Combined effects of high environmental ammonia, starvation and exercise on hormonal and ion-regulatory response in goldfish (Carassius auratus L.).

Due to eutrophication, high environmental ammonia (HEA) has become a frequent problem in aquatic environments, especially in agricultural or densely populated areas. During certain periods, e.g. winter, feed deprivation may occur simultaneously in natural waters. Additionally, under such stressful circumstances, fish may be enforced to swim at a high speed in order to catch prey, avoid predators and so on. Consequently, fish need to cope with all these stressors by altering physiological processes which in turn are controlled by genes expression. Therefore, in the present study, ammonia toxicity was tested in function of nutrient status (fed versus starved) and swimming performance activity (routine versus exhaustive). Goldfish, a relatively tolerant cyprinid, were exposed to HEA (1 mg/L; Flemish water quality guideline for surface water) for a period of 3 h, 12 h, 1 day, 4 days, 10 days, 21 days and 28 days and were either fed (2% body weight) or starved (kept unfed for 7 days prior to sampling). Results showed that the activity of Na⁺/K⁺-ATPase in the gills was stimulated by HEA and disturbance in ion balance was obvious with increases in plasma [Na⁺], [Cl⁻] and [Ca²⁺] after prolonged exposure. Additionally, osmoregulation and metabolism controlling hormones like cortisol and thyroid hormones (T3 and T4) were investigated to understand adaptive responses. The expression kinetics of growth, stress and osmo-regulatory representative genes such as Insulin-like growth factor 1 (IGF-I), growth hormone receptor (GHR), thyroid hormone receptor β (THRβ), prolactin receptor (PRLR), cortisol receptor (CR) and Na⁺/K⁺-ATPase α(3) were examined. Overall effect of HEA was evident since Na⁺/K⁺-ATPase activity, plasma cortisol, Na⁺ and Ca²⁺ concentration, expression level of CR and Na⁺/K⁺-ATPase α₃ mRNA in fed and starved fish were increased. On the contrary, transcript level of PRLR was reduced after 4 days of HEA; additionally T3 level and expression of GHR, IGF-I and THRβ genes were decreased following 10-21 days of HEA. Starvation, the additional challenge in the present study, significantly increased plasma cortisol level and CR transcript level under HEA compared to the fed exposed and control fish. Furthermore, a remarkable reduction in T3 and mRNA levels of THRβ, IGF-I and GHR genes was observed under starvation. The toxic effects in both feeding treatments were exacerbated when imposed to exhaustive swimming with more pronounced effects in starved fish. This confirms that starvation makes fish more vulnerable to external ammonia, especially during exercise.