Urea enrichment and production.
Urea enrichment gradually increased over time in CHO but was constant in MIX and AA. Calculated rates of urea production thus indicated a significant fall during the third hour after CHO but no changes in the other trials. Urea production coincided with Ra of Phe, i.e., both were significantly reduced during the third hour after CHO but did not change over time in the other trials (Fig. 9).
Results from this study expand upon previous findings from this laboratory, which showed an increase in net muscle protein synthesis when a large dose of amino acids was either infused (4) or ingested (12,16) after exercise. A mixture of 6 g EAA plus 35 g glucose also increased net muscle protein synthesis when given either immediately before or at one of three times after exercise, although the anabolic effect was greatest when given before exercise (17). These previous studies did not address the relative importance of components of the EAA/glucose mixture. The results from the current study show that when ∼35 g CHO was combined with ∼6 g of a balanced amino acid mixture (i.e., EAA and nonessential AA (NEAA)) the effect on net muscle protein synthesis was roughly equivalent to the sum of the independent effect of either given alone (Fig. 5). During the 2-h period after ingestion of CHO plus AA (MIX) and amino acids alone (AA), net muscle protein synthesis was significantly greater than CHO alone, but there was no difference between MIX and AA (Fig. 6).
Previous studies have provided some evidence that carbohydrate ingestion alone improves protein metabolism. Rennie et al. showed improved leucine balance during carbohydrate supplemented endurance exercise (13), whereas Roy et al. reported diminished myofibrillar proteolysis when carbohydrate was ingested following resistance exercise (14). Our results are consistent with this latter observation, as both urea production and Phe Ra were significantly lower in the CHO group relative to the predrink value at 30 min, and net balance was greater than zero throughout the 3 h after ingestion (Fig. 6). Implicit in this interpretation of our results is the assumption that in the absence of any intervention the net balance of Phe would have remained constant throughout the time during which the response to the drink was measured. The basis for this assumption is the result from our previous study in which a group of subjects received no intervention until 4 h after exercise (12). In that study there were no changes in net balance over the first 3 h after exercise (12).
Although the net effect of CHO alone on net muscle protein synthesis indicated moderate anabolism (i.e., the rate of synthesis exceeding the rate of breakdown), there was no effect of CHO on Rd, indicating no change in the rate of muscle protein synthesis. This is consistent with our previous observation that local hyperinsulinemia failed to stimulate muscle protein synthesis after exercise (5). When studies assessing the response of muscle protein synthesis to insulin are taken together, it is clear that insulin only stimulates muscle protein synthesis in the setting of adequate amino acid availability (20). Thus, it is possible that after exercise insulin increases the potential for accelerated muscle protein synthesis, but this can only be reflected in an increased production of protein if amino acid availability is increased contemporaneously. Thus, when an increase in insulin was coupled with an increase in amino acid concentration (MIX), there was an interactive effect in which the normal response of muscle protein synthesis to amino acids was amplified by the elevation of insulin, even though an isolated increase in insulin was ineffective.
Our experimental design did not control for caloric intake. Thus, the MIX group, which had the most positive net balance response, received more total calories (164) than the AA (24) or the CHO (140) group. However, it is unlikely that the observed responses can be explained by the caloric imbalances. Thus, when amino acids (24 cal) were added to carbohydrate (140 cal) a significant improvement in net balance resulted (MIX vs CHO). In contrast, when carbohydrate (140 cal) was added to amino acids (24 cal), no significant improvement in net balance resulted over the response to amino acids alone (MIX vs AA).
In contrast to the situation with CHO ingestion, there is little doubt that amino acids alone can stimulate net muscle protein synthesis. This response is elicited by either the infusion or ingestion of amino acids, whether at rest or after exercise (4,15,17). In the current study, the anabolic action of amino acids in both the AA and MIX groups was principally due to a stimulation of muscle protein synthesis, as reflected by Rd (Fig. 8). We previously showed that 40 g of a balanced mixture of amino acids stimulated muscle protein synthesis after exercise (18), but in the current study we gave a total of only two doses of 6 g each (3 g of EAAs and 3 g of NEAAs per dose). When combined with CHO ingestion, this resulted in a total net Phe uptake of 43 ± 8 mg·h−1·leg−1 over the first hour after ingestion of the first drink. This uptake is approximately half the value for the response of net Phe uptake we previously obtained (107 mg·h−1·leg−1) when we gave 6 g of EAAs + 35 g of glucose as a single dose 1 h after exercise (12). These results are consistent with our notion that only the EAAs are effective in stimulating muscle protein synthesis (18).
