Project Summary This application is an Administrative Supplement to 2R01HL103866, entitled "Gut flora metabolism of dietary carnitine and cardiovascular disease" (NOT-AG-20-034 on Alzheimer’s-focused administrative supplements for NIH grants that are not focused on Alzheimer’s disease). The overall proposal is predicated upon the recognition that gut microbiota contribute to both the development and adverse consequences associated with Cardiovascular Disease (CVD), other chronic inflammatory diseases and the recent findings of elevated TMAO levels in the CSF of aging subjects with Alzheimer’s disease (AD), as well as those with mild cognitive impairment (MCI). Elevated TMAO levels have also recently been reported in mouse models of AD. Yet a direct role for gut microbiota-generated TMAO in susceptibility for development of MCI or AD is lacking. Gut microbe- dependent generation of trimethylamine (TMA), and its conversion in the host liver to trimethylamine N-oxide (TMAO), are recognized for being mechanistically and clinically linked to the development of CVD. In prior work we identified microbial participants in conversion of dietary choline and phosphatidylcholine into TMA and TMAO, mechanisms through which TMAO contributes to CVD, and approaches for intervening and suppressing TMAO generation from choline. However, performance of those studies also revealed that microbial TMA/TMAO generation in some subjects, particularly omnivores, can originate from dietary carnitine. Moreover, the microbes and microbial enzymatic machinery involved in gut microbiota-dependent TMAO elevation from either dietary carnitine, or a diet enriched in red meat, are distinct from the microbial participants in choline/phosphatidylcholine conversion into TMA/TMAO. The overarching goal of this administrative supplement is to test whether gut microbial metabolism of dietary trimethylamines and TMAO generation enhances risk for MCI and AD, and whether therapeutic interventions for the TMAO-generating pathway have potential promise for AD. The proposed research studies are multidisciplinary, and encompass a combination of basic, translational and human clinical investigations. In Aim 1 we will define the relevance of the metaorganismal choline TMA TMAO and carnitine gamma-butyrobetaine (gBB) TMA TMAO pathway to MCI and AD through clinical association studies. In Aim 2, we will explore a therapeutic approach for inhibiting dietary choline-dependent TMA and TMAO generation, and relevant behavioral phenotypes in an animal model of AD. Successful completion of the proposed studies will further reveal a potential role for the gut microbial TMAO pathway in MCI and AD development. They also will enable potential advances in both diagnostics and therapeutic approaches for MCI and AD.