MULTIPLE MECHANISMS OF ACTIVATING CA2-CELLS( ENTRY IN FRESHLY ISOLATED RABBIT AORTIC ENDOTHELIAL)

In Fura-2-loaded, freshly isolated rabbit aortic endothelial cells the Ca2+ entry pathway was investigated using the Mn2+-quenching techniqu e. Acetylcholine (ACh) interaction with muscarinic receptors activated Mn2+ influx through the plasma membrane. Sarcoplasmic-endoplasmic ret iculum Ca2+ ATPase blockers such as cyclopiazonic acid (CPA), thapsiga rgin and BHQ, which block the endoplasmic reticulum Ca2+ pump and do n ot interact with receptors, also activated Mn2+ influx. Mn2+ influx ac tivated by either ACh or CPA was blocked by the following agents: SKF9 6365, a receptor-operated Ca2+ channel (ROC) blocker, NCDC, a PLC and ROC blocker, and genistein, a tyrosine kinase inhibitor. D600, the L-t ype Ca2+ channel blocker, had no significant effect on Mn2+ influx. Ca ffeine blocked the ACh-induced Ca2+ release but had no effect on the A Ch-induced Mn2+ influx. Similarly dantrolene, which blocked intracellu lar Ca2+ release induced by ACh, did not affect the ACh-activated Mn2 influx. These data suggest that ACh can activate Ca2+ influx without depletion of the ACh-sensitive intracellular Ca2+ store, It is conclud ed (1) that in freshly isolated endothelial cells depletion of the int racellular Ca2+ store is not necessary for ACh-activated Ca2+ influx, and (2) that receptor activation and intracellular Ca2+ store depletio n may activate the same Ca2+ entry pathway through parallel mechanisms .