Physiological responses of Perccottus glenii to cold stress, freezing, and recovery

Using the cold-tolerant fish Perccottus glenii as the experimental model, this study systematically investigates its physiological responses throughout the entire process of low-temperature stress, freezing, and post-thaw recovery. Furthermore, it elucidates the regulatory mechanisms of melatonin and its receptor-mediated signaling pathways during the recovery phase. The results indicate that during the low-temperature and freezing stages, hepatic glycogen and mitochondrial cristae decrease, while lipid droplets increase. Plasma glucose and glycerol levels, as well as the activities of enzymes related to glycolipid metabolism and antioxidant processes, significantly increase. Cold stress primarily activates glucose and phospholipid metabolic pathways, whereas freezing upregulates genes involved in circadian rhythm, oxidative phosphorylation, and lipid synthesis. During the recovery phase, the number of endoplasmic reticula increased, and immune and anti-apoptotic pathways are activated. In the post-thaw recovery process, melatonin exhibits temporal expression patterns and regulates autophagy, endoplasmic reticulum stress, mitochondrial function, and antioxidant capacity in the liver and brain in a time-dependent manner. In the early phase of recovery, melatonin activates autophagy and enhances mitochondrial activity and antioxidant levels; however, these parameters decline in the later phase. Exogenous melatonin inhibits endogenous melatonin synthesis. In the late recovery phase, significant changes are observed in the expression of key melatonin synthesis enzymes, melatonin receptors, and multiple signaling pathway molecules, including PKC, ERK, PKA, and AKT. In vitro hepatocyte experiments confirm that melatonin receptors regulate PKC expression. In summary, P. glenii demonstrates differentiated adaptive strategies in response to varying low-temperature environments. Melatonin, via its receptor-mediated signaling network, plays a crucial role in the post-thaw physiological repair process. This study offers significant theoretical support for understanding the mechanisms of cold tolerance in fish, as well as implications for animal cryopreservation and related medical research.