ENDOTHELIN AND INCREASED CONTRACTILITY IN ADULT-RAT VENTRICULAR MYOCYTES - ROLE OF INTRACELLULAR ALKALOSIS INDUCED BY ACTIVATION OF THE PROTEIN KINASE-C-DEPENDENT NA+-H+ EXCHANGER

Endothelin, a 21-amino acid vasoactive peptide, is among the most potent positively inotropic agents yet described in mammalian heart. Having demonstrated that endothelin's inotropic effect is due, in part, to an apparent sensitization of cardiac myofilaments to intracellular calcium, we determined whether this could be due to a rise in intracellular pH (pH(i)). In isolated adult rat ventricular cells loaded with the H+-selective fluorescent probe BCECF, 100 pM endothelin increased contractile amplitude to 190+/-26% of baseline and pH(i) by 0.08+/-0.02 (n = 8), whereas 1 nM endothelin increased pH(i) by 0.13+/-0.03 with little further increase in contractility. Amiloride (10(-4) M) prevented the increase in pH(i) in response to endothelin and reduced the inotropic response by 45%, although the inotropic effect could be readily restored by subsequent NH4Cl-induced alkalinization. Similarly, inhibitors of protein kinase C (H-7 and sphingosine) diminished or abolished the rise in pH(i) after endothelin superfusion while causing a decline in its inotropic effect comparable with that observed with amiloride. Pretreatment with pertussis toxin, which we have demonstrated results in complete ADP-ribosylation of the alpha-subunits of G(o) and G(i) GTP-binding proteins and abolition of endothelin's positive inotropic effect, only partially reduced the intracellular alkalinization induced by the peptide, suggesting a complex signal transduction mechanism. Thus, the positive inotropic action of endothelin is due in part to stimulation of the sarcolemmal Na+-H+ exchanger by a protein kinase C-mediated pathway, resulting in a rise pH(i) and sensitization of cardiac myofilaments to intracellular Ca2+.