We can estimate that approximately 50% of ingested Phe was incorporated into whole body muscle if we assume that the Phe incorporated into one leg represented 25% of the total. If another 20% of ingested Phe was incorporated into protein in other tissues, then approximately 30% was oxidized. Comparable percentages for the other EAA in the mixture would be anticipated, and a greater percentage of NEAA intake would be expected to be oxidized. The nitrogen from the oxidized amino acids would be expected to eventually contribute to urea production, yet urea production did not increase in either group receiving amino acids. However, the lack of change in urea production in the groups receiving amino acids has to be interpreted in the context of the reduction in urea production in the group receiving only glucose (Fig. 9) (8). Also, our ability to detect the magnitude of expected change in urea production on the basis of amino acid ingested was marginal, considering the relatively high efficiency of incorporation of ingested amino acids into protein.
The response to ingestion of the second dose of each drink at 2 h after exercise was essentially the same as the response to the first dose (Fig. 6). Whereas the second drink sustained increased concentrations of glucose, Phe, or insulin (depending on the drink) throughout the 3 h following exercise, essentially all measured variables related to muscle protein metabolism returned to baseline by the end of the second hour. The rapid, transient response of amino acid concentrations to a free amino acid bolus drink observed in this study were in contrast to a previous study in which we tested the response to a primed-constant infusion of amino acids that elicited a 1.7-fold increase in amino acid concentrations that was essentially constant for the duration of the 6-h experiment (6). Muscle protein synthesis, however was only increased for the first 2 h of the elevation in amino acid concentrations (6). This diminished response over time implies that during a steady-state increase in amino acid concentrations muscle protein synthesis becomes refractory to the stimulatory effect of the amino acids. One might therefore have anticipated that there would have been a diminished response to the second drink after 1 h because the Phe concentration was still double the basal value, yet net muscle protein synthesis had dropped back to the basal value at that time. However, ingestion of the second drink 1 h after ingestion of the first drink stimulated a similar net uptake of amino acids as seen in response to the first drink (Fig. 6). The practical implication of this finding is that a supplement or beverage containing amino acids taken after exercise would appear not to diminish the stimulation of muscle protein synthesis that would result, for example, from a meal consumed at some point following exercise. From a mechanistic perspective, it may be that muscle protein synthesis responds to a change in blood or intracellular amino acid concentrations, rather than to their absolute values.
Another aspect of timing of the response suggested by our data is that the maximal insulin effect may not coincide with its peak concentration. Both Ra (reflecting muscle protein breakdown) and urea production were only significantly reduced in the third hour after the initial dose of CHO alone (Figs. 7 and 9). This reduction explains the positive net balance in this group during the third hour (Fig. 6). Furthermore, the MIX group had an almost identical response of net Phe uptake to the AA group over the first hour, and it was only in the second and particularly the third hour that the response was more positive than the AA group alone (Fig. 6). If, in fact, there is a delay in insulin action, then the peak amino acid effect may not have coincided in time with the peak insulin effect. If this is the case, amino acids ingested in a more slowly absorbed form (i.e., intact protein) may elicit a greater interactive effect between the amino acids and carbohydrate, so that the peak in amino acid concentration would coincide with the peak action of insulin.
In conclusion, our findings confirm the previously described anabolic effect on muscle protein of amino acids and glucose given after resistance exercise. Further, the results indicate that the combined effects of carbohydrate and amino acid ingestion approximately reflects the sum of their individual effects. When the results of the current study are combined with our previous results (12), it appears that ingestion of only the EAAs are required to stimulate muscle protein synthesis after exercise. Finally, the response to the second drink shows that prior ingestion of a modest amount of amino acids or carbohydrate does not diminish the metabolic response to a second comparable dose ingested 1 h later.
Studies were supported by the General Clinical Research Center at the University of Texas Medical Branch at Galveston, grant M01 RR 00073 from the National Center for Research Resources, National Institutes of Health, United States Public Health Service, National Institutes of Health grant DK 38010, and Shriners Hospital for Children grant 8490.
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Keywords:©2003The American College of Sports Medicine
STABLE ISOTOPES; UREA PRODUCTION; PHENYLALANINE; LEG BALANCE; HUMAN SUBJECTS