Resistance exercise manipulations of load (1,2), volume (3,4), and the time that muscle is experiencing loaded tension (5) have been shown to modify the magnitude and duration of the postexercise muscle protein synthetic response. Based on this work, it has been suggested that common characteristics within resistance exercise prescription to promote maximal postexercise muscle protein synthesis rates, primarily within the myofibrillar protein fraction, is to ensure each exercise set is performed to volitional fatigue (proxy indicator for bulk muscle fiber recruitment (6)) and with sufficient volume load (3). Indeed, hypertrophic muscle protein remodeling through resistance exercise training supports long-term muscle health and physical independence by expanding the myofibril protein pool and facilitating the removal of damaged proteins with the replacement of new muscle proteins (7). These past studies, however, that assessed the influence of contractile manipulation on the regulation of postexercise myofibrillar protein synthesis rates did not specifically seek to assess other important performance-related characteristics such as improvement in power.
Intraset rest is a variable that is manipulated within a resistance exercise prescription to prevent power decrements throughout an exercise session in more advanced training programs (8) but also has relevance to the general population (9,10). Specifically, the incorporation of cluster sets (CLU), whereby a small period of intraset rest is incorporated between “clusters” of repetitions has been shown to produce greater velocity and power during resistance exercise when compared with training with a continuous completion of repetitions (traditional set configuration [TRD]) (10). As such, resistance exercise prescriptions that promote long-term functional status through power preservation (11,12) as well as hypertrophic muscle protein remodeling may optimize muscle health and physical performance for both athletes and the general population. Despite the potential benefit of the CLU configuration on the skeletal muscle adaptive response, the effect of intraset rest on the postexercise regulation of muscle protein synthesis rates has not been investigated.
The mechanisms that underpin the stimulation of postexercise muscle protein synthesis rates have largely been attributed to the activation of signaling molecules within the mammalian target of rapamycin (mTOR) pathway (13). Past efforts have shown that the activation of the mTORC1 pathway is modifiable based on exercise intensity (% of 1 repetition maximum [1RM]) (14) and volume load (repetitions × load) in healthy adults (1,4). However, this hallmark of the postexercise activation of the mTOR pathway seems to be diminished by chronic resistance exercise training (15,16), consequently there is a need to better understand alternative anabolic mechanisms that may be involved in the regulation of changes in postexercise muscle protein synthesis rates. Interestingly, yes-associated protein (YAP) has recently been implicated as an mTORC1-independent mechanosensor involved in hypertrophic muscle protein remodeling in mice (17). However, the extent that resistance exercise mediates YAP expression and/or how intraset rest manipulations impacts mTORC1 pathway activation has not been investigated in humans.
Therefore, the purpose of the study was to examine the extent to which intraset rest manipulations consisting of CLU or TRD set configurations matched for total volume load alters mTORC1 and YAP mediated anabolic signaling events and postexercise myofibrillar protein synthesis rates in resistance trained young adults. Given that CLU configurations are mainly applied with compound exercises in the field of strength and conditioning (10), we utilized barbell back squats as the form of resistance exercise to address the research question. We hypothesized that CLU or TRD conditions would both stimulate postexercise myofibrillar protein synthesis rates to a similar extent when matched for total volume load. We also hypothesized that both CLU and TRD conditions would phosphorylate mTORC1 targets (i.e., p70S6K and 4EBP1) and promote YAP expression during recovery from resistance exercise.
The study was approved by the University of Illinois Institutional Review Board and conformed to standards for the use of human participants in research as outlined in the Declaration of Helsinki. This trial is registered at clinicaltrials.gov as NCT04028726. Each participant was informed of the purpose of the study, the experimental procedures, and all of the potential risks before providing their written consent to participate.
Eight resistance-trained participants (n = 7 males and 1 female; 23 ± 1 yr; 81 ± 4.7 kg; BMI, 26.6 ± 1 kg·m−2; body fat, 18% ± 1.9%) volunteered to participate in this study. Participants were regularly engaged in resistance exercise (mean ± SEM: 5 ± 0.1 sessions per week) for the past 8 ± 1 yr. All participants performed barbell squats frequently in their training with minimum strength eligibility assessed by 1RM ≥ 1.5 × body mass for study inclusion. All participants were considered healthy based on a self-reported medical screening questionnaire and had no prior history of participating in stable isotope amino acid tracer experiments. Volunteers who reported lower body musculoskeletal injury/surgery within the year prior, consumption of dietary supplements and/or ergogenic aids, within the 6 months prior, or any previous use of nonfood anabolic agents, were also excluded from participation.
