The inhibition of the constitutive bovine endothelial nitric oxide synthase by imidazole and indazole agents.
Citrulline formation by the Ca2+ CaM-dependent nitric oxide synthase of bovine endothelium is inhibited reversibly by 7-nitroindazole, 1-phenylimidazole, and imidazole. As measured at 0.67 microM (6R)-5,6,7,8-tetrahydrobiopterin (BH4), IC50 values of 0.8, 200, and 50 microM were determined for 7-nitroindazole, 1-phenylimidazole, and imidazole, respectively. Increasing concentrations of added BH4 cofactor increased the IC50 values for 7-nitroindazole and 1-phenylimidazole but did not alter the IC50 value for imidazole. 7-nitroindazole inhibited citrulline formation by the endothelial cNOS noncompetitively versus arginine substrate but competitively versus BH4 with a Ki value of 0.8 microM. 1-Phenylimidazole inhibited citrulline formation by the endothelial cNOS competitively versus both arginine substrate and BH4 with a Ki value of 50 microM. Imidazole inhibited citrulline formation competitively versus arginine substrate but noncompetitively versus BH4 with a Ki value of 50 microM. Neither 7-nitroindazole, 1-phenylimidazole, nor imidazole inhibited the cytochrome c reductase activity of endothelial cNOS at concentrations up to 5000-fold higher than their Ki values for inhibition of citrulline formation. By comparison with the previously determined kinetic properties of the other nitric oxide synthase isoforms, these observations establish that 1-phenylimidazole displays marked specificity for inhibiting the inducible nitric oxide synthase isoform and, since 7-nitroindazole has been reported not to elevate blood pressure (McCall et al., 1991, Br. J. Pharmacol. 102, 234-238), fails to confirm the expected insensitivity of the constitutive endothelial nitric oxide synthase to inhibition by 7-nitroindazole.
Wolff
D J
DJ
Department of Pharmacology, University of Medicine and Dentistry of New Jersey, Robert Wood Johnson Medical School, Piscataway 08854.
Lubeskie
A
A
Umansky
S
S
D000594:Amino Acid Oxidoreductases / Q000037:antagonists & inhibitors* / Q000302:isolation & purification; D000818:Animals; D001921:Brain / Q000201:enzymology; D002417:Cattle; D002956:Citrulline / Q000032:analysis; D004730:Endothelium, Vascular / Q000201:enzymology*; D007093:Imidazoles / Q000494:pharmacology*; D007191:Indazoles / Q000494:pharmacology; D007527:Isoenzymes / Q000037:antagonists & inhibitors; D007700:Kinetics; D008264:Macrophages / Q000201:enzymology; D019001:Nitric Oxide Synthase; D011651:Pulmonary Artery
D007093:Imidazoles; D007191:Indazoles; D007527:Isoenzymes; D002956:Citrulline; C037738:1-phenylimidazole; C029899:imidazole; D019001:Nitric Oxide Synthase; D000594:Amino Acid Oxidoreductases; C080122:7-nitroindazole