The major classes of glial cells, namely astrocytes, oligodendrocytes, and microglial cells were compared in parallel for their susceptibility to damage after combined hypoxia and hypoglycemia or hypoxia alone. The three glial cell types were isolated from neonatal rat brains, separated, and incubated in N2/CO2-gassed buffer-containing glucose or glucose substitutes, 2-deoxyglucose or mannitol (both nonmetabolizable sugars). The damage to the cells after 6 hours' exposure was determined at 0, 1, 3, 7 days based on release of lactate dehydrogenase and counting of ethidium bromide-stained dead cells, double-stained with cell-type specific markers. When 2-deoxyglucose replaced glucose during 6 hours of hypoxia, both oligodendrocytes and microglia rarely survived (18% and 12%, respectively). Astroglia initially increased the release of lactate dehydrogenase but maintained 98% to 99% viability. When mannitol, a radical scavenger and osmolarity stabilizer, replaced glucose during 6 hours of hypoxia, oligodendrocytes rarely survived (10%), astroglia survival remained at 99%, but microglia survival increased to 50%. After exposure to 6 and 42 hours, respectively, of hypoxic conditions alone, oligodendrocytes exhibited 10% survival whereas microglia and astroglia were only temporarily stressed and subsequently survived. In conclusion, oligodendrocytes, then microglia, are the most vulnerable glial cell types in response to hypoxia or hypoglycemia conditions, whereas astrocytes from the same preparations recover.