Biochemical and multi-omics analyses reveal skeletal muscle adaptation to chronic low-salinity stress of marine medaka (Oryzias melastigma).

Salinity fluctuations are key environmental drivers that shape physiological homeostasis in euryhaline teleosts by altering osmotic balance and energy allocation. In this study, marine medaka (Oryzias melastigma) was used to elucidate skeletal muscle adaptation to chronic low-salinity stress by integrating histology, transmission electron microscopy (TEM), antioxidant indices, transcriptomics, and metabolomics. Offspring (F₁) derived from low-salinity-acclimated parents were reared at a salinity of 2.5‰ and sampled at 4 months, with fish from 25‰ seawater serving as controls. Low salinity significantly reduced the mean cross-sectional area of myofibers while increasing fiber density. TEM revealed a marked widening of the I-band without changes in sarcomere length, suggesting an altered actin-myosin overlap that may help maintain contractile performance under osmotic stress. Biochemical assays showed unchanged catalase activity but significantly elevated total antioxidant capacity (TAC) and malondialdehyde (MDA) levels, indicating enhanced lipid peroxidation accompanied by compensatory activation of the antioxidant defense network. Transcriptome profiling identified 1266 differentially expressed genes (985 downregulated, 281 upregulated), enriched in cytoskeletal and contractile remodeling (desmosomes, myofibrils, myosin filaments) as well as signaling pathways including Wnt/β-catenin and mTOR. Metabolomic analysis detected 112 differential metabolites, including increased POVPC and 3-hydroxycoumarin and decreased uric acid. These metabolites were mainly associated with mTOR signaling, amino acid metabolism, efferocytosis, and the sulfur relay system. Collectively, the results indicate that chronic low salinity shifts skeletal muscle from growth-oriented processes toward structural maintenance, redox regulation, metabolic reallocation, and cellular repair. These findings provide a multi-omics framework for understanding muscle plasticity under hypo-osmotic stress and offer candidate targets for optimizing brackish and freshwater aquaculture practices.