At least 72 h before the first infusion trial, participants reported to the laboratory having refrained from all activities outside of daily living for at least 72 h. Upon arrival, body weight and height were measured followed by body composition assessment via dual energy x-ray absorptiometry (QDR 4500A; Hologic, Marlborough, MA). Finally, the participants were familiarized with exercise equipment before determination of their 1RM for the barbell back squat exercise (mean ± SEM: 150 ± 9.1 kg) using standardized procedures (8).
Participants were instructed to maintain their regular exercise habits between trials and refrain from vigorous physical activity for the 72 h before each infusion trial. In addition, participants were instructed to consume their habitual diet. The Automated Self-Administered Recall System (ASA24 version 2016; National Cancer Institute, Rockville, MD) was used to record their intake for the 48 h before each trial. Recorded dietary intakes before each trial were similar for the TRD (30 ± 4 kcal·kg−1 body weight, 2.7 ± 0.4 g carbohydrate·kg−1, 1.1 ± 0.19 g fat·kg−1, and 2.4 ± 0.4 g protein·kg−1) and CLU condition (31 ± 1 kcal·kg−1 body weight, 3.4 ± 0.3 g carbohydrate·kg−1, 1.1 ± 0.1 g fat·kg−1, and 2.1 ± 0.3 g protein·kg−1). All participants were provided with a standardized meal of the same composition to be consumed the evening before each infusion trial. On completion of the first trial, a copy of the diet records were returned to participants, which they were instructed to replicate for the 48 h leading up to the second trial. Participants were randomly assigned in a counterbalanced fashion to perform the TRD or CLU condition for their first trial. The time between crossover trials was approximately 7 d.
The protocol for the infusion trials is presented in Figure 1. On the infusion trial days, participants reported to the laboratory at 0600 after an overnight fast. A Teflon catheter was inserted into an dorsal vein in the hand for baseline blood sample collection, after which a primed continuous infusion of l-[ring-13C6]phenylalanine [prime: 2 μmol·kg−1 lean body mass (LBM) −1; 0.05 μmol·kg LBM−1·min−1] was initiated and maintained until the end of the trial. A second Teflon catheter was placed in a contralateral heated dorsal hand vein and kept patent by a 0.9% saline drip for repeated arterialized blood sampling. To minimize the number of biopsies collected during the trials and to provide a reference value for the assessment of postabsorptive myofibrillar protein synthesis rates (18), we collected a single resting biopsy after 3 h (t = −60) of infusion. Afterward, participants consumed 25 g of protein (2.6 g of leucine) dissolved in 350 mL of water before the performance of TRD or CLU conditions as represented in Figure 1. The TRD condition consisted of four sets of 10 repetitions (4 × 10) with a 120-s rest between sets. The CLU condition consisted of four sets of 2 × 5 clustered repetitions (4 × [2 × 5]) with 30 s of rest between clusters and 90 s between sets. Barbell load for both sessions was set at 70% 1RM. Volume load (repetitions × load) was matched between the TRD (4059 ± 244 kg) and CLU (4059 ± 244 kg) conditions (P = 1.00). Barbell kinematic data (e.g., mean and peak concentric velocity) were assessed using a commercially available optical encoder (GymAware; Kinetic Performance Technology, Caberra, Australia). Validity and methods of data acquisition for this unit are reported elsewhere (19).
