MICROTUBULAR PROTEOLYSIS IN FOCAL CEREBRAL-ISCHEMIA

Calpain, a neutral protease activated by calcium, may promote microtub ular proteolysis in ischemic brain. We tested this hypothesis in an an imal model of focal cerebral ischemia without reperfusion. The earlies t sign of tissue injury was observed after no more than 15 min of isch emia, with coiling of apical dendrites immunolabeled to show microtubu le-associated protein 2 (MAP2). After 6 h of ischemia, MAP2 immunoreac tivity was markedly diminished in the infarct zone. Quantitative Weste rn analysis demonstrated that MAP2 was almost unmeasurable after 24 h of ischemia. An increase in calpain activity, shown by an antibody rec ognizing calpain-cleaved spectrin fragments, paralleled the loss of MA P2 immunostaining. Double-labeled immunofluorescent studies showed tha t intraneuronal calpain activity preceded evidence of MAP2 proteolysis . Perikaryal immunolabeling of tau protein became increasingly promine nt between 1 and 6 h in neurons located within the transition zone bet ween ischemic and unaffected tissue. Western blot experiments confirme d that dephosphorylation of 7 protein occurred during 24 h of ischemia , but was not associated with significant loss of 7 antigen. We conclu de that focal cerebral ischemia is associated with early microtubular proteolysis caused by calpain.