Glucose Deprivation Increases Hydrogen Peroxide Level in Immunostimulated Rat Primary Astrocytes

Activated astrocytes produce a large amount of bioactive molecules, including reactive oxygen and nitrogen species. Astrocytes are in general resistant to those reactive species. However, we previously reported that immunostimulated astrocytes became highly vulnerable to metabolic insults, such as glucose deprivation. In this study, we investigated whether H ₂O₂ production was associated with the increased vulnerability. Glucose deprivation for up to 8 hr did not change the intracellular level of H₂O₂ in astrocytes. Treatment with lipopolysaccharide plus interferon-γ for 48 hr evoked astroglial H ₂O₂ production; however, no apparent death or injury was observed in immunostimulated astrocytes. Glucose deprivation after 48 hr of immunostimulation markedly increased H₂O₂ level, depleted adenosine triphosphate (ATP), and enhanced lactate dehydrogenase (LDH) release. The ATP depletion and LDH release were in part prevented by catalase, mannitol, and N-acetyl-L-cysteine. The enhanced level of H₂O₂ in glucose-deprived immunostimulated astrocytes appeared to be secondary to the depletion of reduced glutathione. 4-(2-Aminoethyl)bebzenesulfonyl fluoride (AEBSF), an inhibitor of NADPH oxidase, reduced H₂O₂ level and LDH release in glucose-deprived immunostimulated astrocytes. H ₂O₂, either endogenously produced or exogenously added, depolarized mitochondrial transmembrane potential in glucose-deprived astrocytes, leading to their ATP depletion and death. The present results strongly indicate that glucose deprivation causes deterioration of immunostimulated astrocytes by increasing the intracellular concentration of H₂O₂.