RPE and affective responses were also determined after the completion of each exercise set. Participants completed the Feeling Scale (FS) (a single item scale ranging from −5 [very bad] to +5 [very good]) to assess affective valence (20,21) along with Borg’s (22). RPE scale (a single-item scale ranging from 6 [no exertion] to 20 [maximal exertion]), was used to assess perceptions of exertion during the exercise intervals. Participants were also asked to complete the Physical Activity Enjoyment Scale following (5 min post) each conditions. The Physical Activity Enjoyment Scale is an 18-item (Likert style with bipolar anchors) questionnaire that has been deemed a valid tool for measuring activity enjoyment (23). Upon completion of the exercise bout, a muscle biopsy sample was collected (t = 0) followed by the ingestion of a second drink containing 25 g of protein (2.6 g of leucine). All drinks were enriched to 4% with tracer according to the measured phenylalanine content of the whey protein to minimize disturbances in the precursor pools (24). Additional muscle biopsies were collected at 120 and 300 min of the postexercise recovery period. Muscle biopsies were collected from the middle region of the vastus lateralis (∼15 cm above the patella) with a Bergström needle modified for manual suction under local anesthesia (2% xylocaine). All muscle biopsy samples were freed from any visible blood, adipose, and connective tissue, immediately frozen in liquid nitrogen, and stored at −80°C until subsequent analysis. Arterialized blood samples were drawn every 30 or 60 min during the postabsorptive and postprandial states. Blood samples (8 mL) were collected in EDTA-containing tubes and centrifuged at 3000g at 4°C for 10 min. Aliquots of plasma were frozen and stored at −80°C until subsequent analysis.
Glucose and lactate were analyzed with whole blood using an automated biochemical analyzer (YSI 2300 Stat Plus; YSI, Yellow Springs, OH). Plasma insulin concentrations were determined using a commercially available enzyme-linked immunosorbent assay (ELISA) (ALPCO Diagnostics, Salem, NH). For plasma amino acid enrichments and concentrations, the amino acid standard solution (AAS18, Sigma, USA), containing 2.5 μmol·mL−1 each of l-alanine, l-arginine, l-aspartic acid, l-glutamic acid, glycine, l-histidine, l-isoleucine, l-leucine, l-lysine·HCl, l-methionine, l-phenylalanine, l-proline, l-serine, l-threonine, l-tyrosine, and l-valine, and 1.25 μmoL·mL−1l-cystine and a custom mixture containing 2.5 μmol·mL−1 each of l-tryptophane, l-glutamile, l-asparagine, l-citrulline, l-cysteine were used for the calibration curve. Plasma samples (200 μL) were deproteinized with acetone/isopropyl alcohol/water (3:3:2 v/v), centrifuged with after supernatant evaporation in vacuum and resuspended in 1 mL of 0.1% formic acid in water before instrument injection. Twenty microliters of internal standard (ε-aminocaproic acid, 1 mg·mL−1) was added to each sample and standard solution. Samples were analyzed by Thermo Altis Triple Quadrupole LC/MS/MS system. Software TraceFinder 4.1 was used for data acquisition and analysis. The LC separation was performed on a Thermo Accucore Vanquish C18+ column (2.1 × 100 mm, 1.5 μm) with mobile phase A (0.1% formic acid in water) and mobile phase B (0.1% formic acid in acetontrile) and the flow rate was 0.2 mL·min−1. The linear gradient was as follows: 0 to 0.5 min, 0% B; 0.5 to 3.5 min, 60% B; 3.5 to 5.5 min, 100% B; 5.5 to 7.5 min, 0% B. The autosampler and HPLC column chamber were set at 10°C and 50°C, respectively. The injection volume was 1 μL. Mass spectra were acquired under positive electrospray ionization (ESI) with the ion spray voltage of 3500 V. Selected reaction monitoring used for the amino acid quantitation.
