Vol. 281, Issue 3, 1038-1046, 1997

Beta Adrenergic Receptor Stimulated Prostacyclin Synthesis in Rabbit Coronary Endothelial Cells Is Mediated by Selective Activation of Phospholipase D: Inhibition by Adenosine 35-Cyclic Monophosphate¹

Ying Ruan², Hong Kan³ and Kafait U. Malik

Department of Pharmacology, College of Medicine, The University of Tennessee, Memphis, Tennessee

Activation of beta adrenergic receptors in the isolated rabbit heart by catecholamines stimulates prostacyclin (PGI₂) synthesis, which is inhibited by adenosine 35-cyclic monophosphate (cAMP). The purpose of this study was to determine if activation of beta adrenergic receptors in cultured coronary endothelial cells (CEC) of rabbit heart with isoproterenol (ISOP) stimulates PGI₂ synthesis and if cAMP inhibits the synthesis of this prostanoid and to investigate the underlying mechanism. Incubation of CEC with ISOP increased production of cAMP and PGI₂, measured as immunoreactive cAMP and 6-keto-prostaglandin F₁, (6-keto-PGF₁), respectively. Forskolin, an activator of adenylyl cyclase, increased cAMP accumulation and inhibited ISOP-stimulated 6-keto-PGF₁ synthesis. 8-(4-chlorophenylthio) cAMP also inhibited ISOP-induced 6-keto-PGF₁ production. However, miconazole, an inhibitor of adenylyl cyclase, reduced cAMP accumulation and enhanced ISOP-stimulated 6-keto-PGF₁ synthesis in CEC. ISOP-induced 6-keto-PGF₁ synthesis was attenuated by C₂-ceramide, an inhibitor of phospholipase D (PLD) by propranolol, a beta-AR antagonist that also inhibits phosphatidate phosphohydrolase and by the diacylglycerol lipase inhibitor 1,6-bis-(cyclohexyloximinocarbonylamino)-hexane (RHC 80267). Acetylcholine (ACh) induced 6-keto-PGF₁ synthesis was also inhibited by these agents. Both ISOP and ACh increased PLD activity, which was inhibited by C₂-ceramide but not by RHC 80267 or propranolol. ACh but not ISOP increased phospholipase A₂ activity in CEC. ISOP- but not ACh-induced increase in PLD activity was attenuated by forskolin and 8-(4-chlorophenyl-thio)-adenosine 3-5-cyclic monophosphate and augmented by miconazole. These data suggest that beta adrenergic receptors activation promotes PGI₂ synthesis in the CEC by selective activation of PLD and that cAMP decreases PGI₂ synthesis by decreasing PLD activity. Moreover, beta adrenergic receptors activated PLD appears to be distinct from that stimulated by ACh.