Biosynthesis of endothelium-derived relaxing factor: A cytosolic enzyme in porcine aortic endothelial cells Ca2+-dependently converts L-arginine into an activator of soluble guanylyl cyclase

Mayer, Bernd; Schmidt, Kurt; Humbert, Peter; Böhme, Eycke

doi:10.1016/0006-291x(89)91513-1

Cited by 251 publications

(111 citation statements)

References 16 publications

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“…Since this L-NAME-induced potentiation was abolished in endothelium-denuded mesenteric arterial beds, it can be concluded that the action of L-NAME resulted from an inhibitory action on NO synthesis in endothelial cells. The endothelial NO synthase requires Ca2" and NADPH as co-factors for the conversion of L-arginine to NO and L-citrulline (Myers et al, 1989); thus, the failure of L-NAME to potentiate aadrenoceptor-mediated pressor responses obtained during perfusion with Ca2"-free PSS further supports the conclusion that endothelial NO synthase is the target of L-NAME action in the present study.…”

Section: Discussionsupporting

confidence: 85%

The effects of perfusion rate and N^G‐nitro‐L‐arginine methyl ester on cirazoline‐ and KCl‐induced responses in the perfused mesenteric arterial bed of rats

Adeagbo¹,

Tabrizchi²,

Triggle³

1994

British J Pharmacology

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1 The purpose of this study was to characterize the effects of N0-nitro-L-arginine methyl ester (L-NAME) on the perfusion rate/pressure relations, and on the pressor responses induced to cirazoline and KC1 in isolated, perfused mesenteric arterial beds from normotensive and spontaneously hypertensive rats. 2 The basal perfusion pressure of arterial beds perfused with either physiological salt solution (PSS) or PSS containing 1% polyvinylpyrrolidone increased as the perfusion rate increased. L-NAME, in concentrations up to 100 .lM, failed to alter the basal pressure regardless of the perfusion rate and viscosity; however, at 5 gM, it potentiated cirazoline-induced vasoconstriction at each of the perfusion rates. 3 L-NAME but not D-NAME caused a leftward shift of cirazoline concentration-response curves with a marked increase in the maximal response. The potentiating action of L-NAME was abolished in arterial beds perfused with a Ca2"-free physiological salt solution and also in beds denuded of endothelium by an infusion of distilled water for 5 min. 4 In endothelium-intact and -denuded preparations, L-NAME potentiated KCl pressor responses; the endothelium-independent potentiation of KCl pressor activity was stereospecific, time-independent and was not prevented by the presence of dexamethasone (0.511M) in the perfusion medium. However, L-NAME failed to potentiate vasoconstriction obtained to KCl in arterial beds denervated by cold storage (4-5°C) for 2 days. 5 The absence of K+ in the perfusate did not inhibit the ability of L-NAME to potentiate aadrenoceptor-mediated pressor responses, and nor did L-NAME inhibit KCl-induced vasodilatation in preconstricted arteries. It was thus concluded that L-NAME does not affect Na+/K+-ATPase activity. 6 No differences in the potentiating ability of L-NAME on either cirazoline-or KCl-mediated pressor responses were apparent between normotensive Sprague Dawley (SD), Wistar Kyoto (WKY) and spontaneously hypertensive (SHR) rats. 7 Our data thus provide evidence that: the presence of a vasoconstrictor is required for basal nitric oxide (NO) release in the mesenteric arterial bed from either normotensive or spontaneously hypertensive rats; L-NAME causes potentiation of cirazoline-and KCl-induced vasoconstriction respectively by inhibiting endothelial and neuronal NO synthase(s). Furthermore, our data indicate that NO synthase activity is not impaired in the mesenteric arterial bed of spontaneously hypertensive rats.

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Section: Discussionsupporting

confidence: 85%

The effects of perfusion rate and N^G‐nitro‐L‐arginine methyl ester on cirazoline‐ and KCl‐induced responses in the perfused mesenteric arterial bed of rats

Adeagbo¹,

Tabrizchi²,

Triggle³

1994

British J Pharmacology

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“…We found that KCl-insoluble fractions of PAEC contained 80-95% of total NOS with a specific activity ranging from 50 to 100 pm01 L-citrulline x mg-' x min-', whereas NOS activity was below 5 pmol x mg-' x min-' in the supernatants. These results may explain the previously observed poor Larginine-induced stimulation of purified soluble guanylyl cyclase co-incubated with PAEC cytosols [2].…”

Section: Determination Of L-citrulline Formationsupporting

confidence: 60%

“…NO release is probably mediated by intracellular free Ca2+ required for activation of a constitutively expressed, calmodulin-dependent NO synthase (NOS) [2,3]. Whereas cytokine-inducible and neuronal isoforms of NOS are predominantly cytosolic [4,5], the endothelial enzyme is associated with membranes [6] due to posttranslational myristilation at its N-terminus [7,8].…”

Section: Introductionmentioning

confidence: 99%

Reversible inactivation of endothelial nitric oxide synthase by N^G‐nitro‐l‐arginine

et al. 1993

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&'"-Methyl-L-arginine (L-NMA) and AJo-nitro-L-arginine (L-NNA) inhibited NO-induced cGMP accumulation in porcine aortic endothelial cells with half-maximally effective concentrations of 15 and 3.4,~M, respectively. The effects of both compounds were reversible, but the L-NNA-induced inhibition was only reversed by wash-out in the presence of 1 mM L-arginine. In short-term incubations (45 s) of membrane fractions, L-NMA and L-NNA exhibited similar potencies to inhibit endothelial NO synthase, but L-NNA was markedly more potent than L-NMA after prolonged incubation periods (2 3 min) due to induction of a pronounced, reversible enzyme inactivation.

