Extracellular L‐arginine is required for optimal NO synthesis by eNOS and iNOS in the rat mesenteric artery wall

Abstract

1 The formation of NO from endothelial nitric oxide synthase (eNOS) in rat superior mesenteric artery rings was dependent on extracellular l-arginine, and was optimal at a concentration of larginine close to the plasma level (carbachol-stimulated NO: control 15.770.9, l-arginine 100 mm 22.871.3 nm). 2 Enhancement of NO output by l-arginine was stereospecific, required the cationic amino-acid transporter and was dependent on caveolin. 3 Induction of inducible nitric oxide synthase (iNOS) impaired the stimulated NO synthesis from eNOS (100 nm carbachol-stimulated NO: control 5.770.6, iNOS 0.370.3 nm). 4 The interaction between iNOS and eNOS was reversed by the superoxide scavenger MnTMPyP. Impairment of eNOS by iNOS was also prevented by l-arginine 100 mm administered simultaneously with carbachol, but not by l-arginine administered during incubation with lipopolysaccharide. 5 These data provide functional evidence that supplementing l-arginine from the extracellular medium optimises the formation of NO from eNOS and suggests that the impairment of eNOS by iNOS is caused by excess formation of superoxide by NO synthase, which can be prevented by larginine. These results provide an explanation for the observations that extracellular l-arginine can enhance endothelium function only when the endothelium is impaired or when iNOS has been induced.

Physiologically Relevant Measurements of Nitric Oxide in Cardiovascular Research Using Electrochemical Microsensors

“…Simultaneous measurements of NO concentration and relaxation have also revealed that not only iNOS depends on influx of L -arginine, but endothelium-derived NO concentration and vasorelaxation are also increased by L -arginine supplementation in segments of the rat superior mesenteric artery [43]. The extra increase in NO concentration evoked by acetylcholine in the presence of L -arginine appeared to involve L -arginine uptake through the Y + transporter which is converted to NO primarily by NOS localized in caveolae.…”

“…From our experience with the 30-µm diameter polymer-coated carbon microsensor [5, 43], the characteristics vary significantly from one sensor to another and can change on a daily basis. Therefore, each sensor must pass a series of quality tests before being considered suitable for use (table 1).…”

“…NO formation occurred after platelet adhesion had reached a plateau and was able to prevent subsequent recruitment of platelets added at this time [47]. NO formation was prevented by N G -monomethyl- L -arginine [43] in endothelial NOS –/– mice [48] and was reduced in platelets from smokers [43]. Losartan and other angiotensin receptor antagonists were found to stimulate NO release from washed rat platelets, which correlated with inhibition of aggregation [49].…”

“…Introduction of an NO-sensitive microelectrode in the lumen of arterial segments mounted for isometric tension recording allows simultaneous measurements of NO concentration and vascular contractility, and hence, to establish direct relationships between them and address whether the release or response to NO is altered [5, 43, 55, 88] (fig. 3).…”

“…observation]. Therefore, electrodes with 30-µm tips have been applied in almost all the studies of simultaneous measurements of force and vascular contractility [5, 43, 55, 88]. In addition to the microelectrode tip, two holding wires with sizes of 20–100 µm are in the arterial lumen, and hence, exclude simultaneous measurements of NO and vascular contractility in arteries with lumen diameters below 100 µm; so far, simultaneous measurements of NO concentration and contractility have only been reported for arteries with lumen diameters of 200 µm or more [89].…”