Inhibition of nitrovasodilator- and acetylcholine-induced relaxation and cyclic GMP accumulation by the cytochrome P-450 substrate, 7-ethoxyresorufin

We examined the effect of the cytochrome P-450 substrate, 7-ethoxyresorufin (7-ER), and its corresponding product, resorufin, on nitrovasodilator- and endothelium-dependent relaxation of isolated rat aorta. The EC50 value for glyceryl trinitrate (GTN) induced relaxation was increased over 100-fold by 7-ER and less than 3-fold by resorufin. The EC50 value for sodium nitroprusside (SNP) induced relaxation was increased approximately 12-fold by 7-ER, acetylcholine (ACh) induced relaxation was abolished, and relaxation induced by isopropylnorepinephrine was not significantly affected. GTN-, SNP-, and ACh-induced increases in cyclic GMP accumulation were inhibited by 7-ER, as were basal cyclic GMP levels in endothelium-intact, but not endothelium-denuded tissues. 7-ER decreased GTN biotransformation in intact aorta and decreased the regioselective formation of glyceryl-1,2-dinitrate. The activation by GTN and SNP of aortic guanylyl cyclase in broken cell preparations was not affected by 7-ER, indicating that the inhibitory effect of 7-ER is probably not due to a direct interaction with guanylyl cyclase. The inhibitory effect of 7-ER on GTN-induced relaxation was not altered by the addition of superoxide dismutase, suggesting that 7-ER does not act by increasing superoxide anion concentration (which would serve to increase the degradation of nitric oxide (NO) formed during vascular GTN biotransformation). Our data provide further evidence for the role of the cytochrome P-450 – cytochrome P-450 reductase system in the biotransformation of GTN to an activator (presumably nitric oxide) of guanylyl cyclase. The data are consistent with a mode of action of 7-ER involving either competitive inhibition of vascular cytochrome P-450 or uncoupling of vascular cytochrome P-450 reductase from cytochrome P-450. The data also suggest that the cytochrome P-450 system facilitates NO release from SNP and that 7-ER has an inhibitory effect on endothelial nitric oxide synthase.Key words: glyceryl trinitrate, nitrovasodilators, cytochrome P-450, vascular smooth muscle, 7-ethoxyresorufin, endothelium, cyclic GMP.

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Effect of inhibitors of glutathione S-transferase on glyceryl trinitrate activity in isolated rat aorta

Canadian Journal of Physiology and Pharmacology ◽

10.1139/y93-025 ◽

1993 ◽

Vol 71 (2) ◽

pp. 179-184 ◽

Cited By ~ 19

Author(s):

Rita Nigam ◽

Tracy Whiting ◽

Brian M. Bennett

Keyword(s):

Glyceryl Trinitrate ◽

Cyclic Gmp ◽

Guanylyl Cyclase ◽

Rat Aorta ◽

Supernatant Fraction ◽

Organic Nitrates ◽

Transferase Activity ◽

Glutathione S Transferase ◽

Glutathione S Transferases

We investigated the role of glutathione S-transferases (enzymes known to biotransform organic nitrates) in the vascular action of glyceryl trinitrate (GTN). Relaxation of phenylephrine-contracted rat aortic strips was assessed in the presence or absence of the glutathione S-transferase inhibitors Basilen Blue, bromosulfophthalein, Rose Bengal, hematin, chlorotriphenyltin, and (octyloxy)benzoylvinylglutathione. Whereas none of the inhibitors increased the EC50 for GTN relaxation, glutathione S-transferase activity in the 100 000 × g supernatant fraction of rat aorta was inhibited markedly by most of the inhibitors. In addition, GTN-stimulated activation of aortic guanylyl cyclase in broken-cell preparations was attenuated by all of the glutathione S-transferase inhibitors, suggesting a direct inhibitory action on guanylyl cyclase. In other experiments using aortic strips preexposed to phenylephrine, the inhibitors had no effect on GTN-induced cyclic GMP accumulation or on vascular biotransformation of GTN. In contrast, both Basilen Blue and bromosulfophthalein significantly inhibited GTN-induced relaxation of K+-contracted aortic strips, and Basilen Blue significantly inhibited GTN biotransformation in aortic strips preexposed to 25 mM K+. This may be due to a more favourable electrochemical gradient for entry of the inhibitors into membrane-depolarized tissues. We conclude that vascular glutathione S-transferases play a role in mediating the vasodilator actions of GTN in intact tissues in vitro, but that this appears to depend upon the nature of the contractile agent used in such studies.Key words: glyceryl trinitrate, glutathione S-transferase, cyclic GMP, vascular smooth muscle, biotransformation.

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Thiopental inhibits nitric oxide production in rat aorta

Canadian Journal of Physiology and Pharmacology ◽

10.1139/y99-103 ◽

1999 ◽

Vol 77 (12) ◽

pp. 958-966 ◽

Cited By ~ 11

Author(s):

Carlos Castillo ◽

Juan Asbun ◽

Bruno Escalante ◽

Carlos M Villalón ◽

Pedro López ◽

...

