Survival and acute osmoregulatory response of grass carp under salinity stress

Inland saline waters, widely distributed globally, impose stringent constraints on aquaculture owing to their high osmotic pressure, preventing most aquatic organisms from surviving. As the most extensively farmed freshwater fish worldwide, the grass carp (Ctenopharyngodon idella) is confined to freshwater habitats. The adaptation of this species to saline aquaculture holds significant ecological and economic potential; however, the mechanisms underlying its osmotic regulation in hypersaline environments remain poorly understood. In this study, juvenile grass carp with a mean body weight of 15.42 ± 0.96 g were subjected to acute salinity tolerance tests across six gradients (0, 4, 7, 10, 13, and 16 ppt). The 96 h median lethal salinity concentration (LC50) was first determined, followed by the evaluation of osmoregulatory dynamics through physiological-biochemical profiling and targeted quantification of salinity-responsive gene expression under acute stress conditions. Results showed mortality exhibited salinity-dependent escalation, and the LC50-96 h was 10.58 ppt. Under 24 h salinity exposure (0, 4, 7, and 10 ppt), grass carp exhibited salinity-dependent increases in serum electrolytes (Na⁺, Cl^-, K⁺) and cortisol (P < 0.05). Serum osmolality remained stable at 4 ppt and 7 ppt but increased significantly at 10 ppt. Serum prolactin showed progressive decline from 7 ppt. Superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) activities in the liver, kidney, and gills showed salinity-stimulated upregulation (significant at 7 and 10 ppt). Immune-related enzymes, acid phosphatase (ACP) and alkaline phosphatase (AKP), increased at 4 and 7 ppt, whereas AKP activity declined at 10 ppt. Gill Na⁺/K⁺-ATPase (NKA) activity was significantly elevated at 10 ppt (P < 0.01). Moreover, both gill and kidney structures exhibited significant alterations under 7 ppt and 10 ppt salinity stress. Specifically, the gill showed cracks in the filaments, chloride cell hyperplasia, and detachment of flattened epithelial cells; renal tubules were atrophic. The expression of ion transport-related genes NKA and solute carrier family 12 member 2 (SLC12A2) in the gills increased with increasing salinity; both genes showed significant differences at 7 ppt and 10 ppt, indicating their role in regulating osmotic pressure balance. This study provides a theoretical basis for the saline aquaculture technology and salt-tolerant variety development in grass carp.