Daily rhythms of acute stress responses and antioxidant systems in the European sea bass (Dicentrarchus labrax): Effects of the time of the year

Fish reared in aquaculture face various acute stressors, including air exposure during handling. Research on the stress response in fish can provide essential insights into their physiology and help define better aquaculture practices. In this study, we investigated the daily rhythms in the stress-axis response of the European sea bass (Dicentrarchus labrax) subjected to an acute stressor consisting of air exposure (1 min), and how this response is influenced by the time of the day and the season of the year. In addition, rhythms in antioxidant systems were also assessed. The experiments were performed in late Autumn (December) and late Spring (June), with natural photoperiod (10 L:14D and 15 L:9D, respectively) and water temperature (ranging from 19.47 ± 0.17 December to 22.13 ± 0.13 ◦ ◦ C in C in June). Samples were collected throughout a 24-h cycle at Zeitgeber time (ZT) 0.5, 4, 7.5, 12, 16, 20, and 24.5 h at both seasons. At each sampling point, an untreated control (CTRL) group was sampled, while a STRESS group was exposed to air for one minute, returned to the tank, and sampled one hour later. Fish were sacrificed to collect plasma samples, hypothalamus and liver. Plasma samples were analyzed for cortisol, glucose, and lactate. In the hypothalamus, the mRNA expression levels of corticotropin-releasing hormone (crh) and crh-binding protein (crh-bp) were quantified using quantitative RT-PCR (qPCR). In the liver, genes related to antioxidant systems (catalase, superoxide dismutase 1, glutathione peroxidase,and glutathione reductase) and mitochondrial markers of stress (uncoupling protein 1, cytochrome c oxidase IV and peroxiredoxin3) were also analyzed by qPCR. The results revealed that most stress indicators (cortisol, cat, sod1, gsh-px, gsr, ucp1, coxIV) displayed daily rhythms. Furthermore, the stress response was significantly influenced by the time of day and the season in which the stressor was applied. In June, cortisol and glucose responses to stress were higher during the day than at night. The increase observed after stress in genes related to the antioxidant system was more significant in June than in December. Conversely, the response of mitochondrial markers was greater in December. Taken together, these findings highlight that the stress response of the European sea bass is time- dependent, both on a daily and a seasonal basis. This emphasizes the importance of considering cyclic environmental factors and circadian rhythms in aquaculture procedures to enhance fish welfare.