EXCITOTOXICITY IN CIRCULATORY ARREST--BRAIN INJURY

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DESCRIPTION (Verbatim from Applicant's Abstract): The goal of the project is to define the mechanisms of excitotoxic neuronal injury caused by hypothermic circulatory arrest (HCA) and to develop the means to prevent it. In our canine survival model of HCA, replicating clinical experience during cardiac operations, dogs subjected to 2 hours of circulatory arrest at 18degreesC sustain a consistent neurologic deficit and histologic pattern of selective neuronal death. We originally showed that administration of selective glutamate receptor antagonists before and after HCA-induced injury reduced the neuronal necrosis. We have now shown that neuronal death can occur by apoptotic or necrotic mechanisms. We showed that glutamate release after HCA results in accumulation of nitric oxide (NO), which mediates neuronal death and that inhibition of neuronal nitric oxide synthase (NOS) reduces production of NO in the brain and prevents apoptosis. We hypothesize that mitochondrial dysfunction determines the mechanism of delayed excitotoxic neuronal injury after HCA by apoptosis or necrosis and that neuronal apoptosis can be prevented by ischemic preconditioning (IPC), achieved pharmacologically by opening ATP-dependent potassium channels on the inner mitochondrial membrane. We further hypothesize that NO may act as a mediator both of neuronal injury and neuronal protection by IPC. In preliminary experiments we have shown that diazoxide, an ATP-dependent potassium channel opener can produce pharmacologic IPC and that that this agent can prevent apoptosis in cardiomyocytes acting on the inner mitochondrial membrane. In our canine model, diazoxide has shown near total elimination of neurologic deficit following HCA, with reduction in apoptosis in select neuronal populations. We also showed that hypoxia can activate HIF-1 with induction of iNOS and production of NO, a putative molecular pathway of the late form of IPC. We propose to: (1) measure metabolic indicators of mitchondrial dysfunction in specific brain regions using 'H and 'P MRSI following HCA in our canine model and (2) correlate these with neuronal survival, apoptosis and necrosis; (3) to examine pharmacologic IPC with diazoxide as means of neuroprotection and (4) determine how opening the potassium channels in the mitochondrion alters its function to effect IPC; (5) to establish how NO can act as mediator of both injury and protection in the brain. This research will expedite the clinical use of inhibitors in the excitotoxic cascade and facilitate exploitation of IPC by pharmacologic means to provide cerebral protection resulting in better patient outcomes after HCA in cardiovascular operations.