P2 purinoceptor-stimulated conversion of arginine to citrulline in bovine endothelial cells is reduced by inhibition of protein kinase C.
Bovine aortic endothelial cells contain two coexisting receptors for extracellular ATP, named the P2Y and P2U purinoceptors. Previous studies have shown that these receptors are linked to phospholipase C in a manner that is modulated in part by protein kinase C (PKC). In this study, we investigate the influence of PKC in the regulation of endothelial nitric oxide synthase (NOS) by these two purinoceptors. Activation of either P2Y or P2U purinoceptors by either 2-methylthio-ATP or UTP, respectively, stimulated the formation of [3H]-citrulline in [3H]-arginine-labelled cells in a concentration-dependent manner. This stimulation was sensitive to inhibition by NG-nitro-L-arginine. Ten minutes of pretreatment with the PKC activator tetradecanoyl phorbol acetate (TPA) failed to affect NOS activity, either alone or when stimulated with 2-methylthio-ATP or UTP. However, under these conditions TPA caused almost complete translocation of PKC-alpha from the cytosol to the membrane. Ten minutes of pretreatment with the PKC inhibitor Ro 31-8220 significantly inhibited the agonist-induced stimulation of NOS. These results show that both P2Y and P2U purinoceptors stimulate endothelial NOS in a manner that is dependent on PKC activity.
Brown
C A
CA
Department of Cell Physiology and Pharmacology, University of Leicester, UK.
Patel
V
V
Wilkinson
G
G
Boarder
M R
MR
D000818:Animals; D001120:Arginine / Q000378:metabolism*; D002417:Cattle; D002478:Cells, Cultured; D002956:Citrulline / Q000378:metabolism*; D004730:Endothelium, Vascular / Q000378:metabolism*; D009569:Nitric Oxide / Q000378:metabolism; D019001:Nitric Oxide Synthase / Q000378:metabolism; D011493:Protein Kinase C / Q000037:antagonists & inhibitors*; D018048:Receptors, Purinergic P2 / Q000502:physiology*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D018048:Receptors, Purinergic P2; D002956:Citrulline; D009569:Nitric Oxide; D001120:Arginine; D019001:Nitric Oxide Synthase; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate