During brain ischemia in vivo the extracellular concentration of the excitotoxic amino acid, glutamate, increases. This increase could be caused either by an enhanced formation rate of glutamate (from glutamine) or by an impaired re-uptake (or both). This re-uptake occurs to a large extent in astrocytes. In the present study we have determined glutamate uptake and the ability of the cells to maintain their glutamate content during exposure to anoxia, substrate deprivation and combined substrate deprivation and anoxia ('simulated ischemia') for a duration of up to 4 h. Isolated anoxia had no significant effect, whereas both substrate deprivation alone and 'simulated ischemia' reduced glutamate uptake and glutamate content by one-half after 2 h. Under hypothermic conditions (incubation at 32 degrees C), which in in vivo experiments exerts some protection against ischemic cell death in neurons, ischemia of intermediate duration (2 h) decreased glutamate uptake and glutamate content to a less extent than at 37 degrees C. Hypothermia did not have a similar effect during exposure to isolated substrate deprivation.