VITAMIN-E ENHANCES CA2-MEDIATED VULNERABILITY OF IMMATURE CEREBELLAR GRANULE CELLS TO ISCHEMIA()

The effects of vitamin E on lipid peroxidation, intracellular free Ca2 + concentration ([Ca2+](i)), and cell death were investigated in the p ostischemic immature cerebellum. Deprivation of oxygen and glucose for 10-min in a suspension of freshly dissociated granule cells from the cerebellum of 9-day-old male rat pups resulted in a recovery-induced c onsumption of cell nonenzymatic antioxidants (ascorbic acid, glutathio ne, and alpha-tocopherol) and development of membrane lipid peroxidati on as measured by the thiobarbituric acid method. The rate of lipid pe roxidation of the postischemic cells was stimulated, not reduced, by t reatment of the cells with vitamin E (5-30 mu M alpha-tocopherol phosp hate). In flow-cytometric studies a 10-min period of ischemia resulted ina small increase in intracellular calcium concentration, lipid pero xidation products and cell death, but in the presence of alpha-tocophe rol the same treatment caused a dramatic increase in cell death, accom panied by a large increase in [Ca2+](i) and lipid peroxidation product s. Pretreatment of the cells with a mixture of three antioxidants (vit amin C/rutin/ubiquinol-10, 10/5/1) or nickel (Ni2+) reduced the alpha- tocopherol-induced increases in [Ca2+](i), and cell death. Hydrogen pe roxide (1 mM) and the water-soluble analogue of vitamin E, trolox (50 mu M), mimicked the effect of vitamin E on lipid peroxidation in the p ostischemic cells. Pretreatment of the cells with the intracellular Ca 2+ chelator, BAPTA-AM, reduced both the cy-tocopherol-induced increase in [Ca2+](i) and cell death. The effect of vitamin E on [Ca2+](i) was age dependent and decreased abruptly during maturation of the cerebel lum between the first and second weeks of Life. Results of in vitro tr eatment of the immature cerebellar cells with the water-soluble form o f vitamin E (alpha-tocopherol phosphate) suggest that, after consumpti on of cellular co-antioxidants, vitamin E may be converted to an alpha -tocopheroxyl radical, which acts as a toxic prooxidant as cellular bi oenergetics deteriorate. (C) 1998 Elsevier Science Inc.