Abstract: Ammonia nitrogen is an important water quality indicator in fish culture, as concentrations exceeding the safe range can cause stress to fish. The study aimed to investigate the effects of high ammonia exposure on the gill tissue structure, immune indexes, and related genes of yellow catfish (Tachysurus fulvidraco) after pre-acclimatization with low ammonia concentration. Yellow catfish (body weight: 3.13±0.49 g) were exposed to low concentration of ammonia nitrogen (4.7mg/L, 10%96hLC₅₀) after pre-acclimated for 3, 7, and 14d, followed by exposure to high ammonia nitrogen concentrations (23.7mg/L, 50% 96hLC₅₀) for 12h and 48h, respectively. The results showed that: (1) after 48h of high ammonia nitrogen stress, the gill structure of yellow catfish displayed various pathological changes such as vacuolization, shortening, curling, and merging of gill lamellae. However, the pre-acclimated groups (3d, 7d, and 14d) experienced less damage compared to the control group (not pre-acclimated to the low concentration of ammonia nitrogen). The control group showed different pathological changes such as vacuolization, gill lamellae shortening, curling, and merging.In contrast, the gill tissues of the pre-acclimated group were less damaged at both 12h and 48h of exposure to the high concentration of ammonia nitrogen. Specifically, the 14-day pre-acclimated group exhibited the least gill tissue damage at these time points. (2) After 48h of high-concentration ammonia nitrogen stress, the LZM activity and IgM, C3 and C4 contents of yellow catfish increased to varying degrees in both pre-adapted and control groups. In all pre-adapted groups, the LZM activity and IgM, C3 contents showed significantly increased, while C4 contents were only significantly increased only in the pre-adapted 7d and the 14d group. At 12h of high-concentration ammonia nitrogen stress, the LZM activity and C3, C4 contents in the 14d group were significantly increased. Additionally, the LZM activity and C3, C4 contents of the 14d group were significantly increased. C3 and C4 contents were significantly higher than those of other groups at 12h of high concentration ammonia stress (P<0.05). (3) The expression levels of tnf-α and c3 genes were significantly down-regulated at 12h of high ammonia nitrogen stress, while in the expression level of il-1β gene showed no significant change. The expression level of igm gene was significantly up-regulated only in the 7d pre-acclimatization group. The above results indicated that pre-acclimatization to low concentrations of ammonia nitrogen, followed by exposure to high concentration of ammonia nitrogen, resulted in reduced gill tissue damage, increased LZM activity, and elevated IgM, C3 and C4 contents in yellow catfish. This suggests that yellow catifish develop a certain tolerance to high concentrations of ammonia nitrogen after pre-acclimatization to low concentration of ammonia nitrogen.