EFFECT OF BRAIN ISCHEMIA AND REPERFUSION ON THE LOCALIZATION OF PHOSPHORYLATED EUKARYOTIC INITIATION-FACTOR 2-ALPHA

Postischemic brain reperfusion is associated with a substantial and lo ng-lasting reduction of protein synthesis in selectively vulnerable ne urons. Because the overall translation initiation rate is typically re gulated by altering the phosphorylation of serine 51 on the alpha-subu nit of eukaryotic initiation factor 2 (eIF-2 alpha), we used an antibo dy specific to phosphorylated eIF-2 alpha [eIF-2(alpha P)] to study th e regional and cellular distribution of elF-2(alpha P) in normal, isch emic, and reperfused rat brains. Western blots of brain postmitochondr ial supernatants revealed that similar to 1% of all eIF-2 alpha is pho sphorylated in controls, eLF-2(alpha P) is not reduced by up to 30 min utes of ischemia, and eIF-2(alpha P) is increased similar to 20-fold a fter 10 and 90 minutes of reperfusion. Immunohistochemistry shows loca lization of eIF-2(alpha P) to astrocytes in normal brains, a massive i ncrease in eIF-2(alpha P) in the cytoplasm of neurons within the first 10 minutes of reperfusion, accumulation of eIF-2(alpha P) in the nucl ei of selectively vulnerable neurons after 1 hour of reperfusion, and morphology suggesting pyknosis or apoptosis in neuronal nuclei that co ntinue to display eIF-2(alpha P) after 4 hours of reperfusion. These o bservations, together with the fact that eIF-2(alpha P) inhibits trans lation initiation, make a compelling case that eIF-2(alpha P) is respo nsible for reperfusion-induced inhibition of protein synthesis in vuln erable neurons.