A putative role for S-nitrosoglutathione as the source of nitric oxide in photorelaxation of the mouse gastric fundus

Abstract

Mouse gastric fundus strips were relaxed by ultraviolet light (UV) irradiation, exogenous nitric oxide (NO), isoproterenol, S-nitrosoglutathione, S-nitroso-l-cysteine and S-nitroso-N-acetyl-penicillamine. Glutathione did not affect relaxations in response to UV irradiation, exogenous NO and isoproterenol while inhibiting that with S-nitrosoglutathione. l-Cysteine inhibited responses to UV irradiation and exogenous NO, but not in the presence of exogenous Cu²⁺/Zn²⁺ superoxide dismutase. However, l-cysteine alone or in combination with Cu²⁺/Zn²⁺ superoxide dismutase did not affect the relaxations in response to S-nitroso-l-cysteine. Ethacrynic acid and diamide inhibited photorelaxations but not the relaxations with exogenous NO and isoproterenol. This inhibition was prevented by glutathione, but not by l-cysteine. S-nitrosoglutathione-induced relaxations were abolished by diamide and ethacrynic acid, whereas responses to S-nitroso-l-cysteine and S-nitroso-N-acetyl-penicillamine were only inhibited by ethacrynic acid. These results suggest that S-nitrosoglutathione may, at least in part, be the putative S-nitrosothiol, which is converted to NO in response to UV irradiation in mouse gastric fundus strips.

Introduction

In 1955, Furchgott et al. demonstrated that isolated mammalian arteries in a state of active tone contraction relaxed when exposed to ultraviolet light (UV). Three decades later, the photorelaxation of vascular smooth muscle was shown to be independent of endothelium, inhibited by haemoglobin and methylene blue, and accompanied by an increase in guanosine 3′-5′ cyclic monophosphate (cGMP) (Furchgott et al., 1985). In addition, Matsunaga and Furchgott (1989) indicated that photorelaxation might also be due to endogenous nitric oxide (NO) liberated from a photodegradable molecular “store” of NO contained within the vessel wall. Subsequently, this hypothesis was supported by Venturini et al. (1993), who suggested a light-activated, depletable and replenishable NO-yielding store as being responsible for the photorelaxation of vascular smooth muscle in the isolated rabbit aortic strips. A similar conclusion was reached by Kubaszewski et al. (1994), who showed directly the release of NO via UV light by using a porphyrinic sensor in vascular smooth muscle. These authors' observations that repeated treatments with UV light cause diminished vascular smooth muscle relaxation and NO release indicate that UV light causes the release of NO from a depletable store.

In our previous study (Ögülener et al., 1996), UV light caused relaxation in isolated mouse gastric fundus strips. The photorelaxation was inhibited by haemoglobin, hydroxocobalamin, FeSO₄ and methylene blue, whereas N^G-monomethyl-l-arginine had no effect. These observations were interpreted as indicating that release of NO by UV light causes photorelaxation. Recently, Büyükavşar et al. (1999) showed that there could be a store of photosensitive compounds yielding NO in smooth muscle strips of the mouse gastric fundus. However, the origin and anatomic localization of the NO store are not known.

There are several candidates for the source of NO, e.g. iron–sulphur complexes, which are photolysed to release NO (Flitney et al., 1993), and thiol groups, which can react with NO in order to form S-nitrosothiols Stamler et al., 1992, Megson et al., 1995, Lovren and Triggle, 1998. NO is an important mediator of smooth muscle relaxation in the peripheral nervous system Bult et al., 1990, Rand and Li, 1995; however, the short half-life of NO which may limit the efficacy and duration of its biological activity (Wood and Gartwaite, 1994) leads to the suggestion that NO might bind intracellularly to a carrier molecule, resulting in a more stable NO-containing compound from which NO is released at its site of action. It has been proposed that low molecular weight thiols such as glutathione and cysteine, which react with NO or oxides of nitrogen to form S-nitrosothiols, are likely candidates for such a NO-carrier molecule (Ignarro et al., 1981); these thiols are abundantly present in cells (Gow et al., 1997). Endogenous S-nitrosothiols may act as intermediates in the storage and/or transport of NO and consequently enhance the biological potency of NO in vascular Myers et al., 1990, Stamler et al., 1992, Gaston et al., 1994 and nonvascular smooth muscle (Kerr et al., 1992). The biological effects of these compounds is generally attributed to the activity of NO released on homolytic decomposition of the S–NO bond (Butler and Williams, 1993). Also, S-nitrosothiols can decompose photochemically to liberate NO and the corresponding disulfide Williams, 1983, Sexton et al., 1994, Singh et al., 1995, Singh et al., 1996a, Singh et al., 1996b.

Assuming the existence of NO stores in gastric fundus, we investigated whether the source of NO in photorelaxation is S-nitrosothiols in the mouse gastric fundus. For this purpose, we compared the effects of glutathione and l-cysteine, low molecular weight thiols, and ethacrynic acid and diamide, thiol modulators, on the relaxations in response to UV irradiation and S-nitrosothiols, S-nitrosoglutathione, S-nitroso-l-cysteine and S-nitroso-N-acetyl-penicillamine.