Effects of Endothelin-1 on Ca2+Signaling in Guinea-Pig Ventricular Myocytes: Role of Protein Kinase C

The effects of ET-1 on contraction, Ca2+transient and -type Ca2+current (ICa,L) were studied in single cells isolated from ventricles of guinea-pig hearts. The aim of our study was to elucidate the mechanism of the positive inotropic effect during endothelin receptor stimulation by focusing on the role of PKC. ET-1 at concentrations of 5 and 10 n produced a biphasic pattern of inotropism: a first decrease in contraction by 34.4±2.5% of the control followed by a sustained increase in contraction by 66.6±8.4% (mean±SEM,n=9). The Ca2+transient decreased by 13.5±1.0% during the negative inotropic phase, while it increased by 58.1±8.4% (n=10) during the positive inotropic phase. Using the whole-cell voltage-clamp technique with conventional microelectrodes, the application of ET-1 (5 n ) increased theICa,Lby 32.6±5.1% (n=10), which was preceded by a short-lived decrease inICa,L. Incubation of myocytes with pertussis toxin (PTX, at 2μg/ml for >3 h at 35°C) failed to block the ET-1-induced enhancement ofICa,L. The increases in contraction, Ca2+transient, andICa,Lby ET-1 were inhibited by pretreatment with 5-N-methyl-N-isobutyl amiloride (MIA; 10μm), an amiloride analog, and a novel selective Na+/H+exchange inhibitor HOE694 (10μm). To determine whether activation of protein kinase C (PKC) is responsible for the enhancement ofICa,Lby ET-1, we tested a PKC inhibitor, GF109203X, and found that it does exert an inhibitory effect on the ET-1-inducedICa,Lincrease. Our study suggests that during ET receptor stimulation an increase inICa,Ldue to stimulation of Na+/H+exchange via PKC activation causes an increase in Ca2+transients and thereby in the contractile force of the ventricular myocytes.