This experiment was conducted to examine the effect of dietary nucleotides (NT) on fish performance and acute stress response on fingerling rainbow trout (23 g ± 0.01, mean weight ± SEM). Five experimental diets according to different levels of supplemented nucleotides (0, 0.05, 0.1, 0.15, and 0.2%) were assayed on experimental fish for 8 weeks. Growth, hematological parameters (hematocrit, hemoglobin, erythrocyte, lymphocyte, and neutrophil count), serum proteins (globulin, albumin), and plasma enzymatic activity (alkaline phosphatase, ALP; aspartate transaminase, AST; lactate dehydrogenase, LDH; alanine transaminase, ALT) were assayed. At the end of feeding trial, fish fed the control and 0.2% diets were subjected to handling and crowding stress. Modulatory effects of nucleotides on acute stress response (cortisol and glucose) and plasma electrolytes (Na(+), Cl(-), K(+), and Ca(2+)) were studied. The percentage of body weight gain (WG) and feed efficiency (FE) of fish were better when the fish were fed 0.15-0.2% diets. Fish fed the nucleotide-supplemented diets tended to have lower levels of serum enzymes including ALP, AST, LDH, and ALT. Plasma cortisol levels of fish fed on 0.2% diet under handling and crowding stress were significantly lower than fish fed the control diet at all post-stress time intervals. In our study, fish fed nucleotide-supplemented diet had significantly lower concentrations of glucose compared to those fed the basal diet. The concentrations of sodium, chloride, calcium, and potassium of fish fed the control diet were significantly lower than in fish fed nucleotide-supplemented diet. Dietary nucleotides administration seems to promote growth and to enhance resistance against handling and crowding stress in fingerling rainbow trout.