Keyword(s):

Nitric Oxide ◽

Inhibitory Effect ◽

Rat Aorta ◽

Nitric Oxide Production ◽

Nitric Oxide Synthesis ◽

Oxide Production ◽

Nitrite Production ◽

Oxide Synthase ◽

Endothelial Nitric Oxide ◽

Endothelium Dependent Relaxation

We studied whether thiopental affects endothelial nitric oxide dependent vasodilator responses and nitrite production (an indicator of nitric oxide production) elicited by acetylcholine, histamine, and A23187 in rat aorta (artery in which nitric oxide is the main endothelial relaxant factor). In addition, we evaluated the barbiturate effect on nitric oxide synthase (NOS) activity in both rat aorta and kidney homogenates. Thiopental (10-100 µg/mL) reversibly inhibited the endothelium-dependent relaxation elicited by acetylcholine, histamine, and A23187. On the contrary, this anesthetic did not modify the endothelium-independent but cGMP-dependent relaxation elicited by sodium nitroprusside (1 nM - 1 µM) and nitroglycerin (1 nM - 1 µM), thus excluding an effect of thiopental on guanylate cyclase of vascular smooth muscle. Thiopental (100 µg/mL) inhibited both basal (87.8 ± 14.3%) and acetylcholine- or A23187-stimulated (78.6 ± 3.9 and 39.7 ± 5.6%, respectively) production of nitrites in aortic rings. In addition the barbiturate inhibited (100 µg/mL) the NOS (45 ± 4 and 42.8 ± 9%) in aortic and kidney homogenates, respectively (measured as 14C-labeled citrulline production). In conclusion, thiopental inhibition of endothelium-dependent relaxation and nitrite production in aortic rings strongly suggests an inhibitory effect on NOS. Thiopental inhibition of the NOS provides further support to this contention.Key words: thiopental, rat aorta, endothelium-dependent relaxation, nitric oxide synthesis.

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Direct interaction between the reductase domain of endothelial nitric oxide synthase and the ryanodine receptor

FEBS Letters ◽

10.1016/j.febslet.2005.04.078 ◽

2005 ◽

Vol 579 (14) ◽

pp. 3159-3163 ◽

Cited By ~ 13

Author(s):

Mónica Martínez-Moreno ◽

Alberto Álvarez-Barrientos ◽

Fernando Roncal ◽

Juan Pablo Albar ◽

Francisco Gavilanes ◽

...

Keyword(s):

Nitric Oxide ◽

Nitric Oxide Synthase ◽

Ryanodine Receptor ◽

Endothelial Nitric Oxide Synthase ◽

Direct Interaction ◽

Oxide Synthase ◽

Endothelial Nitric Oxide ◽

Reductase Domain

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Upregulation of endothelial nitric oxide synthase in rat aorta after ingestion of fish oil-rich diet

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.01145.2003 ◽

2004 ◽

Vol 287 (2) ◽

pp. H567-H572 ◽

Cited By ~ 34

Author(s):

Diego López ◽

Xavier Orta ◽

Kelly Casós ◽

M. Puy Sáiz ◽

Pere Puig-Parellada ◽

...

Keyword(s):

Nitric Oxide ◽

Nitric Oxide Synthase ◽

Fish Oil ◽

Endothelial Nitric Oxide Synthase ◽

Rat Aorta ◽

Oxide Synthase ◽

Endothelial Nitric Oxide

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EDHF mediates flow-induced dilation in skeletal muscle arterioles of female eNOS-KO mice

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.2001.280.6.h2462 ◽

2001 ◽

Vol 280 (6) ◽

pp. H2462-H2469 ◽

Cited By ~ 93

Author(s):

An Huang ◽

Dong Sun ◽

Mairead A. Carroll ◽

Houli Jiang ◽

Carolyn J. Smith ◽

...

Keyword(s):

Nitric Oxide ◽

Skeletal Muscle ◽

Endothelial Nitric Oxide Synthase ◽

Hexanoic Acid ◽

Gracilis Muscle ◽

Wild Type ◽

Basal Release ◽

Oxide Synthase ◽

Endothelial Nitric Oxide ◽

Cytochrome P 450

Vasodilation to increases in flow was studied in isolated gracilis muscle arterioles of female endothelial nitric oxide synthase (eNOS)-knockout (KO) and female wild-type (WT) mice. Dilation to flow (0–10 μl/min) was similar in the two groups, yet calculated wall shear stress was significantly greater in arterioles of eNOS-KO than in arterioles of WT mice. Indomethacin, which inhibited flow-induced dilation in vessels of WT mice by ∼40%, did not affect the responses of eNOS-KO mice, whereas miconazole and 6-(2-proparglyoxyphenyl)hexanoic acid (PPOH) abolished the responses. Basal release of epoxyeicosatrienonic acids from arterioles was inhibited by PPOH. Iberiotoxin eliminated flow-induced dilation in arterioles of eNOS-KO mice but had no effect on arterioles of WT mice. In WT mice, neither N ω-nitro-l-arginine methyl ester nor miconazole alone affected flow-induced dilation. Combination of both inhibitors inhibited the responses by ∼50%. 1 H-[1,2,4]oxadiazolo[4,3- a]quinoxalin-1-one (ODQ) alone inhibited flow-induced dilation by ∼49%. ODQ + indomethacin eliminated the responses. Thus, in arterioles of female WT mice, nitric oxide and prostaglandins mediate flow-induced dilation. When eNOS is inhibited, endothelium-derived hyperpolarizing factor substitutes for nitric oxide. In female eNOS-KO mice, metabolites of cytochrome P-450, via activation of large-conductance Ca2+-activated K+ channels of smooth muscle, mediate entirely the arteriolar dilation to flow.

