Comparison of effects in sustained and diel-cycling hypoxia on hypoxia tolerance, histology, physiology and expression of clock genes in high latitude fish Phoxinus lagowskii

Phoxinus lagowskii is a popular fish in Chinese cuisine. Though it is found mainly in China's high-latitude regions, where diel-cycling hypoxia (DCH) is known to have unique impacts on aquatic organisms, there is little known about its response to hypoxia. Currently, nothing is known about the changes in blood parameters, gill and liver morphology, glucose and lipid metabolism, or expression of genes involved in clock and glucose metabolism in response to sustained hypoxia (SH) and diel-cycling hypoxia (DCH). To elucidate the influence of sustained and diel-cycling hypoxia on fish hypoxia tolerance, resting oxygen consumption (MO₂) analysis was performed after ten days of hypoxia. This analysis revealed that hypoxia tolerance profoundly improved after ten days of either sustained or diel-cycling hypoxia acclimation, with DCH groups showing greater improvements than SH groups. Additionally, an increase in RBCs was found in P. lagowskii, suggesting an increase in the O₂-carrying capacity of the blood to tolerate hypoxia. Hemoglobin (Hb) concentrations in P. lagowskii were increased at four days of diel-cycling hypoxia, confirming that physiological and metabolic adaptation to hypoxia is based on the duration of O₂ exposure. Increased Hb and hematocrit (Hct) were found in DCH-exposed fish, both of which have been directly linked to high-latitude hypoxia tolerance. In the gills, lamella surface area increased in SH-exposed fish more than DCH-exposed fish, and these increases were accompanied by a decrease in the volume of interlamellar cell mass (ILCM). Histology changes in the liver showed a higher frequency of cytoplasmic vacuolization in DCH-exposed fish. PK increases in SH-exposed fish suggest that fish can use more energy sources in persistent hypoxia. Meanwhile, DCH-exposed fish use TG as an energy source. In SH-exposed fish, self-regulation of Cry1a was observed, whereas Cry1b gene was up-regulated significantly. In DCH-exposed fish, three of eight clock genes studied had increased expression, including Per1a, Clocka, and Cry1b, suggesting that SH and DCH result in different hypoxic responses. This study presents a novel approach to the study of fish responses to hypoxia in high latitude and shows that sustained hypoxia and diel-cycling hypoxia induce large differences in fish physiology.