During hypoxic-ischemic episodes in the brain, the resulting neuronal death is due, in part, to apoptosis, a genetically controlled form of cell death.p53, a tumor suppressor gene, is believed to promote apoptosis.bcl-2, a proto-oncogene, is believed to inhibit apoptosis. However, the role of these two genes in neuronal survival or death during hypoxia is not clear. We hypothesized that: a) neuronal expression of p53 and bcl-2 is altered during hypoxia, b) altered expression of p53 is associated with decreased neuronal survival during hypoxia, and c) altered expression of bcl-2 is associated with prolonged survival during hypoxia. Methods. 7-10 day old cultured rat neocortical neurons were exposed to 0.1,1, or 3% oxygen. Culture dishes were removed at 24-hour intervals for quantitative immunofluorescence staining and immunoblot analyses. Apoptosis was assessed by changes in cell morphology and in situ and genomic DNA fragmentation. To assess the impact of p53 and bcl-2 expression upon neuronal survival during hypoxia, expression of these genes was attenuated by treating selected neuronal culture plates with 3 μM antisense, sense, and nonsense oligonucleotides 24 hours prior to exposure to 0.1% oxygen. Neuronal survival was assessed by Trypan blue exclusion at 24-hour intervals during hypoxic exposure. Results. At each level of hypoxia studied, morphologic changes (cellular shrinkage, nuclear condensation) and in situ and genomic DNA fragmentation consistent with apoptosis was detected. Immunoblot and quantitative immunofluorescence analyses revealed an increase in both neuronal p53 and bcl-2 during hypoxic exposure. After 48 hours at 0.1% oxygen, the proportion of viable neurons was significantly greater in plates treated with p53 antisense oligonucleotide (≈60%) than in untreated plates(35-40%)(n=5;p<0.05). On the other hand, the proportion of viable neurons in plates treated with bcl-2 antisense was significantly lower (≈20%) as compared to untreated plates (≈40%)(n=3; p<0.05). The proportion of viable neurons in sense and nonsense treated plates was not significantly different from that of untreated plates (p>0.05). Conclusions.p53 and bcl-2 are increased in neurons during hypoxic exposure. Expression of p53 is, in part, necessary for hypoxia-induced neuronal apoptosis.bcl-2 expression, in part, prolongs neuronal survival during hypoxia.