The myofibrillar protein-enriched fraction was extracted from approximately 50 mg of wet muscle using an ice-cold homogenizing buffer (10 μL·mg−1; radioimmunoprecipitation assay) supplemented with protease and phosphatase inhibitor tablets (cOmplete and PhosSTOP; Roche Applied Science, Mannheim, Germany). Myofibrillar proteins were isolated by the differential centrifugation whereby a slow speed spin (700g) is used to pellet the myofibrillar and collagen proteins from the muscle homogenate. Subsequently, the myofibrillar proteins were solubilized by adding 0.3 M NaOH and heating at 37°C for 30 min as described previously (25). The remaining muscle homogenate was stored at −80°C for subsequent Western blot analyses. Isolated myofibrillar protein fractions were hydrolyzed overnight in 6 M HCl at 110°C. The resultant free amino acids were purified using action exchange chromatography (Dowex 50W-X8-200 resin; Acros Organics, Geel, Belgium) and dried under vacuum. Free amino acids were re-suspended in 60% methanol and centrifuged before analysis using 5500 QTRAP (Sciex, Redwood City, CA) liquid chromatography-tandem mass spectrometry in accordance with a previously reported methodology (26). The l-[ring-13C6]phenylalanine enrichments were determined by multiple reaction monitoring at m/z 166.0 → 103.0 and 172.0 → 109.0 for unlabeled and labeled l-[ring-13C6]phenylalanine, respectively. Analyst V1.6.2 (Sciex) was used for data acquisition and analysis.
The fractional synthetic rate of myofibrillar proteins were calculated using standard precursor-product methods by dividing the increment in tracer enrichment in the myofibrillar protein fraction by the enrichment of the plasma precursor pool over time as described previously (27). The recruitment of “tracer-naive” participants allowed us to use the single biopsy approach to estimate postabsorptive muscle protein fractional synthetic rate during the first infusion trial as described in detail in our past efforts (18).
Muscle homogenate supernatants from myofibrillar pelleting were stored at −80°C for subsequent Western blot analyses. Total protein concentrations of the muscle homogenate were determined using BCA assays (Pierce, Rockford, IL). Equal amounts of protein from each sample were mixed with loading buffer, denatured at 95°C for 5 min, separated on 7.5% or 10% (w/v) polyacrylamide gels, and then electrophoretically transferred to polyvinylidene fluoride membranes (MilliporeSigma, St. Louis, MO). Membranes were blocked with nonfat milk or bovine serum albumin diluted in Tris-buffered saline with Tween solution for 1 h at room temperature before overnight incubation in primary antibodies at 4°C. Proteins of interest were detected with primary antibodies as follows: rabbit anti-α-tubulin (1:1000; Abcam Cat ab4074, RRID:AB_2288001), rabbit anti-phospho-AMPKα (Thr172) (1:1000; Cell Signaling Technology Cat 2535, RRID:AB_331250), rabbit anti-AMPKα (1:1000; Cell Signaling Technology Cat 5831, RRID:AB_10622186), rabbit anti-phospho-p70S6K (Thr389) (1:500; Cell Signaling Technology Cat 9205, RRID:AB_330944), rabbit anti-p70S6K (1:500; Cell Signaling Technology Cat 9202, RRID:AB_331676), rabbit anti-phospho-4E-BP1 (Thr37/46) (1:1000; Cell Signaling Technology Cat 9459, RRID:AB_330985), rabbit anti-4E-BP1 (1:1000; Cell Signaling Technology Cat 9452, RRID:AB_331692), mouse anti-phospho-Erk1/2 (Thr202/Tyr204) (1:1000; Cell Signaling Technology Cat 9106, RRID:AB_331768), rabbit anti-Erk1/2 (1:1000; Cell Signaling Technology Cat 9102, RRID:AB_330744), rabbit anti-phospho-YAP (Ser127) (1:1000; Cell Signaling Technology Cat 4911, RRID:AB_2218913), and rabbit anti-YAP (1:1000; Cell Signaling Technology Cat 4912, RRID:AB_2218911). After primary incubation, blots were exposed to horseradish-peroxidase-conjugated horse antimouse IgG (1:2000; Cell Signaling Technology Cat 7076, RRID:AB_330924) or goat antirabbit IgG (1:2000–1:10,000; Abcam Cat ab6721, RRID:AB_955447) and detected using the ECL Western Blotting Substrate (Thermo Scientific, Waltham, MA) and the ChemiDoc XRS+ Imaging System (Bio-Rad Laboratories, Hercules, CA). Bands were quantified using ImageJ (National Institute of Health) and then normalized to α-tubulin as the internal control, and a control sample included on each blot to account for interblot variability.
