PROJECT SUMMARY Impaired brain energy metabolism is increasingly recognized as an important target mechanism for Alzheimer’s disease (AD) prevention and treatment. Increasing evidence suggests that manipulating brain energy substrates improves brain health in aging and may be therapeutic in AD through a metabolic shift of the brain. Particularly, the ketogenic diet (KD) shows substantial effects on improving clinical outcomes, including cognitive function, in AD patients, by shifting the major brain energy substrate from glucose to ketones. However, the underlying mechanisms for such approaches are not well understood mainly due to a lack of non-invasive methods to directly assess brain bioenergetics in humans, limiting further progress in developing effective intervention strategies for AD. Thus, we designed a randomized clinical trial to overcome current limitations through the development of advanced 1H and 31P magnetic resonance spectroscopy (MRS) techniques to measure neuroenergetic biomarkers with 3T clinical scanners and to use the developed techniques to test our hypothesis that the KD significantly enhances brain energy metabolism in aging and AD. During the R61 developmental phase, we will develop techniques to measure 1) primary circulating ketone produced in response to a KD, b-hydroxybutyrate (BHB); 2) bioenergetic co-enzymes involved in both cytosolic and mitochondrial metabolism, NAD, NAD+, NADH; 3) the primary energy-carrying molecules, adenosine triphosphate (ATP) and phosphocreatine (PCr) in AD-relevant brain regions. During the R33 Basic Experiment Study involving Humans (BESH) trial, we will investigate the physiological effects of the KD on brain energy metabolism in aging and AD by measuring the 1H/31P MRS neuroenergetic biomarkers that we develop during the R61 phase. Both AD patients and older adults with normal cognition will be randomized to either the 4-week, experimentally-controlled feeding of the KD or therapeutic lifestyles changes (TLC) diet. In Aim 1, we will quantify the degree of brain energetic manipulation via the KD in aging and AD using neuroenergetic imaging. In Aim 2, we will elucidate the association between brain energetic manipulation and brain antioxidant status in aging and AD using antioxidant (GSH) imaging. In Aim 3, we will determine the effect of aging and AD on neuroenergetic status by utilizing baseline data from participants in both R61 and R33 phases (30 healthy adults, 30 older adults with normal cognition, 30 AD patients). We anticipate that this project will provide proof-of-concept that the KD truly manipulates and enhances neuroenergetics in aging and AD for the first time by directly assessing this widely hypothesized mechanism with in vivo neuroimaging biomarkers in humans. The infrastructure established through the R61 phase will provide a critical platform for AD intervention trials that can be extended across the NIA’s network of Alzheimer’s Disease Research Centers (ADRC) for us...