Differential effects of long-term leucine infusion on tissue protein synthesis in neonatal pigs

Abstract

Leucine is unique among the amino acids in its ability to promote protein synthesis by activating translation initiation via the mammalian target of rapamycin (mTOR) pathway. Previously, we showed that leucine infusion acutely stimulates protein synthesis in fast-twitch glycolytic muscle of neonatal pigs but this response cannot be maintained unless the leucine-induced fall in amino acids is prevented. To determine whether leucine can stimulate protein synthesis in muscles of different fiber types and in visceral tissues of the neonate in the long-term if baseline amino acid concentrations are maintained, overnight fasted neonatal pigs were infused for 24 h with saline, leucine (400 micromol kg(-1) h(-1)), or leucine with replacement amino acids to prevent the leucine-induced hypoaminoacidemia. Changes in the fractional rate of protein synthesis and activation of mTOR, as determined by eukaryotic initiation factor 4E binding protein (4E-BP1) and S6 kinase 1 (S6K1) phosphorylation, in the gastrocnemius and masseter muscles, heart, liver, jejunum, kidney, and pancreas were measured. Leucine increased mTOR activation in the gastrocnemius and masseter muscles, liver, and pancreas, in both the absence and presence of amino acid replacement. However, protein synthesis in these tissues was increased only when amino acids were infused to maintain baseline levels. There were no changes in mTOR signaling or protein synthesis in the other tissues we examined. Thus, long-term infusion of leucine stimulates mTOR signaling in skeletal muscle and some visceral tissues but the leucine-induced stimulation of protein synthesis in these tissues requires sustained amino acid availability.

Fig. 1

Fractional rates of muscle protein synthesis (K_s) in a muscle and b visceral tissues after a 24 h infusion of saline or leucine with and without replacement amino acids. Gastroc gastrocnemius muscle, Mass masseter muscle, RH right chamber of heart, and LH left chamber of heart. ANOVA indicates an effect of treatment in the gastrocnemius (P = 0.057), masseter (P = 0.009), liver (P < 0.001) and pancreas (P = 0.003). Results of TUKEY post hoc testing at P < 0.05 denoted by different letters. Values are mean ± SE; n = 6 per treatment

Fig. 2

Phosphorylation of 4E-BP1 (a) and S6K1 (b) in the gastrocnemius, masseter, right and left ventricles after a 24 h infusion of saline or leucine with and without replacement amino acids. ANOVA indicates an effect of treatment on 4E-BP1 for gastrocnemius (P = 0.045) and masseter (P = 0.001), and for S6K1 in the gastrocnemius (P = 0.025) and masseter (P = 0.022). Results of TUKEY post hoc testing at P < 0.05 denoted by different letters. Values are mean ± SE; n = 6 per treatment for all tissues

Fig. 3

Phosphorylation of 4E-BP1 (a) and S6K1 (b) in the liver, pancreas, kidney and jejunum after a 24 h infusion of saline or leucine with and without replacement amino acids. ANOVA indicates an effect of treatment on 4E-BP1 for liver (P = 0.002) and pancreas (P = 0.052), and for S6K1 in the liver (P = 0.003). Results of TUKEY post hoc testing at P < 0.05 denoted by different letters. Values are mean ± SE; n = 6 per treatment for all tissues except pancreas n = 3 per treatment