INVOLVEMENT OF BCL-2 FAMILY AND CASPASE-3-LIKE PROTEASE IN NO-MEDIATED NEURONAL APOPTOSIS

To clarify mechanisms of neuronal death in the postischemic brain, we examined whether astrocytes exposed to hypoxia/reoxygenation exert a n eurotoxic effect, using a coculture system. Neurons cocultured with as trocytes subjected to hypoxia/reoxygenation underwent apoptotic cell d eath, the effect enhanced by a combination of interleukin-1 beta with hypoxia. The synergistic neurotoxic activity of hypoxia and interleuki n-1 beta was dependent on de novo expression of inducible nitric oxide synthase (iNOS) and on nitric oxide (NO) production in astrocytes, Fu rther analysis to determine the neurotoxic mechanism revealed decrease d Bcl-2 and increased Bar expression together with caspase-3 activatio n in cortical neurons cocultured with NO-producing astrocytes. Inhibit ion of NO production in astrocytes by N-G-monomethyl-L-arginine, an in hibitor of NOS, significantly inhibited neuronal death together with c hanges in Bcl-2 and Bar protein levels and in caspase-3-like activity, Moreover, treatment of neurons with a bar antisense oligonucleotide i nhibited the caspase-3-like activation and neuronal death induced by a n NO donor, sodium nitroprusside. These data suggest that NO produced by astrocytes after hypoxic insult induces apoptotic death of neurons through mechanisms involving the caspase-3 activation after down-regul ation of Bcl-2 and up-regulation of Bar protein levels.