Glycolytic compensation rather than NAD⁺/NADH balance sustains neuronal function during mitochondrial stress

Neurons engage compensatory pathways that promote survival when confronted with mitochondrial dysfunction. Indeed, we recently showed that Drosophila neurons upregulate Ldh transcription to help survive the loss of the key mitochondrial fusion gene Opa1. Here, we further characterize this metabolic flexibility and show that it reflects a more general increase in glycolytic activity. A distinct mitochondrial perturbation, TFAM overexpression, similarly induces glycolytic gene expression including Ldh and also elevates lactate levels. LDH is also required to maintain neuronal function under TFAM overexpression. Notably, raising NAD ⁺ /NADH ratio by expressing the bacterial NADH oxidase LbNOX does not substitute for LDH function. On the contrary, it further compromises neuronal function in Opa1 -deficient and TFAM -overexpressing neurons. Moreover, mitochondria-targeted LbNOX expression alone induces mitochondrial dysfunction and the compensatory glycolytic response. Together, these findings indicate that LDH-mediated rescue does not reflect an increase in NAD ⁺ /NADH ratio but is part of a broader neuroprotective metabolic reprogramming which enables neurons to withstand diverse forms of mitochondrial impairment.