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Cytochrome P-450 mediated biotransformation of organic nitrates

Canadian Journal of Physiology and Pharmacology ◽

10.1139/y90-236 ◽

1990 ◽

Vol 68 (12) ◽

pp. 1552-1557 ◽

Cited By ~ 70

Author(s):

Bernard J. McDonald ◽

Brian M. Bennett

Keyword(s):

Glyceryl Trinitrate ◽

Microsomal Fraction ◽

Direct Interaction ◽

Organic Nitrate ◽

Organic Nitrates ◽

Aerobic Conditions ◽

First Order ◽

Preferential Formation ◽

First Order Kinetics ◽

Cytochrome P 450

The vascular biotransformation of organic nitrates appears to be a prerequisite for their action as vasodilators. In the current study, we assessed the involvement of cytochrome P-450 in the denitration of glyceryl trinitrate and the enantiomers of isoidide dinitrate. Denitration of organic nitrates by the microsomal fraction of rat liver was NADPH dependent and followed apparent first-order kinetics. Under aerobic conditions, the t1/2 of D-isoidide dinitrate was significantly shorter than that of L-isoidide dinitrate (11.9 vs. 14.1 min, p ≤ 0.05), which is consistent with the greater potency of the D-enantiomer for vasodilation. Under anaerobic conditions, the denitration of glyceryl trinitrate was very rapid (t1/2 approximately 30 s). Organic nitrate biotransformation was inhibited by carbon monoxide, SKF 525A, and dioxygen. This suggests that the biotransformation of organic nitrates can occur through the direct interaction with the heme moiety of cytochrome P-450. The biotransformation of glyceryl trinitrate was catalyzed preferentially by those isoenzymes induced by phenobarbital. The biotransformation of glyceryl trinitrate was regioselective for 1,3-glyceryl dinitrate formation except in phenobarbital-induced microsomes under aerobic conditions, in which preferential formation of 1,2-glyceryl dinitrate occurred. These data suggest that cytochrome P-450 is involved in the biotransformation of organic nitrates and raises the possibility that vascular cytochrome P-450 may play a role in the mechanism-based biotransformation of organic nitrates, the result of which is vascular smooth muscle relaxation.Key words: cytochrome P-450, glyceryl trinitrate, isoidide dinitrate, biotransformation, liver.

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Nitric oxide stimulates cyclic GMP in human thyrocytes

Journal of Molecular Endocrinology ◽

10.1677/jme.0.0100163 ◽

1993 ◽

Vol 10 (2) ◽

pp. 163-169 ◽

Cited By ~ 21

Author(s):

L J Millatt ◽

R Jackson ◽

B C Williams ◽

G StJ Whitley

Keyword(s):

Nitric Oxide ◽

Sodium Nitroprusside ◽

Cyclic Gmp ◽

Cell Types ◽

Inhibitory Effect ◽

Human Thyroid ◽

Thyroid Cell ◽

Camp Production ◽

Cgmp Production ◽

Thyroid Cell Line

ABSTRACT Sodium nitroprusside spontaneously breaks down in solution to produce the vasodilator nitric oxide. In many cell types, this stimulates the cytosolic form of the enzyme guanylate cyclase, resulting in the elevation of cyclic GMP (cGMP). We have investigated the effect of sodium nitroprusside on the generation of cGMP in primary human thyrocytes and the SV40-transfected human thyroid cell line SGHTL-189. A dose-dependent increase in cGMP was obtained and the maximum response was observed with concentrations above 10 μm sodium nitroprusside in both cell types. Methylene blue (50 μm) had no significant effect on basal cGMP production but inhibited the effect of sodium nitroprusside at all concentrations tested, thus demonstrating that the effect was due to nitric oxide. Sodium nitroprusside had no effect on cyclic AMP (cAMP) production in these cells. TSH at 100 and 1000 μU/ml significantly stimulated the production of cAMP, but not that of cGMP, in primary human thyrocytes. Sodium nitroprusside had no significant effect on basal or TSH-stimulated triiodothyronine secretion in primary human thyrocytes. Forskolin (10 μm) significantly stimulated cAMP production in both primary thyrocytes and SGHTL-189 cells. Although forskolin had no significant effect on basal cGMP production, sodium nitroprusside-stimulated cGMP production was significantly reduced by forskolin. However, this inhibitory effect was not related to the production of cAMP.

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