Chronic nitrate exposure cause alteration of blood physiological parameters, redox status and apoptosis of juvenile turbot (Scophthalmus maximus)

Nitrate (NO₃⁻) is one of the common inorganic nitrogen compound pollutants in natural ecosystems, which may have serious risks for aquatic organisms. However, its toxicological mechanism remains unclear. In the current study, juvenile turbot (Scophthalmus maximus) were exposed to different concentrations of NO₃⁻ (CK− 3.57 ± 0.16, LN − 60.80 ± 1.21, MN − 203.13 ± 10.97 and HN − 414.16 ± 15.22 mg/L NO₃–N) for 60 d. The blood biochemical assays results revealed that elevated NO₃⁻ exposure significantly increased the concentrations of plasma NO₃⁻, NO₂⁻, MetHb, K⁺, cortisol, glucose, triglyceride, lactate, while significantly decreased the concentrations of plasma Hb, Na⁺ and Cl⁻, which meant that NO₃⁻ caused hypoxic stress and further affected the osmoregulation and metabolism in fish. Besides, exposure to MN and HN induced a significant decrease in the level of antioxidants, including SOD (Point: 60th day, MN, HN v.s. CK: 258.36, 203.73 v.s. 326.95 U/mL), CAT (1.97, 1.17 v.s. 2.37 U/mL), GSH (25.38, 20.74 v.s. 37.00 μmol/L), and GPx (85.32, 71.46 v.s. 129.36 U/mL), and a significant increase of MDA (7.54, 9.73 v.s. 5.27 nmol/L), suggesting that NO₃⁻ exposure leading to a disruption of the redox status in fish. Also, further research revealed that NO₃⁻ exposure altered the mRNA levels of p53 (HN: up to 4.28 folds) and p53-regulated downstream genes such as Bcl-2 (inferior to 0.44 folds), caspase-3 (up to 2.90 folds) and caspase-7 (up to 3.49 folds), indicating that NO₃⁻ exposure induced abnormal apoptosis in the fish gills. Moreover, IBRv2 analysis showed that the toxicity of NO₃⁻ exposure to turbot was dose-dependent, and the toxicity peaked on the 15th day. In short, NO₃⁻ is an environmental toxicological factor that cannot be ignored, because its toxic effects are long-term and could cause irreversible damage to fish. These results would be beneficial to improve our understanding of the toxicity mechanism of NO₃⁻ to fish, which provides baseline evidence for the risk assessment of environmental NO₃⁻ in aquatic ecosystems.