Based on previous research (27,28), our power analysis showed that a sample size of 8 participants was sufficient to detect differences in postexercise muscle protein synthesis between conditions when using a 2-sided statistical test (P < 0.05, 80% power, f = 1.2; G*power version 22.214.171.124). Differences in blood metabolites, muscle protein synthesis rates, and intramuscular signaling were tested using a two-factor (condition × time) ANOVA with repeated measures on time. Acute exercise variables (repetitions, load, velocity, and power) were analyzed using a one-factor ANOVA. Where significant interactions were identified in the ANOVA, Tukey post hoc test was performed to determine differences between means for all significant main effects and interactions. Pearson r product moment correlation was used to examine the relationship between different variables (i.e., muscle protein synthesis and anabolic signaling molecules). Linear regression lines were fitted to plasma enrichments to assess the existence of any deviation in enrichment indicated by lines with a significant positive or negative slope. The level of statistical significance was set at P < 0.05 for all analyses. The data are expressed as mean ± SEM. For RPE and affective responses, Cohen’s d (d) was calculated as a measure of effect size (0.20 = small effect size; 0.50 = medium effect size; 0.80 = large effect size) for these variables.
Acute resistance exercise variables
There were no differences in repetitions (40 ± 0) performed or load lifted in sets 1–4 between TRD or CLU conditions (P = 1.00). The CLU condition resulted in greater mean (Fig. 2A) and peak concentric velocity (Fig. 2B) for sets 1–4 when compared to the TRD condition (main effect of condition: P < 0.001). Similarly, CLU produced greater average and peak power for sets 1 to 4 when compared with the TRD condition (Fig. 2; main effect of condition: P < 0.001).
RPE and affective responses
Perceptions of exertion, based on RPE responses, were significantly different between conditions (P = 0.047) and across time (P = 0.004), but the condition–time interaction was not significant (P = 0.17). The RPE responses were higher in the TRD condition (16.3) compared with the CLU condition (15.8) and increased over time in both conditions (14.6 to 17.4). Although the interaction was not significant, following the second set, perceptions of exertion were approximately 1 unit less in the CLU condition (17.3 vs 16.1, 17.9 vs 16.9 following the third and fourth sets, respectively). There were no significant condition (P = 0.29), time (P = 0.21), or condition–time interaction (P = 0.11) effects for the affective valence measures (which is a measure of how pleasant or unpleasant the individual feels). Overall, affect was slightly worse in TRD (1.6) than CLU (1.9), but it stayed fairly stable in TRD (~1.5 after each set); for CLU, affect started higher (2.6) and then declined, with the largest decline after the second set compared with after the third set. Finally, when the participants were asked to rate their enjoyment of the two approaches, while not significant, enjoyment was greater for the CLU (100.8) than for the TRD (96.3) (Cohen’s d effect size = 0.51).
Blood glucose concentration increased above baseline immediately after resistance exercise with no differences between TRD or CLU conditions (Table 1; main effect of time: P = 0.03). Blood lactate concentration was higher after TRD when compared with CLU during recovery from resistance exercise (Table 1; main effect of condition: P < 0.001). Plasma insulin concentration reached peak values (P = 0.01) at 30 min after the ingestion of the second protein bolus and returned to basal values by 1.5 h, irrespective of condition (Table 1). Plasma EAA concentrations reached peaked values at 1 h (P < 0.001) after the ingestion of the second protein bolus and returned to baseline values by 2 h of the postexercise period irrespective of condition (Table 1). Plasma phenylalanine concentrations reached peak values of 70.8 ± 3.6 μM and 70.2 ± 3.8 μM at 0.5 h after the ingestion of the second protein bolus and retuned to baseline values of 61.0 ± 3.7 μM and 58.4 ± 3.5 μM at 1.5 h of the postexercise period in both the TRD and CLU conditions, respectively. Basal plasma l-[ring-13C6]phenylalanine enrichments did not differ between the TRD (averaged 0.053 ± 0.006 tracer·tracee−1; TTR) or CLU (0.048 ± 0.007 TTR) conditions (P > 0.05). Moreover, postprandial plasma l-[ring-13C6]phenylalanine enrichments did not differ between the TRD or CLU conditions (P = 0.26). Specifically, plasma l-[ring-13C6]phenylalanine enrichments increased (P < 0.05) after the ingestion of the second protein bolus during the late postexercise phase (2–5 h) in the TRD (0.072 ± 0.007 TTR) and CLU condition (0.073 ± 0.008 TTR). Linear regression analysis indicated that the slopes of the plasma enrichments were not significantly different from zero, regardless of condition, during the early (0–2 h) or late phase (2–5 h) of the postexercise period, demonstrating isotopic plateau during these measurement periods.
