Modulation of agonist-induced calcium mobilisation in bovine aortic endothelial cells by phorbol myristate acetate and cyclic AMP but not cyclic GMP.
1. In bovine aortic endothelial cells (BAEC), thrombin (1 mu ml-1), bradykinin (1-10 nM) and adenosine triphosphate (ATP) (0.3 microM-100 microM) each induced a biphasic elevation of cytosolic calcium ([Ca2+]i), consisting of an initial transient followed by a sustained plateau phase. 2. Pretreatment of BAEC with 4 beta-phorbol 12-myristate 13-acetate (PMA; 100 nM) reduced the magnitude of the initial transient elevation of [Ca2+]i, induced by thrombin (1 mu ml-1), low concentrations of bradykinin (1 nM) or ATP (0.3 microM, 3 microM), but not by higher concentrations of the latter two agonists. Addition of PMA (100 nM) during the plateau phase of the increase in [Ca2+]i induced by thrombin (1 mu ml-1), bradykinin (10 nM) or ATP (30 microM) resulted in a fall in [Ca2+]i. 3. The inhibitory effects of PMA (100 nM) were inhibited by staurosporine (100 nM) but not mimicked by the inactive phorbol ester, 4 alpha-phorbol 12,13-didecanoate (4 alpha-PDD; 100 nM). Furthermore, staurosporine (100 nM) increased [Ca2+]i when added during the plateau phase of the increase in [Ca2+]i induced by thrombin or bradykinin. In contrast, staurosporine (100 nM) reduced [Ca2+]i when added during the plateau phase of the increase in [Ca2+]i induced by ATP (30 microM). 4. Pretreatment with forskolin (10 microM) had no effect on the magnitude of the initial transient elevation of [Ca2+]i induced by thrombin (1 mu ml-1), bradykinin (1 nM and 10 nM) or ATP (30 microM). In contrast, forskolin (10 microM) and isoprenaline (10 microM) each induced biphasic elevations of [Ca21]i when added during the plateau phase of the increase in [Ca2+]i induced by the three agonists. Furthermore, in the presence of the inhibitor of calcium influx, nickel chloride (4mM), these biphasic elevations were reduced to monophasic transient elevations. 5. 8 Bromo cyclic GMP (30 microM), a membrane-permeant analogue of guanosine 3': 5'-cyclic monophosphate (cyclic GMP), had no effect on the magnitude of the initial transient elevation of [Ca21]i induced by thrombin (1 u ml 1), bradykinin (10 nM) or ATP (3 microM). Furthermore, 8 bromo cyclic GMP (30 microM) and sodium nitroprusside (1 microM), had no effect when added during the plateau phase of the increase in [Ca2+]i induced by the three agonists. 6. NG nitro-L-arginine (50,microM), an inhibitor of nitric oxide synthase, had no effect on the magnitude of the initial transient elevation of [Ca21]i induced by thrombin (1 uml- ), bradykinin (1 nM) or ATP (3,microM), and had no effect on the plateau phase of the increase in [Ca2+]i induced by these agents. 7. These findings suggest that while activation of protein kinase C inhibits and elevation of adenosine 3': 5'-cyclic monophosphate (cyclic AMP) augments calcium mobilisation in bovine aortic endothelial cells, elevation of cyclic GMP appears to have no effect.
Buchan
K W
KW
Department of Pharmacology, University of Glasgow.
Martin
W
W
D000818:Animals; D001011:Aorta / Q000187:drug effects / Q000378:metabolism; D001120:Arginine / Q000031:analogs & derivatives / Q000494:pharmacology; D001920:Bradykinin / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D002120:Calcium Channel Agonists / Q000494:pharmacology*; D002417:Cattle; D002478:Cells, Cultured; D005576:Colforsin / Q000494:pharmacology; D000242:Cyclic AMP / Q000494:pharmacology*; D006152:Cyclic GMP / Q000031:analogs & derivatives / Q000494:pharmacology*; D004730:Endothelium, Vascular / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D007545:Isoproterenol / Q000494:pharmacology; D019335:Nitroarginine; D009599:Nitroprusside / Q000494:pharmacology; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*; D013917:Thrombin / Q000494:pharmacology
D002120:Calcium Channel Agonists; D009599:Nitroprusside; D005576:Colforsin; D019335:Nitroarginine; C016276:8-bromocyclic GMP; D001120:Arginine; D000242:Cyclic AMP; D013917:Thrombin; D006152:Cyclic GMP; D007545:Isoproterenol; D013755:Tetradecanoylphorbol Acetate; D001920:Bradykinin; D002118:Calcium