Mechanism of endothelin-1-induced cytosolic Ca2+ mobility in cultured H9c2 myocardiac ventricular cells

The effect of endothelin-1 (ET-1) on the intracellular free Ca2+ ([Ca2+]i) mobility in cultured H9c2 myocardiac ventricular cells was studied after loading with fura-2-AM. In Ca2+-containing buffer, ET-1 induced [Ca2+]i rise from 10−7 to 10−9 M. ET-1 induced [Ca2+]i, which was composed of a first small peak and a secondary persistent plateau. In Ca2+-free buffer, pretreatment with 10−7 M ET-1 inhibited the thapsigargin and carbonylcyanide m-chlorophenylhydrazone (CCCP)-induced [Ca2+]i increase. Meanwhile, pretreatment with thapsigargin and CCCP also inhibited ET-1-induced [Ca2+]i rise. In Ca2+-containing buffer, the ETA receptor antagonist (BQ123) completely abolished the secondary rising peak and plateau. Conversely, the ETB receptor antagonist (BQ788) completely inhibited the first small peak and secondary peak plateau. Nifedipine and La3+ also abolished the 10−7 M ET-1-induced [Ca2+]i in the first rising peak. The internal Ca2+ release induced by ET-1 was inhibited by U73122 (phospholipase C inhibitor), propranolol (phospholipase D inhibitor) and aristolochic acid (phospholipase A2 inhibitor). After incubation of 10−7 M ET-1 in Ca2+-free buffer, the addition of 5 mM CaCl2 increased Ca2+ influx, implying that release of Ca2+ from internal stores further induces capacitative Ca2+ entry. Taken together, these results suggest that both ETA and ETB receptors are involved in ET-1-induced [Ca2+]i rise in H9c2 myocardiac ventricular cells. Whereas ETB receptor seems to mediate the initial Ca2+ influx via L-type Ca2+ channel, ETA receptor appears to be involved in the subsequent Ca2+ release from endoplasmic reticulum and mitochondria Ca2+ stores.