REGULATION OF PHOSPHOLIPASE-C AND PHOSPHOLIPASE-D IN RAT VENTRICULAR MYOCYTES - STIMULATION BY ENDOTHELIN-1, BRADYKININ AND PHENYLEPHRINE

The physiological activator of protein kinase C (PKC), diacylglycerol, is formed by hydrolysis of phosphoinositides (PI) by phospholipase C (PLC) or phosphatidylcholine by phospholipase D (PLD). We have measure d activation of these phospholipases by endothelin-1 (ET-1), bradykini n (BK), or phenylephrine (PE) in ventricular myocytes cultured from ne onatal rat. The stimulation of PI hydrolysis after 10 min by 0.1 mu M ET-1 (about 12-fold) was much greater than for BK or PE (each about fo ur-fold), and did not correlate with translocation of nPKC delta or nP KC epsilon (Clerk A, Bogoyevitch MA, Andersson MB, Sugden PH, 1994. J Biol Chem 269: 32848-32857; Clerk A, Gillespie-Brown J, Fuller SJ, Sug den PH, 1996. Biochem J 317: 109-118). However, ET-1 and BK stimulated a similar rapid increase in [H-3]InsP(3) formation (<30 s), which was much greater than that seen with PE. This early phase correlated with PKC translocation. Acute or chronic exposure to 12-O-tetradecanoylpho rbol-13-acetate (TPA) or treatment with Ro-31-8220 showed that the sti mulation of PI hydrolysis by PE, but not ET-1 or BK, was inhibited by activation of PKC. Furthermore, ET-1 and BK heterologously desensitize d the stimulation of PI hydrolysis by PE. ET-1 or BK homologously unco upled their own receptors from [H-3]InsP(3) formation, but there was n o evidence of heterologous desensitization with these two agonists. An omalously, chronic exposure to TPA increased the stimulation of PI hyd rolysis by BK, but this probably resulted from an increase in BK recep tor density. PLD was also rapidly activated by TPA, ET-1, BK or PE. Ex periments with Ro-31-8220 showed that the stimulation of PLD by ET-1 a nd BK was mediated through activation of PKC. We discuss the character istics of the activation of PI hydrolysis and PLD by ET-1, BK, and PE with respect to the translocation of PKC. (C) 1997 Academic Press Limi ted.