Research Training Program: Applicant: Dr. Jonathon M. Sullivan is committed to academic medicine and research in the area of brain ischemia and reperfusion. He completed his Emergency Medicine Research Fellowship at Wayne State University, and should complete his Physiology doctoral dissertation within 12 months. Institution: Research training at Wayne State University is strong. It has the nation's largest public graduate school, the third largest medical school, and is ranked in the top 2.5% of all U.S. colleges and universities by the Carnegie Classification """"""""Research University I."""""""" The sponsors of the this application are experienced in studying the pathophysiology of brain ischemia and reperfusion and will provide Dr. Sullivan with extensive training in this area and assure that he has additional training in academic processes such as manuscript preparation, journal manuscript review, grant review for the Emergency Medicine Foundation, didactic presentations in both basic science and clinical disciplines, and clinical teaching. Research Investigation Program: It has been known since 1973 that protein synthesis is inhibited during reperfusion in selectively vulnerable neurons; however, until very recently the mechanisms involved in this were unknown. Our laboratory has found during ischemia calpain-mediated proteolysis of eIF-4G (crucial for cap- binding-dependent translation) and during reperfusion an about20-fold increase in ser-51(P)-eIF-2alpha, which inhibits the generation of eIF-2-GTP that is required for all translation initiation. Our Preliminary Data shows that we have developed an antibody specific for ser-51(P)-eIF-2alpha and that Dr. Sullivan has utilized this antibody to map after 1 hour reperfusion the accumulation of ser-51(P)-eIF-2alpha to vulnerable neurons, found evidence that insulin reduces the accumulation of ser-51(P)-eIF-2alpha in vulnerable hippocampal neurons, and developed a microautoradiographic technique to closely assess new protein synthesis in vulnerable neurons. The hypotheses proposed are that insulin administration during brain reperfusion (1) is associated with dephosphorylation of ser-51 on eIF- 2alpha and (2) will induce restoration of cap-independent translation in selectively vulnerable neurons. Dr. Sullivan will utilize an in vivo model of complete brain ischemia and reperfusion by cardiac arrest and resuscitation for time course histopathologic studies during reperfusion of (I) the location and progression of ser-51(P)-eIF- 2alpha, (ii) the location and progression of general protein synthesis, (iii) the location and progression of synthesis of HSP-70, and (iv) the effects of insulin administered at the onset of reperfusion on these parameters. Examination of the effects of insulin during reperfusion on general synthesis, cap-binding-independent synthesis of HSP-70, and neuronal levels of ser-51(P)-eIF-2alpha could connect the known neuroprotective effects of insulin directly to signal transduction mechanisms affecting translation initiation and message selection.
