PROJECT SUMMARY Alzheimer?s disease (AD) is a societal burden, costing the nation 277 billion dollars per year and is the sixth leading cause of death. As the search for disease modifying therapies for AD continues, a deeper understanding of cellular metabolism in the AD brain is critical to guide research. The apolipoprotein E (APOE) gene is the strongest genetic predictor of late onset AD (LOAD). In humans, there are three major isoforms of apoE: E2, E3, and E4. E3 is the major isoform expressed in humans (~60% of population). E4 confers between a 2 (heterozygous) to 15-fold (homozygous) increase in risk of LOAD compared to E3. Apolipoprotein E (apoE) is primarily secreted by astrocytes as a primary lipid carrier in the brain, and has gained traction as a regulator of cerebral metabolism. For example, E4 carriers exhibit decreased glucose uptake as measured by 18fludeoxy- glucose positron emission tomography (FDG-PET). FDG-PET is similarly lower in AD individuals, however E4 carriers show low FDG-PET decades before the onset of symptomatic cognitive decline. We recently showed that E4 expressing astrocytes accumulate significantly more lipid in the form of lipid droplets (LDs) compared to E3, an intriguing finding in light of the initial 1907 report from Alzheimer that implicated glial lipid accumulation as a hallmark of disease. Since LDs have been shown to be involved in metabolic disease ? including regulation of glucose uptake ? we hypothesize that over accumulation of astrocyte LDs drive the impaired glucose uptake seen in E4 and AD brains. Therefore, this proposal tests a mechanism by which an LD-associated protein sequesters a critical mediator of glucose transporter trafficking, leading to a decrease in astrocyte glucose uptake. We will test this mechanism (Aim 1A) in vivo in mice homozygous for human E3 and E4, and (Aim 1B- C) in vitro in primary astrocytes that express human E3 and E4. We will also translate our findings (Aim 2) by characterizing LD formation in APOE genotyped human post-mortem brain tissue, correlating lipid abundance with clinical data from the UK Alzheimer?s Disease Center cohort. The key focus of this fellowship is to uncover a cellular mechanism driving impaired glucose uptake in E4 astrocytes, as well as equip the trainee with skills in mass spectrometry, confocal microscopy, immunohistochemistry, radioactive tracing, translational human specimen handling, data analysis, and more. These skills and excellent mentorship will be instrumental in the trainee?s path to become an independent physician-scientist.