PARP in focal ischemia

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Poly(ADP-ribose) polymerase (PARP) is an abundant nuclear enzyme which helps to maintain genomic integrity in neurons and numerous other cell types. Inhibition of the enzyme with pharmacological agents or PARP deficiency generated via transgenic paradigms result in reduced injury after cerebral ischemia. Our preliminary studies show that infarction after middle cerebral artery occlusion (MCAO) in PARP null mutants (PARP-/-) is strikingly reduced relative to wild type mice. Aim 1 will fully examine this apparent neuroprotection observed in PARP deficient mice, characterizing extent and longevity of protection in the intact animal over the days of evolving infarction. We will use magnetic resonance imaging (MRI) to non-invasively examine the injured mouse brain and combine these data with functional behavior evaluation over time and terminal histopathology. Aim 2 will establish the importance of nitric oxide generation to PARP activation in vivo and subsequent injury during MCAO. The relative consequences of inhibiting PARP versus neuronal or inducible nitric oxide synthase are examined in the mouse MCAO model. Lastly, using a novel strategy or transfecting virus carrying PARP mutant at two distinct catalytic and cleavage sites, we will begin to examine the molecular mechanism of PARP's important in stroke. The role of the C-terminal NAD binding domain in PARP- mediated neuroprotection will be determined by comparison of stroke outcomes in PARP null mice with or without transfection of a mutant form of PARP with inactive catalytic domain. In addition, we will examine cysteine apartase (CASPASE)-PARP interactions of comparison of stroke outcomes in PARP null mice with and without transfection of a mutant form of PARP lacking in active CASPASE 3 cleavage site (D214G) mutant. These experiments will further our understanding of PARP-mediated mechanisms of in vivo ischemic brain injury.