The reactions catalyzed by the delta-5 and delta-6 desaturases (D5D/D6D), key enzymes responsible for highly unsaturated fatty acid (HUFA) synthesis, regenerate NAD from NADH. Here, we show that D5D/D6D provide a mechanism for glycolytic NAD recycling that permits ongoing glycolysis and cell viability when the cytosolic NAD/NADH ratio is reduced, analogous to lactate fermentation. Although lesser in magnitude than lactate production, this desaturase-mediated NAD recycling is acutely adaptive when aerobic respiration is impaired in vivo. Notably, inhibition of either HUFA synthesis or lactate fermentation increases the other, underscoring their interdependence. Consistent with this, a type 2 diabetes risk haplotype in SLC16A11 that reduces pyruvate transport (thus limiting lactate production) increases D5D/D6D activity in vitro and in humans, demonstrating a chronic effect of desaturase-mediated NAD recycling. These findings highlight key biologic roles for D5D/D6D activity independent of their HUFA end products and expand the current paradigm of glycolytic NAD regeneration.