Aerobic glycolysis accounts for approximately 80%-90% of glucose used by adult photoreceptors (PRs); yet, the importance of aerobic glycolysis for PR function or survival remains unclear. Here, we further established the role of aerobic glycolysis in murine rod and cone PRs. We show that loss of hexokinase-2 (HK2), a key aerobic glycolysis enzyme, does not affect PR survival or structure but is required for normal rod function. Rods with HK2 loss increase their mitochondrial number, suggesting an adaptation to the inhibition of aerobic glycolysis. In contrast, cones adapt without increased mitochondrial number but require HK2 to adapt to metabolic stress conditions such as those encountered in retinitis pigmentosa, where the loss of rods causes a nutrient shortage in cones. The data support a model where aerobic glycolysis in PRs is not a necessity but rather a metabolic choice that maximizes PR function and adaptability to nutrient stress conditions.

Vectors based on the adeno-associated virus (AAV) are currently the preferred tools for delivering genes to photoreceptors (PR) in small and large animals. AAVs have been applied successfully in various models of PR dystrophies. However, unknown barriers still limit AAV's efficient application in several forms of severe PR degenerations due to insufficient transgene expression and/or treated cells at the time of injection. Optimizations of PR gene therapy strategies will likely benefit from the identification of the cellular factors that influence PR transduction. Interestingly, recent studies have shown that the AAV transduction profile of PRs differs significantly between neonatal and adult mouse retinas after subretinal injection. This phenomenon may provide clues to identify host factors that influence the efficiency of AAV-mediated PR transduction. This study demonstrates that rod outer segments are critical modulators of efficient AAV-mediated rod transduction. During retinal development, rod transduction correlated temporally and spatially with the differentiation order of PRs when vectors were introduced subretinally but not when introduced intravitreally. All subretinally injected vectors had an initial preference to transduce cones in the absence of formed rod outer segments and then displayed a preference for rods as the cells matured, independently of the expression cassette or AAV serotype. Consistent with this observation, altered development of rod outer segments was associated with a strong reduction of rod transduction and an increase in the percentage of transduced cones by 2- to 2.8-fold. A similar increase of cone transduction was observed in the adult retinal degeneration 1 (rd1) retina compared to wild-type mice. These results suggest that the loss of rod outer segments in diseased retinas could markedly affect gene transfer efficiency of AAV vectors by limiting the ability of AAVs to infect dying rods efficiently. This information could be exploited for the development of more efficient AAV-based PR gene delivery procedures.