PHOSPHOLIPASE-D IN RAT MYOCARDIUM - FORMATION OF LIPID MESSENGERS ANDSYNERGISTIC ACTIVATION BY G-PROTEIN AND PROTEIN-KINASE-C

Activation of phospholipase D (PLD) and phosphoinositide-specific phos pholipase C (PI-PLC) by fluoride, to stimulate heterotrimeric G-protei ns, and by phorbol esters, to stimulate protein kinase C (PKC), was st udied in rat atria. Fluoride and 4 beta-phorbol-12 beta,13 alpha-dibut yrate (PDB), in contrast to 4 beta-phorbol-13 alpha-acetate (PAc), act ivated PLD, catalyzing the formation of [H-3]-phosphatidylethanol ([H- 3]-PETH), [H-3]-phosphatidic acid ([H-3]-PA), choline and sn-1,2-diacy lglycerol (DAG). Basal PLD activity was resistant to drastic changes i n Ca2+ and to Ro 31-8220, a PKC inhibitor, but was decreased by genist ein, an inhibitor of tyrosine kinase, and increased by vanadate, a tyr osine phosphatase inhibitor; both effects were, however, very small. F luoride-evoked PLD activity was resistant to Po 31-8220 and to geniste in, but was Ca2+-dependent. The rate of fluoride induced PLD activatio n was maintained for at least 60 min. In contrast, PDB-mediated PLD ac tivity was blocked by Ro 31-8220 and was resistant to extracellular Ca 2+-depletion and desensitized within ca. 15 min. PDB markedly potentia ted the fluoride evoked generation of [H-3]-phosphatidylethanol and of choline, but inhibited the formation of [H-3]-inositol phosphates ([H -3]-IP1-3). Ethanol (2%) blocked the PDB-evoked generation of both [H- 3]-phosphatidic acid and of sn-l,2-diacylglycerol, whereas fluoride-ev oked responses were reduced only to approximately 50%. In conclusion, the trimeric G-protein-PLD pathway in heart tissue did not enclose PKC activation and was long-lasting and Ca2+-dependent; there was no evid ence for an involvement of tyrosine phosphorylation. However, PKC acti vation modulated G-protein-coupled PLD and PI-PLC activities in opposi te directions. PLD activity significantly contributed to the mass prod uction of sn-1,2-diacylglycerol in the heart. The evidence for a patho physiological role of PLD activation in cardiac hypertrophy and in isc hemic preconditioning is discussed. BIOCHEM PHARMACOL 56;7:799-805, 19 98. (C) 1998 Elsevier Science Inc.