Angiotensin II-induced negative inotropy in rat ventricular myocytes: role of reactive oxygen species and p38 MAPK

Abstract

The octapeptide angiotensin II (ANG II) can modulate cardiac contractility and is increased in heart failure, where contractile function is impaired. In rat cardiac myocytes, 1 μM of ANG II produces a negative inotropic effect (NIE) (24.6 ± 5% reduction). However, the subcellular signaling involved in this effect remains elusive. We examined the mechanisms and signaling events involved in the reduction in contractile function induced by the peptide in indo-1-loaded rat cardiomyocytes. The results showed that the NIE of ANG II was not associated with a parallel decrease in the intracellular Ca2+transient, indicating that a decrease in myofilament responsiveness to Ca2+underlies the reduction in contractility. We assessed the role of PKC, tyrosine kinases, reactive oxygen species (ROS), and mitogen-activated protein kinases (MAPKs) in the NIE of the peptide. Pretreatment of cells with the NAD(P)H oxidase inhibitor diphenyleneiodonium chloride or with the superoxide scavenger 4,5-dihydroxy-1,3-benzene-disulfonic acid did not affect the ANG II-induced NIE. Moreover, ANG II-induced ROS production, after 20 min of incubation with the peptide, could not be detected with the use of either the fluorophore 5-(6)-chloromethyl-2′, 7′-dichlorodihydrofluorecein diacetate or lucigenin-enhanced chemiluminescence. In contrast, the ANG II-induced NIE was abrogated by the inhibitors of PKC (calphostin C), tyrosine kinase (genistein), and p38 MAPK (SB-202190). Furthermore, the NIE was significantly exacerbated (60 ± 10% reduction) by p38 MAPK overexpression. These results exclude the participation of ROS in the NIE of the peptide and point to PKC and tyrosine kinase as upstream mediators. Furthermore, they reveal p38 MAPK as the putative effector of the reduction in myofilament responsiveness to Ca2+and the decrease in contractility induced by the peptide.