IRREVERSIBLE anoxic injury of axons in the rat optic nerve requires the presence of extracellular Ca2+. To test the hypothesis that Ca2+enters an intracellular compartment during anoxia we monitored [Ca2+]0in this CNS white matter tract using ion-sensitive microelectrodes. Periods of anoxia lasting 15 min resulted in a rapid, reversible increase in [Ca2+]0accompanied by transient loss of nerve conduction. This increase in [Ca2+]0was apparently the result of extracellular space shrinkage. Anoxic periods lasting 60 min resulted in an initial rise followed by a sustained fall in [Ca2+]0, indicative of net influx of Ca2+into an intracellular compartment. Following reoxygenation after 60 min of anoxia, [Ca2+]0slowly returned toward control levels but nerve conduction recovered incompletely, indicating irreversible loss of function. Removal of bath Ca2+lowered [Ca2+]0to about 100 μM, prevented the anoxia-induced fall in [Ca2+]0, and protected against irreversible loss of the compound action potential.