Cellular and subcellular calcium accumulation during glutamate-induced injury in cerebellar granule neurons

We have investigated the role of Ca²⁺ accumulation and neuronal injury in cerebellar granule neurons after glutamate receptor overactivation. After the removal of the free cytosolic Ca²⁺ we identified an extensive second Ca²⁺ fraction (SCF) that is retained within the neurons after glutamate receptor overactivation. The SCF reaches a plateau within 10 min with the magnitude of this SCF accumulation reflecting the extent of the neuronal injury that occurs within the neurons. The existence of this SCF is sensitive to both NMDA receptor antagonists and mitochondrial inhibitors but is unaffected by agents that deplete endoplasmic reticulum Ca²⁺, indicating that this Ca²⁺ fraction may be located within the mitochondria. Through the isolation of mitochondria from cerebellar granule neurons treated with glutamate we have shown that the majority of the SCF is mitochondrial in location. On the removal of the glutamate stimulus the SCF recovers at a slower rate than the free Ca²⁺ concentration within the neuron. This is intriguing, as it implies a capacity to remember previous excitatory events. Most significantly we have shown that a short pre-application of subthreshold glutamate or kainate blocks both SCF Ca²⁺ accumulation and extensive neuronal injury in response to high concentrations of glutamate. These findings may be relevant to the observations of pre-conditioning in the brain and heart.