Excessive water temperature rise increases the energy consumption of the fish and thus requires additional energy supply, thereby increasing the cost of growth in fish (Little et al., 2020).
The formation of ROS and the increase in metabolic demand under the rise of CO2 in seawater as well as water temperature generally lead to damage caused by peroxidation and increased activity of antioxidant enzymes, and additional energy consumption is unavoidable (Rosa et al. 2016). Contrarily, in the case of a species with high adaptability (resilient) to environmental stressors, it also prevents cell damage (lipid peroxidation) by upregulating the action of antioxidant enzymes to cope with oxidative stress caused by warming (Sampaio et al. 2018).
The complex environment of ocean acidification and warming generates ROS and antioxidant enzymes act to remove ROS, but excessive production of ROS exceeds the action capacity of antioxidant enzymes, so it can have a negative effect on fish. Araújo et al. (2018) found that the effects of ocean acidification and the combined effects of two complex environments (ocean warming and acidification) have negative synergistic effects such as increased energy demand, anaerobic metabolism, and impaired proteolysis in sea bream Sparus aurata. It is reported to induce cytotoxic effects and pose a serious threat to sea bream populations. Rosa et al. (2016) reported that when juvenile bamboo sharks Chiloscyllium punctatum were exposed to a combined environment of ocean warming and acidification, antioxidant enzymes detoxified the ROS. However, upregulation of these antioxidant enzymes was insufficient to minimize the increase in cholinergic neurotransmitters caused by peroxidative damage and stress response in the brain.
In our study, it was found that under the warming conditions (higher water temperature), the stress response and the activity of antioxidant enzymes were high, and the appearance of apoptosis was also high. This findings are consistent with that of a similar study by Madeira et al. (2016) who reported that excessive oxidative stress induced in the muscles, liver, and brain of Gilt-head bream Sparus aurata at high water temperature (30°C) increased the activity of antioxidant enzymes (SOD, CAT, GST) to counteract the effect of increasing ROS. Additionally, it was similar to the results of Kim et al. (2016) who reported that a high water temperature environment increased the antioxidant enzyme activity of olive flounder and induced apoptosis in liver tissue by increasing the activity of CASP3. Additionally, in our study, the stress response, the antioxidant enzyme activity, and the appearance of apoptosis were found to be high in the combined warming (high water temperature) and acidification (pH decrease) conditions, similar to the overall warming conditions. This result was confirmed in a study by Rosa et al. (2016) and Araújo et al. (2018) who state that the combined effect of the two environments appears to act negatively to induce cytotoxicity, which results in increased emergence of apoptosis. Specifically, this implies that the antioxidant enzyme to remove ROS acted not just in a high temperature environment but also in a complex environment with warming (high water temperature) and acidification (pH decrease), although the appearance of apoptosis was not inhibited because the action capacity of the antioxidant enzyme exceeded due to a large amount of ROS.
The activity of CAT in our study was consistently high under the conditions in which warming and acidification were combined. This is described in Carney Almroth et al. (2019) who state that Atlantic halibut Hippoglossus maintained a significant correlation of high SOD and CAT activity in a warming (high water temperature) environment but not in a high CO2 concentration (low pH level) environment. Fish may have increased amounts of hydrogen ions (H+) in their body to normalize the pH levels around blood cells by increasing HCO3− in an environment exposed to CO2 (Heuer and Grosell, 2014). This excess presence of H+ in the blood can combine with H2O to form H2O2 (Sampaio et al., 2018), which is believed to induce a sustained increase in CAT, the key enzyme that generally scavenges H2O2.
The results of this study under the combined environmental conditions of warming and acidification, stress and antioxidant enzyme activity, and apoptosis all tend to be high at 20°C. Grans et al. (2014) found that warming and acidification conditions increased oxygen demand and cardiac performance in Atlantic halibut Hippoglossus hippoglossus, although these increases were not interpreted as growth improvements. Instead, it was reported that Atlantic halibut was more affected by acidification under lower water temperature conditions than the optimum growth water temperature, and hence the growth decreased. When cells are exposed to oxidative stress, ROS is also generated in the electron transport chain of mitochondria, but considering the generation of ROS due to increased respiration in a high-temperature environment, in our study, the effects of acidification under the combined warming and acidification conditions do not exceed the effects of warming. Rather, the effect of acidification appears to be greater in low-temperature than in high-temperature conditions.
Summarizing the results of our study on juvenile olive flounder, 1) Oxidative stress and apoptosis tend to increase under both the warming (high water temperature), combined warming (high water temperature), and acidification (pH decrease) conditions. 2) Considering the increase in ROS due to oxidative stress as well as the increase in ROS generation due to increased respiration in the high water temperature condition, the effect on acidification in the combined warming and acidification condition does not seem to exceed the effect of warming. 3) It is implied that the effect on acidification is greater at a relatively low water temperature (20°C) than at a high temperature condition.
Excessive induction of oxidative stress due to warming and acidification may lead to a decrease in the growth of fish, induce disease due to reduced immunity, and shorten their lifespan. Therefore, understanding the physiological responses of marine organisms in combination with the prediction of future changes in the marine environment is important for predicting the direct impact of climate change on marine organisms and their vulnerability. The data thus obtained can also be very important for predicting the sustainability of the fishery and aquaculture industries. Additionally, in the mid-latitude regions where Korea is located, seawater temperature fluctuates greatly in many cases due to seasonal changes. Therefore, we suggest that additional research on oxidative stress and antioxidant responses of marine organisms is needed in an environment in which low water temperature and acidification conditions coexist in winter, which cannot guarantee sufficient nutritional status due to poor food intake.