Excitatory amino acid‐induced alterations of cytoplasmic free Ca 2+ individual cerebellar granule neurons: Role in neurotoxicity

The effects of glutamate on intracellular free Ca 2+ [Ca 2+ ] i , and neurotoxicity were compared in cerebellar granule neurons in vitro. [Ca 2+ ] i , was measured with fura‐2 and digital fluorescence imaging microscopy; neurotoxicity was monitored using a vital dye and colorimetric analysis. Glutamate produced dosedependent increases in [Ca 2+ ] XSi transient for glutamate concentrations in a range of : 0.01–0.5 μM and sustained for higher levels of glutamate. The ED 50 for the [Ca 2+ ] i response to glutamate was. 6 μM. The LQ 50 for glutamate‐induced neurotoxicity was similar, i.e., 10 μM The effect of glutamate on [Ca 2+ ] i was greatly dmiinished when external Ca 2+ was removed and blocked by Mg 2+ + N‐methyl‐D‐aspartate (NMDA)‐type receptor antagonists. The latter conditions as well as preloading granule neurons with the intracellular Ca 2+ chelator quin2 largely prevented glutamate cytotoxicity. The neurotoxic effect of glutamate required incubations with the stimulus for 10–20 min at 25°C. Withdrawal of glutamate after this period was accompanied by a prolonged alteration in [Ca 2+ ] i . Pretreatment cells with the ganglioside GM1 reduced this late increase in [Ca 2+ ] i , as well as the neurotoxic effects of glutamate. This indicates that glutamate‐induced neurotoxicity results from a composite of diverse temporal alterations in Ca 2+ homeostasis and that blunting any of these components reduces excitotoxicity.