Muscle anabolic signaling
Phosphorylated, total, or phosphorylated-to-total ratios of AMPKα, P70S6K, and 4E-BP1 were unaffected by both conditions and did not increase from baseline values at any time point during recovery (Fig. 3; all P > 0.05). Phosphorylated (Fig. 4A) and total expression of YAP (Fig. 4B) were elevated (P < 0.05) immediately postexercise regardless of condition (main effect for time: P = 0.006 and 0.009, respectively) with no change to the phosphorylated-to-total ratio (P = 0.134; Fig. 4C). Similarly, phosphorylation of ERK1/2Thr202/Tyr204 and phosphorylated-to-total ERK1/2 ratio were significantly elevated immediately postexercise with no differences between condition (main effect for time: P = 0.004 and 0.006, respectively; Fig. 3B).
Myofibrillar protein synthesis
Myofibrillar protein synthesis rates increased above baseline by approximately 4- and 3-fold during the early phase (0–2 h) of postexercise recovery for the TRD and CLU conditions, respectively (both P < 0.001; Fig. 5A). TRD tended (P = 0.096) to increase the postexercise myofibrillar protein synthetic response to a greater extent during the early phase of recovery when compared with the CLU condition. However, the increase in the cumulative myofibrillar protein synthetic response calculated over the entire 5 h postexercise phase did not differ (P = 0.48) between the TRD (0.074 ± 0.013%·h−1) or CLU (0.063 ± 0.018%·h−1) condition.
Our study is the first to characterize the impact of intraset rest on the postexercise regulation of myofibrillar protein synthesis rates and anabolic signaling in trained young adults. We show that the CLU configuration resulted in higher power output and a similar stimulation of the cumulative (0–5 h) postexercise myofibrillar protein synthetic response when compared with TRD with total volume load equated. In addition, we show that resistance exercise, regardless of condition, induced an increase in total and phosphorylated YAP expression without altering mTORC1-related signaling events (i.e., p70S6k and 4EBP1 phosphorylation).
It is indeed interesting that our results demonstrated a tendency (P = 0.09) for the postexercise myofibrillar protein synthetic response to be greater during the early (0–2 h) phase of recovery in the TRD compared with the CLU condition (Fig. 5A). We speculate this could be related to the decrease in concentric velocity (Fig. 2) resulting in greater time the muscle is experiencing loaded tension (10), which likely would have increased surface EMG activity in the TRD when compared with CLU condition, had it been measured in our study (29). This notion is consistent with the thesis that motor unit recruitment and activation modulates the postexercise stimulation of myofibrillar protein synthesis rates (3,5). What is noteworthy is that the postexercise myofibrillar protein synthetic response, irrespective of condition, returned to the basal-state by 3 to 5 h of recovery. The short-lived nature of the postexercise myofibrillar protein synthetic response is likely linked to the trained status of the volunteers. For example, it has been shown that the stimulation of postexercise muscle protein synthesis rates is attenuated both in terms of amplitude and duration in trained versus novice weight lifters (30–32).
The influence of acute resistance exercise on the activation-status of the mTORC1 pathway has been shown to be intensity (%1RM) (14) and volume load (1) sensitive in humans. Here, we show that the mTORC1 pathway, as noted by the extent p70SK1 and 4E-BP1 phosphorylation, remained unchanged from preexercise to postexercise regardless of the intraset rest manipulation. Similarly, we show AMPK phosphorylation, a cellular energy sensor (33), was not altered during recovery from resistance exercise. Again, these findings are likely related to the trained nature of the participants whereby the phosphorylation of these proteins has been shown to be diminished by regular resistance exercise (15,16). Moreover, it is also possible that other relevant mTORC1 regulatory events, such as protein–protein interactions or intracellular redistribution to the sarcolemma, occurred and were not captured by our more traditional Western blot readout of protein phosphorylation (34).
