Calpatatin in brain contusion and axonal injury: a novel therapeutic approach

Calpains are activated acutely and for prolonged periods in regions of axonal injury and neuronal cell death after traumatic brain injury (T81), despite colocalization of calpains with their endogenous inhibitor, calpastatin. While little is known about the role of calpastatin in the traumatically injured brain, work in other CNS injury paradigms suggests that endogenous calpastatin expression may be insufficient to adequately inhibit excessive or prolonged "pathological" calpain activation. Therefore, we hypothesize that overexpression of neuronal calpastatin will attenuate calpain activation initiated by contusive brain injury or traumatic axonal injury, leading to neuroprotection and improved functional outcome. We propose to test this hypothesis using transgenic (Tg) mice that overexpress human calpastatin (hCAST) in CNS neurons, resulting in a 3-fold enhancement of calpastatin activity. In parallel, we will utilize calpastatin-deficient (CAST -/-) mice to probe the inherent role of calpastatin in the pathology of contusion and axonal injury. The coordinated evaluation of the role and therapeutic potential of calpastatin, in two models of traumatic injury is motivated by the increasing appreciation that the pathobiology of contusion (gray matter) injury and white matter injury (e.g. diffuse axonal injury) may be distinctly different, requiring tailored therapeutic strategies. Specifically, we will quantify the extent of acute cortical and hippocampal spectrin proteolysis by ca/pains, measure subacute cortical and hippocampal cell death, and monitor cognitive and motor function in the days and weeks after controlled cortical impact brain injury to hCAST Tg and CAST _/_ mice (Aim 1). We will then evaluate the temporal course of cytoskeletal proteolysis and assess impairment of retrograde axonal transport in optic nerves from hCAST Tg and CAST -/- mice subjected to optic nerve stretch injury (Aim 2). The proposed studies will, for the first time, establish a functional role for caJpastatin in the pathology of T81 and will provide guidance for the development of new therapeutic approaches focused on (a) directly or indirectly enhancing calpastatin expression or activity or (b) mimicking the actions of calpastatin through novel synthetic peptides.