In vitro models differentiating between direct and indirect effects of ischemia on astrocytes

Abstract

Mouse astrocytes in primary cultures were subjected to an in vitro model of ischemia (hypoxia combined with substrate deprivation, excess potassium, or elevated glutamate) and examined with the light (phase) and electron microscope. Three hours of hypoxia alone or in combination with the other insults had little effect upon the morphology of astrocytes but did cause disaggregation of polyribosomes. With reoxygenation, polyribosomes reformed and many mitochondria changed from the orthodox to the condensed configuration. Notably, there was little swelling. Excess (50 m M) potassium, added (as KCl) to a normal isotonic medium, also caused no swelling. However, when 50 m M potassium was substituted for a similar amount of sodium, marked astrocyte swelling did occur. A morphologically similar swelling was seen when glutamate (50 μ M to 1 m M) was added to the culture medium, both with or without hypoxia with or without substrate deprivation. Potassium or glutamate-induced swelling was reversible with 1 h of recovery in normal medium. These results show that alterations in postischemic astrocytic morphology in vivo to a large extent can be reproduced in astrocytes in primary cultures. In addition, they suggest that postischemic astrocyte swelling is related to alterations in extracellular milieu, including accumulation of glutamate and/or alterations in the potassium/sodium ratios with increased potassium and decreased sodium. In contrast, morphologic alterations in polyribosomes and in mitochondria appear to be a direct response to ischemia itself.