Given that training status (15) and regular resistance exercise training (16) have been shown to reduce mTORC1 pathway activation, we specifically sought to investigate other potential anabolic mechanisms that may modulate the postexercise stimulation of muscle protein synthesis rates in trained volunteers. Recently, YAP has been implicated as a mechanosensor involved in hypertrophic muscle protein remodeling (17). In this study, we show that total and phosphorylated YAP on Ser127 is increased immediately after acute resistance exercise irrespective of condition. Our results are consistent with those of Goodman et al. (17) who demonstrated that mechanical loading increased total and phosphorylated YAP on Ser112 (corresponding to Ser127 in humans) in hypertrophic mice muscle. Indeed, it could be speculated that the immediate intramuscular adjustment of total YAP protein implies it is translationally regulated and/or stabilized through reduction of YAP degradation rates, thereby leading to YAP accumulation. In any case, the underlying mechanism in which YAP transduces mechanical signals to regulate muscle mass has not been completely deciphered (17,35), but it has been speculated that YAP expression is not involved in the contraction-induced activation of mTORC1 as originally assumed (17).
Interestingly, we observed a relationship between total (r = 0.67; P < 0.0001) and phosphorylated (r = 0.58; P < 0.0001) human YAP with Erk1/2 phosphorylation immediately after resistance exercise regardless of the TRD or CLU configuration. The increase in Erk1/2 phosphorylation in our study is consistent with past efforts that have shown the MAPK pathway is sensitive to acute resistance exercise (36–38); however, our results suggest there may be a connection between hippo/YAP signaling and Erk1/2 phosphorylation in human skeletal muscle as shown in other cell models (39). This notion is consistent with results that have shown Erk1/2 phosphorylation is mTORC1 independent in rodents (40,41). From our study design, it is not possible to decipher the influence of training status per se, but it is possible that mTORC1-dependent and -independent events are differentially modulated in trained versus untrained muscle in response to acute resistance exercise (42). Moreover, it is possible that YAP localization, rather than phosphorylation, may be more relevant as a mediator of the anabolic response as suggested by a cell-based assay (43). These are points for future work to explore.
Overall, our results implicate the CLU configuration as an easily manipulated resistance exercise variable to maximize power development as well as facilitate muscle protein remodeling as indicated by similar increases in the cumulative (0–5 h) postexercise myofibrillar protein synthetic response between the TRD and CLU conditions with equal total volume load. In support, Oliver et al. (2013) showed that 12 wk of training with CLU or TRD configurations resulted in similar increases lean body mass in trained young men. From a psychological perspective, the CLU approach was perceived as less effortful, particularly on the later sets, yet rated the same affectively and was somewhat more enjoyable. This would seem to suggest that either the TRD or CLU approach could be used depending on the individual’s preference without detrimental affective or perceptual consequences. Certainly, future efforts would be required to directly determine the translation of our findings into other populations (e.g., aging) and other exercise selections (e.g., leg extension) that are often commonplace within a program of resistance exercise training, especially in the general population.
In conclusion, intraset rest manipulations of CLU or TRD configurations stimulated a similar increase in cumulative (0–5 h) myofibrillar protein synthesis rates. This result was primarily driven by the early (0–2 h) stimulation of myofibrillar protein synthesis rates as the response waned in the later phase (2–5 h) of recovery from resistance exercise. These results demonstrate that intraset rest is an exercise variable that can be manipulated to augment power output and support the acute skeletal muscle adaptive response without altering total volume load or the duration of the exercise bout. Finally, our findings support the notion that YAP may be involved in resistance exercise-induced muscle protein remodeling in healthy trained young adults.
The authors wish to thank volunteers for their time and effort. Funding for this work was supported by internal funds from the University of Illinois. A. F. Salvador is supported by Coordination for the Improvement of Higher Education Personnel (CAPES).
Conflict of Interest. No authors have any conflicts of interest, financial or otherwise, to declare. The results of the present study do not constitute endorsement by the American College of Sports Medicine and are presented clearly, honestly, and without fabrication, falsification, or inappropriate data manipulation.
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