduced. We show, for the first time, that in rabbit ventricular myocytes, UcnII activates cAMP/PKA and PI3K/Akt signaling and that both pathways converge in the phosphorylation of eNOS to stimulate NO production and contribute to the increased contractility.