Omega 3 fatty acids, acute neuroprotection via mitochondria

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Project Summary Neonatal stroke and hypoxia-ischemia (HI) brain injury remains a leading cause of a lifelong neurological handicap. The only currently accepted approach is post-ischemic hypothermia. However, no exact therapeutic mechanisms of this approach are known. A growing body of experimental evidence showed that omega-3 (n-3) fatty acids (FAs) and their bioactive metabolites protect developing brain against HI-reperfusion injury. Using triglyceride emulsions we showed that docosahexaenoic acid (DHA) provided strong neuroprotection against HI brain injury in different rodent models. This neuroprotection was associated with increased DHA content in cerebral mitochondria, preserved mitochondrial function in reperfusion and coupled with a dramatic (3 to 15 fold) elevation of DHA metabolites, neuroprotectin D1 (NPD1) and other specialized pro-resolving mediators (SPMs), specifically in the ischemic brain. We have reasoned that beneficial action of DHA may be exerted by its metabolites. Among tested SPMs, only NPD1 significantly attenuated HI brain damage in mice. Administration of NPD1 also suppressed activation of Ca2+ induced mitochondrial membrane permeability transition pore (mPTP) and apoptotic death pathway by blocking BAX-mitochondria interaction. In this renewal application, we hypothesize that DHA-derived bioactive mediator, NPD1 a) modifies the matrix mitochondrial membrane properties that attenuate activation of mPTP, b) prevents mitochondrial outer membrane interaction with BAX. These effects attenuate post-ischemic secondary energy failure and mitochondrial apoptotic cell death pathway, contributing/explaining the neuroprotective mechanisms afforded by DHA. This hypothesis will be tested in three specific aims. Aim 1: To determine the role of exogenous NPD1 and DHA in attenuation of BAX-driven mitochondrial cell death following HI brain injury. In this aim, we will compare the potency of DHA or NPD1 in limiting BAX-mediated cellular death. Given that both compounds demonstrated beneficial changes in mitochondrial functions (Ca2+ buffering and respiration (DHA) and Ca2+ buffering (NPD1)) after HI insult, we were focused on determining the fate of exogenous DHA and its metabolite, NPD1 by tracing deuterated 2H10-DHA in the ischemic brain, and mitochondria/mitoplasts isolated from the ischemic hemisphere. Aim 2: To determine the origin of increased brain NPD1 after acute injection of DHA and distribution of NPD1 and exogenous DHA following HI injury). Finally, in Aim 3, we will determine potential actions of NPD1 or/and DHA on preserving an integrity of the inner mitochondrial membrane, directly testing electrical conductance driven by the presence or absence of permeability transition. Our proposal is a translational study with a focus on specific mechanisms of neuroprotection targeting post-ischemic mitochondria using innovative approaches toward an understanding a temporal role of mitochondrial permeability transition in secondary energy failure and cell injury. Translational impacts are defined by the neuroprotective strength and clinical utility of DHA metabolites, rather than DHA itself.