Validation of Adenylosuccinate as a Novel Endogenous Pro-Angiogenic Factor in the Brain

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Adult brain angiogenesis is required for adaptation to low energy conditions e.g. hypoxia and ischemia, and for recovery after brain injury. Decreased brain angiogenesis may cause brain ischemia, neurodegeneration, and damage during aging while increased angiogenesis is linked to tumorigenesis including glioblastoma among other pathologies. Thus, an understanding of the mechanisms regulating adult cerebral angiogenesis has both basic neuroscience and clinical/translational importance to therapeutic vessel modulation for these diseases. The present basic neuroscience project is designed to investigate a previously unrecognized mechanism for brain angiogenesis regulation through a novel endogenous pro-angiogenic factor, adenylosuccinate (AdSucc). Our preliminary studies have revealed that AdSucc has an important cellular signaling role in the brain rather than functioning just as a metabolic intermediate metabolite. We demonstrated, for the first time, that AdSucc activates angiogenesis in assays both in vitro and in vivo, and that AdSucc induces phospholipase C dependent increase in cytoplasmic calcium, suggesting a receptor dependent action. Both of these effects were activated under AdSucc endogenous concentrations found under low energy conditions in the brain, which are conditions that require angiogenesis for long term compensation. Based on our preliminary data, our central hypothesis is that AdSucc is a brain endogenous pro-angiogenic factor. Our long term objective is to better understand metabolic regulation for brain angiogenesis, and determine if AdSucc is a valid therapeutic target for treating conditions where aberrant angiogenesis is involved. To test our hypothesis, we have established and characterized a novel AdSucc synthase (ADSS) conditional knockdown (KO) mouse model that allows for spatial and temporal control of AdSucc attenuation. We have also established in our laboratory a minimally invasive assay to quantify brain angiogenesis in a living animal in vivo using two photon microscopy. Our central hypothesis will be tested via the following Specific Aims: 1. Determine the role of AdSucc in brain angiogenesis under chronic hypoxia. We will use brain vascular imaging in vivo to determine angiogenesis in wild type and ADSS conditional KO mice under normoxia and chronic hypoxia, and confirm data with histochemical and functional assays. 2. Determine the role of AdSucc in brain angiogenesis under normoxia. In this aim, we will determine if AdSucc induces brain angiogenesis independently of oxygen availability. Successful completion of these aims will test a novel mechanism for regulation of brain angiogenesis.