Vision loss affects 1.3 billion people worldwide, with 253 million of those people experiencing vision loss which ranges from severe to total blindness. The etiology of diseases which can cause vision loss are highly complex, as more genes have been identified which can be mutated to cause blindness compared to any other disease. However, many of these genes are involved in metabolic processes. This suggests that it is imperative to have a complete understanding of metabolism in a healthy retina so we can better understand how to treat it when it becomes dysregulated in disease. One aspect of retinal metabolism which seems to be very important for maintaining vision is a high level of aerobic glycolysis performed by photoreceptors. However, we are still far from understanding how photoreceptors maintain this high rate pyruvate to lactate conversion since they also contain functional and underused mitochondria which could use pyruvate to more efficiently make ATP. In our preliminary investigations into this question, we have observed that photoreceptors use GAPDH to maintain a uniquely high cytoplasmic NADH/NAD+ ratio and that high levels of NADH could be sufficient to drive lactate production. However, previously published results have shown that GAPDH is not overexpressed in photoreceptors, so it is still unclear what is driving its high activity.
In aim 1 of this proposal, I will address how retinas maintain a uniquely high cytoplasmic NADH/NAD+ ratio by performing a comparative analysis of flux through each step in glycolysis between retinas and oxidative tissues. I predict that I will identify steps early in glycolysis which are rate-limiting for flux through GAPDH and thus cytosolic NADH production in oxidative tissues, but not in retinas.
In aim 2 of this proposal, I will use an alternative oxidase to specifically lower the cytoplasmic NADH/NAD+ ratio in cone photoreceptors, which will allow us to address both the importance of cytoplasmic NADH for maintaining aerobic glycolysis, as well as the role of photoreceptor lactate production on the health of the retinal ecosystem as a whole.
Aerobic glycolysis is key to maintaining photoreceptor health and function, yet how photoreceptors convert pyruvate to lactate instead of using it to more efficiently make ATP in oxidative phosphorylation is still unknown. A high cytosolic NADH/NAD+ ratio could explain how pyruvate is converted to lactate before it has a chance to be oxidized in mitochondria, but a causative link between a high NADH/NAD+ ratio and aerobic glycolysis has not yet been shown. This project proposes the use of well-validated metabolic flux methods in order to identify how glycolytic retinas but not oxidative tissues maintain a high cytoplasmic NADH/NAD+ ratio, and how photoreceptor metabolism is altered when this ratio is lowered.