Muscle memory of exercise optimizes mitochondrial metabolism to support skeletal muscle growth.

Exercise protects against age-related declines in skeletal muscle mass and function while improving overall health. Exercise can also prime long-term muscle health to enhance adaptations upon exercise retraining, a phenomenon termed muscle memory that remains largely understudied. To assess how prior endurance training elicits a lasting metabolic memory in skeletal muscle, we used C57BL/6 mice fed either a control (CD) or obesogenic diet [high-fat diet (HFD)] that underwent 4-wk training, detraining, and retraining periods. Our results show that exercise retraining attenuated weight gain and potentiated muscle growth, even with reduced voluntary running volumes. Training increased fiber size [fiber cross-sectional area (fCSA)], which disappeared with detraining and was recovered with retraining regardless of diet, pointing to a glycolytic-to-oxidative fiber shift. Transcriptomic analysis (bulk RNA-Seq) of the retrained muscle revealed a robust enhancement of mitochondrial oxidative phosphorylation (OxPhos) and mitoribosomal genes, paralleled by increases in OxPhos protein complex IV levels, higher long-chain fatty acid oxidative capacity [acyl-CoA dehydrogenase, long chain (ACADL)], and sustained citrate synthase activity 1 wk after retraining, reinforcing the optimization of mitochondrial metabolism. Although transcriptomic evidence revealed a major overlap between HFD- and CD-fed mice, discrepancies in protein abundance emerged, which point to an intricate regulation of mitochondrial programming that supports the muscle memory of growth. Our study identifies common and selective mechanisms by which the muscle memory of exercise overrides dietary challenges and promotes fiber hypertrophy, offering insight into potential mechanisms to leverage to promote healthy aging.NEW & NOTEWORTHY Here we provide evidence that exercise memory in skeletal muscle fine-tunes mitochondrial metabolism to respond to dietary challenges and support muscle growth. Using physiological, RNA sequencing, and biochemical approaches, we show that exercise retraining optimizes mitochondrial metabolism to increase fatty acid oxidative capacity. These findings enhance our understanding of how prior exercise primes muscle for enhanced adaptations, offering insights into strategies to promote healthy aging.