Cystine Uptake and Glutathione Level in Fetal Brain Cells in Primary Culture and in Suspension

Abstract— The glutathione level and the factors affecting this level were investigated in fetal rat brain cells in a primary culture. Early in the culture, the glutathione level of the brain cells decreased, but after 5 h it began to increase. This increase was not observed in a cystine-free medium and was prevented by excess glutamate. Cystine was taken up in freshly isolated brain cell suspensions, and its rate increased during the culture. The cystine uptake was mediated by a Na⁺-independent, glutamate-sensitive route previously found in various types of cells and designated as system x⁻_c. The uptake of cystine is a crucial factor in maintaining the glutathione level of the cells under culture, because it provides cysteine for the cells for glutathione synthesis. Cysteine was undetectable in the medium before the culture, but it appeared, though at a very low level, when the brain cells were cultured there. The source of this cysteine was the cystine in the medium. Presumably the decrease in the glutathione level of the cells in the early stage of the culture resulted from the fact that the medium did not contain cysteine. The enhancement of the cystine uptake during culture may constitute a protective mechanism against the oxidative stress to which the cultured cells are exposed. Regulation of the glutathione level in fetal brain cells in vivo by the transport of cystine and cysteine is discussed.     

Age-Related Regional Changes in Hydroxyl Radical Stress and Antioxidants in Gerbil Brain

Abstract— The levels of hydroxyl radicals and oxidized GSH have been examined as indices of oxidative stress in young (3 months), middle-aged (15 months), and old (20–24 months) gerbil brain hippocampus, cortex, and striaturn. The hydroxyl radical stress was estimated by measuring the salicylate hydroxyl radical trapping products 2,5-and 2,3-dihydroxybenzoic acid. The stress was significantly higher in all three brain regions in middle-aged and old gerbils versus young animals (66.0%). Regional comparisons showed that the stress was significantly higher in cortex than in either the hippocampus or striatum of the middle-aged and old gerbils (32.0%). The ratio of oxidized to total GSH also increased progressively in middle-aged and old animals in all three brain regions (p < 0.05, 41.1%), further indicating a general age-related increase in oxidative stress. Parallel to this age-related increase in oxidative stress, a significant, albeit slight (8%), decrease in neuronal number in hippocampal CA1 region was observed in both the middle-aged and old animals. Possible differences in antioxidant levels were also examined. Total GSH levels were similar across age groups (variance <12%). However, the regional comparison showed that it was highest in striatum in all age groups. The levels of a-tocopherol (vitamin E) were significantly higher in the middle-aged and old animals in all three regions (70.4%). Vitamin E was highest in the hippocampus and the differences between the hippocampus and the cortex and striatum increased with age. Although of a lesser magnitude, significant increases in hippocampal total ascorbic acid level were also noted with age (p < 0.05, 10%). Ascorbic acid was the most regionally specific of the three antioxidants examined, with hippocampus > cortex > striatum for all age groups. The difference in ascorbic acid level between hippocampus and cortex also increased with age (64.4%). The results suggest that the general age-related, regionally specific increases in oxidative stress stimulate the accumulation of antioxidants. It is interesting that the hippocampus, which is selectively vulnerable to various insults such as ischemia, epilepsy, and insulin-induced hypoglycemia, exhibits the greatest age-related increase in vitamin E and ascorbic acid, perhaps reflective of a greater impact of the progressive increase in baseline oxidative stress.