APOE ε4 allele is the strongest genetic risk factor for developing Alzheimer's disease (AD). The proposed project will examine the effect of APOE genotype on cerebrospinal fluid (CSF) DHA levels and on changes in structural and functional brain connectivity in cognitively healthy older individuals in response to DHA supplementation. DHA is an essential omega-3 fatty acid critical to neuronal functions, and is not formed in sufficient amounts de novo. DHA is highly enriched in cortical grey matter and is more concentrated at synapses where it plays a role in synaptic plasticity. DHA is depleted in AD brains. Randomized clinical trials have yielded mixed results on the effect DHA on cognitive outcomes. This study asks the critical question of whether DHA gets into the brain in sufficient amounts after supplementation, and whether APOE genotype affects brain penetrance. Mice carrying the human APOE ε4 allele have decreased brain delivery of DHA compared to mice expressing the APOE ε2 or the APOE ε3 allele. High dose DHA supplementation prevents AD pathology in APOE ɛ4 transgenic mouse models. Our preliminary data indicate lower DHA concentrations in the CSF of cognitively healthy APOE ε4 carriers compared to non-carriers. We hypothesize that APOE ε4 carriers have reduced delivery of DHA to the CSF that can be reversed upon high dose DHA supplementation. To address this hypothesis, we propose a double-blind placebo-controlled clinical trial of high dose (2 grams/day) of DHA over 6 months in 160 cognitively healthy participants stratified by APOE status (ε4 vs. non ε4 carriers). The primary outcome is the effect of APOE genotype on CSF DHA levels in response to DHA supplementation. We will examine red blood cell DHA concentrations as a peripheral biomarker following the 24-week trial. Our secondary outcomes are changes in brain structural and functional connectivity assessed by resting state functional MRI, and changes in cognition. APOE ε4 is associated with blood-brain barrier breakdown, hypolipidated apoE HDL, and brain amyloid deposition. To provide insights into mechanisms regulating DHA brain delivery, our second aim is to examine the association of the change in CSF DHA levels during supplementation with measures of blood-brain barrier integrity (assessed by the CSF albumin quotient), DHA content of apoE particles in CSF, and CSF Aβ42 levels. The results of these studies will provide novel information that can be used clinically to design personalized approaches for the prevention of AD in high-risk individuals. Given the safety profile, availability, and affordability of DHA, refining a DHA intervention in APOE ε4 carriers can have significant impact on reducing AD incidence.