The general aim of this study is to show whether the failure of insulin infusion is to stimulate muscle protein synthesis in adult humans is the result of an inadequate supply of amino acids. Previous studies of this phenomenon have concluded that infusion of amino acids to prevent the hypoaminoacidemia resulting from insulin infusion causes a stimulation of muscle protein synthesis [1]. They measured the exchange of two labeled tracers, leucine and phenylalanine, across the leg, but the results were ambiguous, as the increase in protein synthesis was only indicated by phenylalanine not by leucine. However, the uptake of label in such experiments can be altered not only by changes in the rate of protein synthesis, but also by changes in the labeling of the precursor amino acid resulting from changes in the concentration of the tracers. We therefore proposed the following hypothesis: that the apparent stimulation of muscle protein synthesis in adults humans, brought about by infusion of insulin plus amino acids, results from an increase in the labeling of the infused tracer at the site of protein synthesis.
Specific Aim 1 is to demonstrate in human volunteers infused with insulin plus amino acids plus three different tracers, that the incorporation of each labeled amino acid into protein can be varied independently by altering its concentration in the plasma. The incorporation of three labeled amino acids (L-[1-13C]leucine,L-[15N2]lysine and L- [2H5]phenylalanine) into muscle protein will be determined in volunteers infused intravenously with insulin and glucose, plus an amino acid mixture sufficient to maintain the plasma level of leucine at its pre-insulin value. The protocol will be repeated with the addition of an infusion of either unlabeled lysine or unlabeled phenylalanine to increase with plasma concentrations. The premise is that any real change in protein synthesis would be indicated similarly by all tracers, as lysine and phenylalanine are not believed to have any specific effect on muscle protein synthesis [2,3]. If the three tracers show discrepant changes in incorporation due to insulin and amino acid infusion., then this will be taken as indirect evidence of a change in precursor enrichment. As additional confirmation, the response of muscle protein synthesis to insulin plus amino acid infusion will be measured by 'flooding' with [2H5]phenylalanine, method which aims to minimize artifacts arising from changes in the enrichment of the precursor.
Specific Aim 2 will investigate the incorporation of the three tracers, L- [1-13C]leucine, L- [15N2]lysine and L-[2H5]phenylalanine, during insulin infusion with hyperaminoacidemia. The concentrations of the tracer amino acids in human subjects will be varied independently by altering the mixtures of amino acids infused as in Specific Aim 1, but the levels will be at the upper end of the physiological range rather than at levels comparable to the post-absorptive state. The combination of increased amino acids plus insulin is more representative of the fed state and it is also important under these conditions to differentiate between changes in protein synthesis and changes in precursor labeling.
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