The rainbow trout (Oncorhynchus mykiss) exhibits high dietary amino acid requirements and an apparent inefficiency to use dietary carbohydrates. Using this species, we investigated the metabolic consequences of long-term high carbohydrates/low protein feeding. Fish were fed two experimental diets containing either 20 % carbohydrates/50 % proteins (C20P50), or high levels of carbohydrates at the expense of proteins (35 % carbohydrates/35 % proteins – C35P35). The expression of genes related to hepatic and muscle glycolysis (glucokinase (GK), pyruvate kinase and hexokinase) illustrates the poor utilisation of carbohydrates irrespective of their dietary levels. The increased postprandial GK activity and the absence of inhibition of the gluconeogenic enzyme glucose-6-phosphatase activity support the hypothesis of the existence of a futile cycle around glucose phosphorylation extending postprandial hyperglycaemia. After 9 weeks of feeding, the C35P35-fed trout displayed lower body weight and feed efficiency and reduced protein and fat gains than those fed C20P50. The reduced activation of eukaryotic translation initiation factor 4-E binding protein 1 (4E-BP1) in the muscle in this C35P35 group suggests a reduction in protein synthesis, possibly contributing to the reduction in N gain. An increase in the dietary carbohydrate:protein ratio decreased the expression of genes involved in amino acid catabolism (serine dehydratase and branched-chain α-keto acid dehydrogenase E1α and E1β), and increased that of carnitine palmitoyltransferase 1, suggesting a higher reliance on lipids as energy source in fish fed high-carbohydrate and low-protein diets. This probably also contributes to the lower fat gain. Together, these results show that different metabolic pathways are affected by a high-carbohydrate/low-protein diet in rainbow trout.