PROJECT SUMMARY The electron transport chain (ETC) in the inner mitochondrial membrane is essential for efficient ATP production in the cell. ETC function can be disrupted both genetically and pharmacologically by mitochondrial diseases and metabolic poisons, respectively. Inefficient ATP production affects all tissues, but particularly the brain with its high energy demands, thus making this a deadly neurological problem. ETC dysfunction is often associated with buildup of the initial electron donor, NADH, within the mitochondria. This increase in NADH corresponds with a decrease in its oxidized form, NAD+, contributing to an overall NAD+/NADH ratio imbalance. Since NAD+ and NADH are cofactors for many redox reactions in the cell, this results in redox imbalance and reduced metabolic flux, intensifying the energy production problem. Cyanide, a pharmacological ETC inhibitor, and Leigh syndrome, a collection of fatal genetic mitochondrial diseases, both result in inefficient ATP production, NAD+/NADH ratio imbalances, and severe neurological consequences. Few cyanide antidotes exist, all of which have a limited efficacious time window for administration. No cure is available for any form of Leigh syndrome. We hypothesize that normalization of the NAD+/NADH ratio will improve outcomes for both cyanide poisoning and Leigh syndrome. Within Aim 1 of this proposal, we will investigate 1) if restoring NAD+/NADH ratio is sufficient for rescuing cyanide poisoning and 2) what occurs downstream of restoring NAD+/NADH ratio in the context of cyanide poisoning. Within Aim 2, we will determine if redox modulators can rescue disease phenotypes in an array of zebrafish models of Leigh syndrome designed to cover a wide breadth of mutations associated with Leigh syndrome. These disease phenotypes include glial reactivity and deficits in locomotion, oxygen consumption, and associative learning. Collectively, these experiments will evaluate the therapeutic value of restoring the NAD+/NADH ratio for different forms of mitochondrial dysfunction, and will focus on redox modulators that improve neurological outcomes.