The aim of this study was to assess the metabolic fate of AA (endogenous or export protein synthesis, gluconeogenesis, or oxidation) after an imbalanced supply of energy and N in the diet of growing lambs. Eighteen INRA 401 lambs (3 mo old, 29.7 +/- 0.45 kg of BW) were fed 3 experimental diets, one providing a N and energy supply according to recommended allowances (control), one with 23% less N supply relative to energy (LN), and one with 19% less ME supply relative to N (LE). Animals were assigned to 6 blocks of 3, with each animal receiving 1 of the 3 diets, and the animals from each block were slaughtered on the same day. Liver slices from these lambs were incubated in a minimum salt medium (Krebs-Henseleit) containing physiological concentrations of propionate and AA as energy and N sources, similarly across all 3 treatments. Protein synthesis (endogenous and export) using [U-(14)C]valine and [(35)S]methionine, gluconeogenesis from [1-(14)C]propionate and [U-(14)C]alanine, and oxidation were measured. A relative sparing of AA at the liver level was observed with the LN diet because of reduced urinary N (-42%, LN vs. control, P < 0.001). The AA were also directed toward anabolic purposes in the LN diet via an increased endogenous and total export protein synthesis (+51%, LN vs. control, P = 0.01; also observed for fibrinogen synthesis, but not for albumin or transferrin) associated with a tendency for increased gluconeogenesis from alanine (+58%, LN vs. LE, P = 0.08). On the other hand, the LE diet resulted in a marked loss of N in urine (+24%, LE vs. control, P < 0.05), but no notable effect of the LE diet was demonstrated for protein synthesis or gluconeogenesis ex vivo. These data demonstrate a more efficient utilization of AA for anabolic purposes in the lambs fed LN, probably via an activation of some AA transport systems, to address the shortage of nitrogenous nutrients in the LN diet. By contrast, no such adaptation occurred in the LE lambs, probably because the regulatory mechanisms that prevailed in this case were the nutrient supply or hormones, which were not altered in our ex vivo experimental model.