Abstract: In the present study, the effects of isoleucine on the mammalian target of rapamycin signaling (mTOR) pathway and nitrogen metabolism were investigated through Intraventricular (ICV) administration in Chinese perch (Siniperca chuatsi). In one hand, the ammonia excretion significantly increased, and mRNA levels of nitrogen metabolism genes involving glutamate dehydrogenase (GDH), glutamic oxaloacetic transaminase (GOT), and adenosine monophosphate deaminase (AMPD) remarkably increased after ICV injection of isoleucine as well (P<0.05). On the other hand, the results showed that blood glucose levels markedly decreased at 0.5h post-injection. The expression of liver ribosomal protein S6 was also notably enhanced, demonstrating that mTOR signaling pathway was activated. The results also indicated that isoleucine could activate the mTOR signaling pathway, mediate amino acid metabolism, and enhance ammonia-N excretion via nitrogen metabolism genes in Chinese perch.