Selective modifiers of glutathione biosynthesis and ‘repriming’ of vascular smooth muscle photorelaxation

Abstract

1 Photorelaxation of vascular smooth muscle (VSM) is caused by the release of nitric oxide (NO) from a ®nite molecular store that can be depleted by irradiating pre-contracted arteries with visible light. The ability of an`exhausted' vessel to respond to a further period of illumination is lost temporarily but then recovers slowly as the photosensitive store is reconstituted in the dark. The recovery process, termed repriming, displays an absolute requirement for endothelium-derived NO and is inhibited by pre-treating arteries with ethacrynic acid, a thiol-alkylating agent. Here we demonstrate that agents that up-or down-regulate glutathione (GSH) biosynthesis in¯uence the extent to which the store is regenerated in the dark. 2 Isolated rat tail arteries (RTAs) were perfused internally with Krebs solution containing phenylephrine (PE; mean [PE]+s.e.mean: 5.78+0.46 mM) and periodically exposed to laser light (l=514.5 nm, 6.3 mW cm 72 for 6 min). Photorelaxations of control RTAs were compared with those from either (a) vessels taken from animals previously injected i.p. with buthionine sulphoximine (BSO), an inhibitor of g-glutamylcysteine synthetase (three injections, 100 mg kg 71 at 8 h intervals); or (b) isolated RTAs that were perfused ex vivo with oxothiazolidine (OXO), a precursor of cysteine (10 74 M OXO for 60 min). RTAs from BSO-treated animals exhibited attenuated photorelaxations: the mean (+s.e.mean) amplitude of the response recorded after 72 min recovery in the dark was 12.4+1.6% versus 21.4+2.9% for control arteries (n=5; P50.01). Conversely RTAs treated with OXO and allowed to recover for a similar period showed enhanced photorelaxations, 32.6+6.3% as compared to 21.4+2.9% for control arteries (n=5; P50.01). A hyperbolic curve ®t to repriming curves for BSO-treated and control arteries returned asymptote values (maximum photorelaxations) of (mean+s.e.mean) 24.2+3.2% and 55.2+8.5%, respectively. 3 The level of GSH in RTA extracts was measured by high-pressure liquid chromatography (HPLC). Injecting animals with BSO decreased GSH to 85% of control levels (P50.05) while treatment of isolated vessels with OXO resulted in a 31% increase above control levels (P50.05). Thus, drug-induced changes in RTA GSH levels were positively correlated with altered photorelaxations. 4 The results lead us to postulate that the photosensitive store in VSM is generated, at least in part, from intracellular GSH which becomes converted to S-nitrosoglutathione (GSNO) by nitrosating species that are formed ultimately from endothelium-derived NO. The possible physiological signi®cance of a photolabile store of NO in VSM is discussed brie¯y.

H2S and HS− donor NaHS releases nitric oxide from nitrosothiols, metal nitrosyl complex, brain homogenate and murine L1210 leukaemia cells

“…It has been proposed that low molecular weight thiols such as glutathione and cysteine, which react with nitric oxide or oxides of nitrogen to form nitrosothiols, are potential candidates for a nitric oxide-carrier molecule [16,17]. Endogenous nitrosothiols, e.g.…”

Carbon Monoxide: Endogenous Production, Physiological Functions, and Pharmacological Applications

“…S-nitrosoglutathione, generated from the interaction of GSH with peroxynitrite, releases NO in a time-dependent manner from vascular tissues (Balazy et al, 1998) or in a light-sensitive manner from cultured leukemia cells (Sexton et al, 1994). In fact, even GSH alone can be a light-sensitive intracellular NO pool (Megson et al, 2000).…”

Chemical nature of nitric oxide storage forms in rat vascular tissue

“…Indeed, nitrite ions do undergo NO photolysis, showing a maximum around 354 nm (15). Other studies claimed to have identified RSNOs (16)(17)(18) as the chemical species responsible for photorelaxation. However, they possess a UVabsorption band peaking in the 330-to 340-nm range (19,20), which does not coincide with any of the action spectrum peaks reported in previous studies.…”