Reduction of intracellular glutathione levels produces sustained arterial narrowing

Although lipid peroxidation and alterations in endogenous antioxidants have been hypothesized to contribute to cerebral vasospasm after subarachnoid hemorrhage, there has been no direct evidence demonstrating the relationship between oxidative stress and delayed arterial narrowing. To elaborate the role of the endogenous intracellular antioxidant and electron exchanger glutathione (GSH) in cerebral vasospasm, rat femoral arteries were treated with perivascular application of I-buthionine-(SR)-sulfoximine (BSO), which inhibits the synthesis of GSH. To determine the dose-response relationship, BSO at doses of 10 to 100 mg/ml, in platelet-rich plasma, was applied for 7 days to rat femoral arteries in vivo. Vessels were then perfusion-fixed for morphometric analysis of luminal cross-sectional area. To determine the time course of arterial narrowing, BSO (75 mg/ml) was applied to femoral arteries for 1, 3, 7, or 21 days before histological analysis, as described above. With rats treated with 50 to 100 mg/ml BSO, exogenous GSH (100 mg/kg) was administered, by intraperitoneal injection, daily for 7 days. To demonstrate the mechanism of BSO effects in smooth muscle cells (SMCs), cultured rat aortic SMCs were treated with 1 mmol/l BSO for 24 hours and assayed for intracellular levels of GSH and two products of lipid peroxidation, malondialdehyde and 4-hydroxyalkenal. Compared with control arteries treated with platelet-rich plasma alone, perivascularly administered BSO applied for periods of 1 to 21 days produced sustained and reversible narrowing of rat femoral arteries with a time course, severity, and histological appearance analogous to those observed after perivascular application of whole blood. BSO-induced arterial narrowing was dose-dependent, with 60% reductions in the luminal cross-sectional area being noted at 75 and 100 mg/ml (P < 0.005). Systemic administration of exogenous GSH slightly inhibited the effect of BSO on arterial narrowing, although the inhibition was not statistically significant. Cultured rat aortic SMCs exposed to BSO for 24 hours showed a 70% decrease in intracellular GSH levels (P = 0.03); levels of two products of lipid peroxidation, malondialdehyde and 4-hydroxyalkenal, were increased by 25% (P = 0.24) and 38% (P = 0.09), respectively. These data support the hypothesis that diminished intracellular levels of GSH may produce delayed chronic arterial narrowing after subarachnoid hemorrhage. The specific mechanism by which GSH levels m