higher density and rearing them with a high-protein diet. This scenario, along with limited renewal of clean water (e.g., in recirculatory and pond-farming), eventually results in a build-up of toxic ammonia in the water. Within the aquaculture system, there is no fast remedy to alleviate high ammonia-induced toxicity, and prevention rather than correction should be the focus.
Therefore, we tested whether pre-acclimation to a low concentration of ammonia could enable the fish to develop an “ammonia stress-avoidance” memory, enhancing their tolerance to subsequent sub-lethal ammonia threats. To test this, we selected channel catfish (Ictalurus punctatus), a key species in the U.S. aquaculture industry, as the model organism. Catfish were pre-exposed to 2.17 mg/L (total) ammonia (10% of determined 10-day LC50) for 14 and 21 days. Thereafter, each of these pre-exposed and parallel control (without pre-exposure “naïve”) groups were recovered for 7 days in clean water. Following this recovery phase, each group was subsequently exposed to sub-lethal 12.2 mg/L ammonia (~25% 96 h LC50) for 14 days. Plasma ion analyses revealed that 14-day pre-exposed fish maintained more stable levels of Na⁺, K⁺, Ca²⁺, Mg²⁺, and Mn⁺, indicating improved ion balance, whereas 21-day pre-exposed and naïve fish exhibited greater ionic disturbances. Correspondingly, gill ion-transporting enzymes demonstrated higher activities of Na⁺/K⁺-ATPase and H⁺-ATPase, in the 14-day pre-exposed group, supporting enhanced ion exchange and acid-base regulation during subsequent ammonia stress. Energy reserves followed this pattern: 14-day pre-exposed fish
preserved glycogen, protein, and lipids under stress, whereas naïve and 21-day groups showed faster depletion, indicating higher metabolic costs. Gene expression profiling revealed that ammonia transport-related genes Rhbg and Rhcg were significantly upregulated in the gills of 14 days of pre-exposed fish, (Fig. 1), suggesting an adaptive transcriptional adjustment to enhance ammonia excretion. In summary, 14-day pre-acclimation effectively trains channel catfish to withstand ammonia toxicity by enhancing survival, preserving ion balance, sustaining energy metabolism, and regulating excretion-related genes. Longer exposure (21 days) offered no added benefits. Overall, we suggest that strategic pre-conditioning can be a promising approach to mitigate ammonia toxicity in aquaculture systems, with 14-day pre-exposure emerging as the optimal duration for channel catfish.