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“…N0-methyl-L-arginine (L-NMMA) was used to demonstrate the precursor role of L-arginine in NO formation by activated macrophages (Hibbs et al, 1987) and vascular endothelial cells (Palmer et al, 1988). L-NMMA was shown to attenuate endothelium-dependent relaxations both in vivo and in vitro (Rees et al, 1989a, b) and to block NO synthesis by endothelial cell homogenates (Mayer et al, 1989;Palmer & Moncada, 1989). Since then, many more L-argininebased NOS inhibitors have been described (Fukuto & Chaudhuri, 1995).…”

Section: Introductionmentioning

confidence: 99%

Inhibition of nitric oxide synthesis by N^G‐nitro‐L‐arginine methyl ester (L‐NAME): requirement for bioactivation to the free acid, N^G‐nitro‐L‐arginine

Pfeiffer

Leopold

Schmidt

et al. 1996

British J Pharmacology

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212

125

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1 The L-arginine derivatives N0-nitro-L-arginine (L-NOARG) and NG-nitro-L-arginine methyl ester (L-NAME) have been widely used to inhibit constitutive NO synthase (NOS) in different biological systems. This work was carried out to investigate whether L-NAME is a direct inhibitor of NOS or requires preceding hydrolytic bioactivation to L-NOARG for inhibition of the enzyme. 2 A bolus of L-NAME and L-NOARG (0.25 Mmol) increased coronary perfusion pressure of rat isolated hearts to the same extent (21 +0.8 mmHg; n = 5), but the effect developed more rapidly following addition of L-NOARG than L-NAME (mean half-time: 0.7 vs. 4.2 min). The time-dependent onset of the inhibitory effect of L-NAME was paralleled by the appearance of L-NOARG in the coronary effluent. 3 Freshly dissolved L-NAME was a 50 fold less potent inhibitor of purified brain NOS (mean IC50 = 70 gM) than L-NOARG (IC50= 1.4 giM), but the apparent inhibitory potency of L-NAME approached that of L-NOARG upon prolonged incubation at neutral or alkaline pH. H.p.l.c. analyses revealed that NOS inhibition by L-NAME closely correlated with hydrolysis of the drug to L-NOARG. 4 Freshly dissolved L-NAME contained 2% of L-NOARG and was hydrolyzed with a half-life of 365 + 11.2 min in buffer (pH 7.4), 207 ± 1.7 min in human plasma, and 29 + 2.2 min in whole blood (n = 3 in each case). When L-NAME was preincubated in plasma or buffer, inhibition of NOS was proportional to formation of L-NOARG, but in blood the inhibition was much less than expected from the rates of L-NAME hydrolysis. This was explained by accumulation of L-NOARG in blood cells. 5 These results suggest that L-NAME represents a prodrug lacking NOS inhibitory activity unless it is hydrolyzed to L-NOARG. Bioactivation of L-NAME proceeds at moderate rates in physiological buffers, but is markedly accelerated in tissues such as blood or vascular endothelium.

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Biosynthesis of endothelium-derived relaxing factor: A cytosolic enzyme in porcine aortic endothelial cells Ca2+-dependently converts L-arginine into an activator of soluble guanylyl cyclase

Cited by 251 publications

References 16 publications

The effects of perfusion rate and N^G‐nitro‐L‐arginine methyl ester on cirazoline‐ and KCl‐induced responses in the perfused mesenteric arterial bed of rats

The effects of perfusion rate and N^G‐nitro‐L‐arginine methyl ester on cirazoline‐ and KCl‐induced responses in the perfused mesenteric arterial bed of rats

Reversible inactivation of endothelial nitric oxide synthase by N^G‐nitro‐l‐arginine

Inhibition of nitric oxide synthesis by N^G‐nitro‐L‐arginine methyl ester (L‐NAME): requirement for bioactivation to the free acid, N^G‐nitro‐L‐arginine

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Biosynthesis of endothelium-derived relaxing factor: A cytosolic enzyme in porcine aortic endothelial cells Ca2+-dependently converts L-arginine into an activator of soluble guanylyl cyclase

Cited by 251 publications

References 16 publications

The effects of perfusion rate and NG‐nitro‐L‐arginine methyl ester on cirazoline‐ and KCl‐induced responses in the perfused mesenteric arterial bed of rats

The effects of perfusion rate and NG‐nitro‐L‐arginine methyl ester on cirazoline‐ and KCl‐induced responses in the perfused mesenteric arterial bed of rats

Reversible inactivation of endothelial nitric oxide synthase by NG‐nitro‐l‐arginine

Inhibition of nitric oxide synthesis by NG‐nitro‐L‐arginine methyl ester (L‐NAME): requirement for bioactivation to the free acid, NG‐nitro‐L‐arginine

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The effects of perfusion rate and N^G‐nitro‐L‐arginine methyl ester on cirazoline‐ and KCl‐induced responses in the perfused mesenteric arterial bed of rats

The effects of perfusion rate and N^G‐nitro‐L‐arginine methyl ester on cirazoline‐ and KCl‐induced responses in the perfused mesenteric arterial bed of rats

Reversible inactivation of endothelial nitric oxide synthase by N^G‐nitro‐l‐arginine

Inhibition of nitric oxide synthesis by N^G‐nitro‐L‐arginine methyl ester (L‐NAME): requirement for bioactivation to the free acid, N^G‐nitro‐L‐arginine