HYDROGEN-PEROXIDE ACTIVATES AGONIST-SENSITIVE CA2-FLUX PATHWAYS IN CANINE VENOUS ENDOTHELIAL-CELLS()

The effect of the biological oxidant H2O2 on purinergic-receptor-stimu lated Ca2+ signalling was determined in canine venous endothelial cell s. H2O2 increased cytosolic free [Ca2+] ([Ca2+](i)), the rate of rise of which was dose-dependently related to H2O2 concentration. The respo nse of [Ca2+](i) to H2O2 resulted in part from release of Ca2+ from in ternal stores. The H2O2-sensitive intracellular Ca2+ pool was characte rized in cells suspended in Ca2+-free/EGTA buffer and stimulated in se quence with H2O2 and ionomycin or ATP. Under this condition, the rank order of apparent compartment size sensitive to each compound was iono mycin>H2O2>ATP. Stimulation of cells with H2O2 eliminated any response of [Ca2+](i) to subsequent addition of ATP. To test more directly whe ther H2O2 accesses the inositol trisphosphate-sensitive Ca2+ store, ce lls were pretreated with thapsigargin, a selective inhibitor of that s tore's Ca2+ pump. Release of Ca2+ from internal Ca2+ stores by H2O2 de clined as the interval after thapsigargin addition increased, a findin g that supports the contention that H2O2 accesses the inositol trispho sphate-sensitive Ca2+ store. H2O2-stimulated Ca2+ influx across the ce ll membrane was sensitive to Ni2+, La3+, and xyphenyl)propoxyl-4-metho xyphenethyl}-1H-imidazole HC1 (SKF-96365), a selective inhibitor of th e agonist stimulated Ca2+-influx pathway. Ca2+ entry triggered by H2O2 appears to occur via the agonist-sensitive Ca2+ influx pathway. Toget her, these results suggest that H2O2, is normally secreted by activate d neutrophils and monocytes, may act as an intercellular messenger and stimulate Ca2+ signalling in target endothelial cells.