Bradykinin stimulates phospholipase D in PC12 cells by a mechanism which is independent of increases in intracellular Ca2

These experiments were designed to learn the role of bradykinin induced changes in intracellular Ca2+ in the activation of phospholipase D activity in PC12 cells. Ionomycin at a concentration of 0.1 microM caused an increase in intracellular Ca2+ comparable to bradykinin, but had no effect on phospholipase D activity. Carbachol, ATP, and thapsigargin also increased intracellular Ca2+ but had no effect on phospholipase D activity. Increases in intracellular Ca2+ may be a necessary but not a sufficient factor in the activation of phospholipase D. To investigate this issue, the bradykinin induced increase in intracellular Ca2+ was blocked by preincubating the cells in Ca(2+)-free media plus EGTA or in media containing the intracellular Ca2+ chelator BAPTA/AM. These preincubations completely blocked the bradykinin induced increase in intracellular Ca2+ but only attenuated the bradykinin mediated activation of phospholipase D. Physiological increases in intracellular Ca2+ apparently do not mediate the effect of bradykinin on phospholipase D.