Neuronal cell death and reactive oxygen species

1. We have investigated the role of reactive oxygen species (ROS) in cell d eath induced by ischemia or application of the excitatory amino acid agonis t, N-methyl-D-aspartate (NMDA) or kainate (KA), in acutely isolated rat cer ebellar granule cell neurons, studied by flow cytometry. Various fluorescen t dyes were used to monitor intracellular calcium concentration, ROS concen tration, membrane potential, and viability in acutely dissociated neurons s ubjected to ischemia and reoxygenation alone, NMDA or kainate alone, and is chemia and reoxygenation plus NMDA or kainate. 2. With ischemia followed by reoxygenation, ROS concentrations rose slightl y and there was only a modest increase in cell death after 60 min. 3. When NMDA or kainate alone was applied to the cells there was a large in crease in ROS and in intracellular calcium concentration but only a small l oss of cellular viability. However, when NMDA or kainate was applied during the reoxygenation period there was a large loss of viability, accompanied by membrane depolarization, but the elevations of ROS and intracellular cal cium concentration were not greater than seen with the excitatory amino aci ds alone. 4. These observations indicate that other factors beyond ROS and intracellu lar calcium concentration contribute to cell death in cerebellar granule ce ll neurons.