PROJECT SUMMARY Late onset Alzheimer’s disease (LOAD) is a neurodegenerative disease with a multifactorial etiology and intersecting genetic and environmental risks, making it a complex systems challenge. Brain functions are highly energy-dependent, with most of which generated by mitochondria via oxidative phosphorylation. While the association between LOAD and an early decline in brain glucose metabolism and changes in mitochondrial function is well-established, therapeutics that universally enhance brain mitochondrial function have yet resulted in favorable outcomes. As the greatest genetic risk factor for LOAD, the e4 variant of APOE (APOE4) was also found to affect brain bioenergetics and lipid metabolism via incompletely understood mechanisms. Considering the metabolically diverse cellular composition of the brain and APOE as an inter-cellular lipid carrier, we propose that cell type-specific intra-cellular bioenergetic shifts and inter-cellular metabolic uncoupling of fatty acid (FA) metabolism underlie APOE4-driven AD relevant metabolic phenotypes in the brain. Specifically, we hypothesize that APOE4-induced disruption to astrocytic clearance of neuronal FAs and subsequent degradation in astrocytic mitochondria could elicit lipid dysregulation, neuronal dysfunction, neuroinflammation and cognitive decline. Program of research proposed herein will determine the mechanisms, at the cellular level, by which APOE polymorphism alters brain bioenergetics and lipid homeostasis, and eventually LOAD risk. To test our hypotheses, we will implement three levels of investigations to understand the complex mechanisms underlying APOE regulation of metabolic coupling between neuron and astrocytes. Aim 1 will determine and differentiate the effect of APOE isoforms on cellular metabolic shift in neurons and astrocytes using single-cell transcriptomics and in vitro functional assessment. Using a neuron-astrocyte co-culture system, Aim 2 is designed to investigate the impact and mechanism by which different isoforms and origins (neuronal- vs. astrocytic) of APOE affect neuron-astrocyte metabolic coupling, focusing on fatty acid metabolism. Aim 3 will test determine how perturbations to neuron-astrocyte metabolic coupling mediate APOE4-induced LOAD at-risk phenotypes during aging in vivo. Projected outcomes from this research will elucidate how cell types with distinctive bioenergetic phenotypes jointly maintain the brain metabolic homeostasis, and how APOE4 increases risk of LOAD by disrupting the metabolic system of the brain. Translationally, this research will shed light on selective cell vulnerability in AD development and has the potential to identify APOE genotype-specific and cell type- specific therapeutic targets to sustain or restore a bioenergetic equilibrium and lipid homeostasis in the brain that are resilient against synaptic- and cognitive declines in LOAD.