| During global brain ischemia and for at least a short time after perfusion is reestablished, there is a profound inhibition of new protein synthesis in neurons. This is reversible in many areas of the brain, but persists in neurons known as selectively vulnerable neurons (SVNs). Previous work in our lab has shown that phosphorylation of eukaryotic initiation factor-2α [eIF2α(P)] accounts for most if not all of this inhibition of protein synthesis, and that dephosphorylation of eIF2α rescues the neuron's competence to synthesize new protein.; Moreover, even though SVNs do not physically disintegrate before 48–72 hours after ischemia, early apoptotic biochemical events such as release of cytochrome c into the cytoplasm are discernible by as early as 3 hours. There is considerable evidence for causal linkage between eIF2α(P) and induction of apoptosis which in vulnerable neurons appears to involve mitochondrial release of cytochrome c and caspase-9.; Utilizing a rat model of cardiac arrest and resuscitation, this study examined the phosphorylation state of eIF2α throughout the entire brain at 10 minutes and four hours of post-ischemic reperfusion; and the colocalization at 4 hours reperfusion of persistent eIF2α(P) with cytoplasmic cytochrome c in vulnerable brain neurons.; The immunohistochemical data show that early in reperfusion, phosphorylated eIF2α is present in neuronal populations in almost all brain regions; those that are selectively vulnerable, as well as those more resistant to death. By four hours of reperfusion, eIF2α(P) immunostaining appears little changed from the 10 minute reperfused brains, with exceptions in the CA1 and CA3 regions of the hippocampus, and some brainstem nuclei, which exhibit a noticeable reduction of label. However, results of immunoblotting brain homogenates at 4 hours reperfusion shows a nearly 60% reduction of total eIF2α(P) levels in the brain, suggesting that the DAB-visualized immunohistochemistry may not be sensitive enough to distinguish any less-than-profound differences in eIF2α(P) concentrations. Immunofluorescent studies of both cytochrome c and eIF2α(P) localization at 4 hours reperfusion revealed that cytosolic cytochrome c was present only in neuronal populations which also had persistence of eIF2α(P). Double immunofluorescence in CA1, CA3, and striatum showed this colocalization in the same neurons. |
