ROLE OF EICOSANOIDS IN VASOPRESSIN-INDUCED CALCIUM MOBILIZATION IN A7R5 VASCULAR SMOOTH-MUSCLE CELLS

The role of arachidonic acid (AA) and its metabolites in vasopressin (AVP)-induced calcium mobilization in A7r5 aortic smooth muscle cells was explored by intracellular calcium monitoring, [C-14]AA labeling, and high-performance liquid chromatography (HPLC) techniques. In fura 2-loaded A7r5 cells, AA potentiated AVP-stimulated increase in intracellular free Ca2+ ([Ca2+]i). The cyclooxygenase inhibitor indomethacin reduced both the AA- and AVP-induced influx of extracellular Ca2+. AVP-induced [Ca2+], transients were not altered by lipoxygenase inhibitors but were reduced in a dose-dependent fashion by ketoconazole, an inhibitor of cytochrome P-450 monooxygenases. Among several epoxygenase metabolites of AA tested, 5,6-epoxyeicosatrienoic acid potentiated AVP-induced [Ca2+]i transients. Reverse-phase HPLC analysis of lipid extracts from A7r5 cells prelabeled with [C-14]AA isolated a radioactive peak that did not coelute with established products of cyclooxygenase-, lipoxygenase-, or cytochrome P-450-catalyzed oxidations of AA. This peak was significantly increased after AVP stimulation and was completely blocked by preincubation with ketoconazole. Thus the stimulation of V1-vascular AVP receptors of A7r5 cells triggers several cytoplasmic signaling pathways involving AA metabolite formation through the cyclooxygenase and epoxygenase pathways.