Eukaryotic initiation factor 2 alpha kinase and phosphatase activity during postischemic brain reperfusion

When ischemic brain is reperfused, there is in vulnerable neurons immediate inhibition of protein synthesis associated with a large increase in phosph orylation of the alpha-subunit of eukaryotic initiation factor 2 [eIF2 alph a, phosphorylated form eIF2 alpha(P)]. We examined eIF2 alpha kinase and eI P2 alpha(P) phosphatase activity in brain homogenate postmitochondrial supe rnatants obtained from rats after 3 to 30 min of global brain ischemia (car diac arrest), after 5 min of ischemia and 5 min of reperfusion (5R), and af ter 10 min of ischemia and 90 min reperfusion (90R), Because it has been su ggested that PKR might be specifically responsible for producing eIF2 alpha (P) during reperfusion, we also examined in brain homogenates from wild-typ e and PKR0/0 C57BL/6J x 129/SV mice the effect of 5 min of ischemia and 5 m in of reperfusion on eIF2 alpha(P). Cytosolic brain elF2 alpha(P) in the 5R and 90R rats was 18- and 23-fold that of nonischemic controls without any change in the rate of elF2 alpha(P) dephosphorylation. There was no change in eIF2 alpha kinase activity between 3 and 30 min of ischemia but an 85% d ecrease in the 5R group; the 90R group was similar to controls. In wild-typ e and PKR0/0 mice total eIF2 alpha was identical, and there was an identica l le-fold increase in eIF2 alpha(P) at 5 min of reperfusion. Our observatio ns contradict hypotheses that PKR activation, loss of eIF2 alpha(P) phospha tase activity, or any general increase in eIF2 alpha kinase activity are re sponsible for reperfusion-induced phosphorylation of eIF2 alpha, and we sug gest that the mechanism may involve regulation of the availability of eIF2 alpha to a kinase. (C) 1999 Academic Press.