Abstract This new R21 application is submitted in response to RFA-OD-21-007 (INvestigation of Co-occurring conditions across the Lifespan to Understand Down syndromE (INCLUDE). The overarching goal of this pilot study is to apply and evaluate [18F]-BCPP-EF Positron Emission Tomography (PET) imaging for the in vivo assessment of Mitochondrial Complex I (MC-I) function in individuals with Down syndrome (DS). Impairment or disruption of the electron transport chain (ETC) is common in several neurodegenerative diseases. Research has shown that alterations in mitochondrial ATP production and redox homeostasis (that strongly supports neural function) are associated with DS and Alzheimer's disease (AD) pathogenesis. The Mitochondrial Complex I (MC-I) is the first enzyme complex of the ETC that plays an important role in oxidative phosphorylation and radical oxygen species (ROS) generation in mitochondria, and it is the largest complex in the respiratory chain. As a result, there is much interest in potential therapeutics for DS that target MC-I. [18F]-BCPP-EF has been developed as a high-affinity PET radioligand for in vivo imaging of MC-I activity. Prior [18F]-BCPP-EF studies showed its feasibility for in vivo human imaging and robust test- retest performance in healthy controls. [18F]-BCPP-EF studies in mild AD subjects yielded evidence of: [1] reduced MC-I activity in cortical and subcortical brain areas; [2] mitochondrial-related energy failure preceding glycolysis-related FDG hypometabolism in the parahippocampus; and [3] links between reductions in MC-I activity and hypometabolism, atrophy, and cognitive decline, but not Aβ load. Recent [18F]-BCPP-EF studies provide a strong basis for future MC-I imaging studies in DS. Our primary aim is to perform [18F]-BCPP-EF PET imaging in DS individuals and control participants and map the distribution of MC-I activity throughout DS brain and relative to that observed in control subjects. We hypothesize that MC-I activity will be reduced in cortical and subcortical regions of DS subjects (relative to controls) in a manner that exceeds differences in cerebral atrophy. We will generate quantitative and simplified regional [18F]-BCPP-EF outcomes to evaluate the kinetics of nonspecific uptake, and further evaluate potential reference regions (as it appears that no region is fully devoid of MC- I and cerebellum is an area for which atrophy can be extensive in DS). Maps of MC-I activity will be generated for the DS and controls subjects and corresponding difference maps will be evaluated. Innovation, Impact, Significance: [18F]-BCPP-EF PET imaging in Down syndrome is novel and, to our knowledge, has not yet been demonstrated. This pilot research (and data sharing) has the potential to: [1] provide new in vivo evidence of mitochondrial dysfunction in DS; [2] support future advanced R01 research of mitochondrial energetics in DS; and [3] further inform drug discovery efforts that target MC-I.