INABILITY TO RESTORE RESTING INTRACELLULAR CALCIUM LEVELS AS AN EARLYINDICATOR OF DELAYED NEURONAL CELL-DEATH

The hippocampus is especially vulnerable to excitotoxicity and delayed neuronal cell death. Chronic elevations in free intracellular calcium concentration ([Ca2+](i)) following glutamate-induced excitotoxicity have been implicated in contributing to delayed neuronal cell death. H owever, no direct correlation between delayed cell death and prolonged increases in [Ca2+](i) has been determined in mature hippocampal neur ons in culture. This investigation was initiated to determine the stat istical relationship between delayed neuronal cell death and prolonged increases in [Ca2+](i) in mature hippocampal neurons in culture. Usin g indo-1 confocal fluorescence microscopy, we observed that glutamate induced a rapid increase in [Ca2+](i) that persisted after the removal of glutamate. Following excitotoxic glutamate exposure, neurons exhib ited prolonged increases in [Ca2+](i), and significant delayed neurona l cell death was observed. The N-methyl-D-aspartate (NMDA) channel ant agonist MK-801 blocked the prolonged increases in [Ca2+](i) and cell d eath. Depolarization of neurons with potassium chloride (KCI) resulted in increases in [Ca2+](i), but these increases were buffered immediat ely upon removal of the KCL, and no cell death occurred. Linear regres sion analysis revealed a strong correlation (R = 0.973) between glutam ate-induced prolonged increases in [Ca2+](i) and delayed cell death. T hese data suggest that excitotoxic glutamate exposure results in an NM DA-induced inability to restore resting [Ca2+](i) (IRRC) that is a sta tistically significant indicator of delayed neuronal cell death.