nitroarginine and nickel-chloride

1. To investigate the mechanism underlying the trypsin-induced endothelium-dependent relaxation, cytosolic Ca(2+) concentration ([Ca(2+)](i)) and tension development of smooth muscle were simultaneously monitored in the porcine coronary artery, and [Ca(2+)](i) of in situ endothelial cells were monitored in the porcine aortic valvular strips, using fura-2 fluorometry. 2. During the contraction induced by 30 nM U46619, a thromboxane A(2) analogue, 100 nM trypsin induced a rapid transient significant decrease in both [Ca(2+)](i) (from 67.9+/-5.1 to 15.7+/-4.4%) and tension (from 97.5+/-9.2 to 16.8+/-3.5%) of smooth muscle only in the presence of endothelium (100% level was assigned to the level obtained with the 118 mM K(+)-induced contraction). [Ca(2+)](i) and the tension thus returned to the levels prior to the application of trypsin by 5 and 10 min, respectively. 3. The initial phase of this relaxation was partly inhibited by 100 microM N(omega)-nitro-L-arginine (L-NOARG), and was completely inhibited by L-NOARG plus 40 mM K(+) or L-NOARG plus 100 nM charybdotoxin and 100 nM apamin, while the late phase of the relaxation was inhibited by L-NOARG alone. 4. Trypsin induced a transient [Ca(2+)](i) elevation in the endothelial cells mainly due to the Ca(2+) release from the intracellular stores, at the concentrations (1 - 100 nM) similar to those required to induce relaxation. 5. In conclusion, trypsin induced an elevation in [Ca(2+)](i) mainly due to Ca(2+) release in endothelial cells, and thereby caused endothelium-dependent relaxation. The early phase of relaxation was due to nitric oxide and hyperpolarizing factors, while the late phase was mainly due to nitric oxide in the porcine coronary artery.

Topics: 15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; Adenosine Triphosphate; Animals; Aortic Valve; Apamin; Biological Factors; Calcium; Charybdotoxin; Coronary Vessels; Dose-Response Relationship, Drug; Endothelium, Vascular; Enzyme Inhibitors; In Vitro Techniques; Indomethacin; Muscle, Smooth, Vascular; Nickel; Nitric Oxide; Nitric Oxide Synthase; Nitroarginine; Oligopeptides; Potassium Channel Blockers; Receptor, PAR-2; Receptors, Thrombin; Substance P; Swine; Time Factors; Trypsin; Vasoconstrictor Agents; Vasodilation

The glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is induced by hypoxia in endothelial cells (EC). To define the mechanisms by which GAPDH is regulated by hypoxia, EC were exposed to cobalt, other transition metals, carbon monoxide (CO), deferoxamine, or cycloheximide in the presence or absence of hypoxia for 24 h, and GAPDH protein and mRNA levels were measured. GAPDH was induced in cells by the transition metals cobalt, nickel, and manganese and by deferoxamine, and GAPDH mRNA induction by hypoxia was blocked by cycloheximide. GAPDH induction by hypoxia, unlike that of other hypoxia-regulated genes, was not inhibited by CO or by 4,6-dioxoheptanoic acid, an inhibitor of heme synthesis. GAPDH induction was not altered by mediators of protein phosphorylation, a calcium channel blocker, a calcium ionophore, or alterations in redox state. GAPDH induction by hypoxia or transitional metals was partially blocked by sodium nitroprusside but was not altered by the inhibitor of nitric oxide synthase N omega-nitro-L-arginine. These findings suggest that GAPDH induction by hypoxia in EC occurs via mechanisms other than those involved in other hypoxia-responsive systems.

Topics: Animals; Antidotes; Calcium; Cattle; Cell Hypoxia; Cells, Cultured; Chlorides; Cobalt; Deferoxamine; Endothelium, Vascular; Enzyme Induction; Enzyme Inhibitors; Ferrous Compounds; Gene Expression Regulation, Enzymologic; Glyceraldehyde-3-Phosphate Dehydrogenases; Heptanoates; Manganese Compounds; Nickel; Nitric Oxide; Nitroarginine; Nitroprusside; Oxidation-Reduction; RNA, Messenger; Up-Regulation; Zinc Compounds