Exploring new pharmacological strategies to protect brain is the long- term goal, using primarily in vitro models. The plan proposed here is aimed at investigating one avenue by which an endogenous agent, glutamate, can exacerbate toxicity in the acute sequelae of brain damage such as stroke, cerebral ischemia, head trauma, seizures and hypoglycemia. The current proposal's goal is to characterize the pharmacology of calcium-permeable AMPA-preferring subtypes of glutamate receptors in the rat hippocampus as a way of understanding how to protect neurons from such damage. In particular, the mechanism of AMPA receptor- mediated neurotoxicity will be characterized by examining the link between AMPA receptors and intracellular calcium, and a possible second messenger such as nitric oxide. The current research design uses the cultured rat hippocampal slice as an in vitro model that shows selective vulnerability to excitotoxic damage mediated by the AMPA subtype of glutamate receptor. Methods will include assessment of neuronal damage by staining with vital dyes such as propidium iodide, trypan blue and morphological assessment using Nissl stains. The role of calcium in excitotoxicity will be studied using histochemical methods and microfluoro-videometry with the calcium indicators fura-2. Immunocytochemistry will be used to elucidate the relationship between receptor subunit composition and observed localization of function. Three years of postdoctoral training with Dr. David Greenberg at San Francisco General Hospital, University of California has provide an invaluable foundation of expertise in fluorescent techniques and experience in excitotoxicity entirely pertinent to the goals stated here. The environment at SUNY Health Science Center includes superb researchers in pharmacology such as Dr. Robert Wong, Dr. Robert Furchgott and Dr. Peter Bergold, whose expertise is extremely valuable. Support in the form of laboratorIes, equipment and support personnel within the Departments of Neurology and Pharmacology provide the resources needed to successfully complete the proposed projects.
