The purpose of this study was to elucidate the mechanism by which bradykinin (BK) enhances prostacyclin (PGI2) production in human umbilical vein endothelial cells (HUVEC). BK-induced enhancement of PGI2 synthesis was observed in a dose- and time-dependent manner, and it also increased Ca2+i followed by enhancement of cytosolic phospholipase A2 (cPLA2) activity. The PKC inhibitors GF109203X and H7 attenuated the BK-induced increase in Ca2+i and inhibited the BK-induced PGI2 synthesis. Phorbol 12-myristate 13-acetate increased cPLA2 activity and PGI2 synthesis but failed to alter Ca2+i. BK increased cPLA2 mRNA eightfold by 15 min, and this increase was inhibited by pretreatment with the PKC inhibitors. In response to cycloheximide pretreatment, cPLA2 mRNA was superinduced. These results suggest that BK stimulates PGI2 synthesis in HUVEC by activation of cPLA2 by dual mechanisms: an elevation of Ca2+i and a PKC-dependent pathway. Moreover, changes in calcium kinetics and expression of cPLA2 mRNA may underlie the BK-induced PGI2 enhancement in these cells.