Effect of ammonia to fish neutrophils and its permissible level

将大肠杆菌溶液（大肠杆菌，5mg/ml） 0.2ml注射到罗非鱼鱼鳔中，并在25‰的温度下注射。注射后24小时，用注射器将渗出液从膀胱腔中抽出。液体中含有大量白细胞，其中中性粒细胞占97%。这种中性粒细胞的分离也适用于其他淡水和海水鱼类。在中性粒细胞-汉克斯平衡盐水溶液中加入浓度为0,1和2ppm的氨（NH4+。0,0.011和0.023ppm为NH3）。在体外氨毒性实验中，氨抑制了中性粒细胞的吞噬能力。在体内实验中，罗非鱼在25℃的氨水中（NH4+分别为0、1、2ppm， NH3分别为0、0.003、0.006ppm）保存一周。分别在第0、1、3、7天分离中性粒细胞进行生理分析。在本试验中，中性粒细胞的生理不受环境氨浓度的影响。在整个试验期间，暴露在含氨水中的鱼的血氨水平几乎相同，尽管水中氨浓度不同。这一结果可能是在氨水中饲养的鱼对中性粒细胞生理没有影响的原因。在体内氨毒性试验过程中，我们经常看到优势鱼对从属鱼的攻击行为。与优势鱼相比，从属鱼的中性粒细胞功能明显下降。由于这一行为方面在鱼类免疫学研究中一直被忽视，免疫学家应进一步发展。

0.2ml of bacterial solution (E.coli, 5mg/ml) was injected into a swim bladder of tilapia under 25ﾟC-WT.At 24 hrs post-injection, the exudative fluid was withdrawn from the bladder lumen by a syringe. The fluid contained large numbers of leucocytes, in which neutrophils occupied >97%. This neutrophil isolation was also available to the other fresh- and seawater fishes.Ammonia was added to the neutrophils-Hanks balanced saline solution at the concentration of 0,1, and 2ppm (as NH4+.0,0.011, and 0.023ppm as NH3). In this in vitro ammonia toxicity, the ammonia depressed the phagocytic ability of neutrophils. For in vivo test, tilapia was kept in ammonia water (0,1, and 2ppm as NH4+, 0,0.003, and 0.006ppm as NH3) for one week under 25ﾟC.The neutrophils were isolated to analyze their physiology in the 0,1st, 3rd, and 7th day. In this test, the neutrophil physiology was not affected by the ambient ammonia concentrations. The blood ammonia levels of the fish exposed to the ammonia water were almost same through the whole test period despite the different water ammonia concentraions. This result is probably responsible for the noinfluence on the neutrophil physiology of the fish kept in ammonia water.On the process of in vivo ammonia toxicity test, we frequently saw aggressive behavior of a dominant fish to a subordinate one. The neutrophil function was drastically depressed in the subordinate fish compared to the dominant. Because this behavioral aspect has been overlooked in fish immunology so far, the immunologist should develop it in future.