Traumatic brain injury (TBI) is the leading cause of death among individuals under the age of 45 in the United States and survivors are faced with chronic brain damage leading to debilitating behavioral dysfunction. Brain damage and behavioral dysfunction may be, in part, due to neuronal death following TBI. The hypothesis to be tested in this proposal is that post-traumatic neuronal cell death is a result of activation of the pro-apoptotic caspase family of cysteine proteases. The objectives of this proposal are to elucidate (1) the association between neuronal death in experimental TBI and in postmortem tissue from head-injured patients and the activation of the "executor" caspase-3, (2) whether caspase-3 activation occurs directly as a result of activation of the "initiator" caspase-8, and/or indirectly as a result of mitochondrial pathway which requires Bax translocation, cytochrome c release and caspase-9 activation, (3) whether caspase-8 activation occurs as a result of activation of the tumor necrosis factor family of death receptors, (4) the role of Bax in mediating trauma-induced caspase-9 activation and subsequent caspase-3 activation and cell death, and (5) whether post-traumatic inhibition of caspases-3, -8 and -9 will reduce the extent of injury-induced cell death. Immunoblot and immunohistochemical analyses using specific and selective antibodies will be utilized to temporal and regional patterns of activation of caspases-3, -8 and -9, resdistribution of Bax and cytochrome c, and, apoptotic neuronal damage, as indicated by the presence of cellular DNA fragmentation and morphologic analyses. Mice deficient in TNF will be used to determine the role of TNF in mediating activation of caspases-8 and -3, and eventual apoptotic cell death by using immunoblot, immunohistochemical and histological analyses. The role of Bax in trauma-induced activation of caspase-9 and -3, cytochrome c redistribution, apoptotic neuronal death and behavioral dysfunction will be examined using immunoblot and immunohistochemical techniques, and by testing cognitive and motor function in brain-injured, Bax-deficient mice, followed by histological analysis of cell death. The effect of post-traumatic treatment with peptide inhibitors selective for caspases-3 (DEVD), -8 (IETD) and -9 (LEHD), on the extent of regional cell death and behavioral dysfunction will provide the mechanistic link between caspase activation and TBI-induced pathology.