Accentuated Eccentric Loading and Cluster Set Configurations in the Back Squat: A Kinetic and Kinematic Analysis.

ABSTRACT: Wagle, JP, Cunanan, AJ, Carroll, KM, Sams, ML, Wetmore, A, Bingham, GE, Taber, CB, DeWeese, BH, Sato, K, Stuart, CA, and Stone, MH. Accentuated eccentric loading and cluster set configurations in the back squat: a kinetic and kinematic analysis. J Strength Cond Res 35(2): 420-427, 2021-This study examined the kinetic and kinematic differences between accentuated eccentric loading (AEL) and cluster sets in trained male subjects (age = 26.1 ± 4.1 years, height = 183.5 ± 4.3 cm, body mass = 92.5 ± 10.5 kg, and back squat to body mass ratio = 1.8 ± 0.3). Four load condition sessions consisted of traditionally loaded (TL) "straight sets," TL cluster (TLC) sets, AEL cluster (AEC) sets, and AEL "straight sets" where only the first repetition had eccentric overload (AEL1). An interrepetition rest interval of 30 seconds was prescribed for both TLC and AEC. Concentric intensity for all load conditions was 80% 1 repetition maximum (1RM). Accentuated eccentric loading was applied to repetitions using weight releasers with total eccentric load equivalent to 105% of concentric 1RM. Traditionally loaded cluster had statistically greater concentric outputs than TL. Furthermore, statistically greater eccentric and concentric outputs were observed during AEC compared with TL with the exception of peak power. Statistically greater concentric characteristics were observed in TLC compared with AEL1, but statistically greater eccentric outputs were observed in AEL1. In the 2 cluster set conditions, statistically greater concentric rate of force development (RFDCON) (d = 0.470, p < 0.001) and average velocity (vavg) (d = 0.560, p < 0.001) in TLC compared with AEC were observed. However, statistically greater eccentric work (WECC) (d = 2.096, p < 0.001) and eccentric RFD (RFDECC) (d = 0.424, p < 0.001) were observed in AEC compared with TLC. Overall, eccentric overload demonstrated efficacy as a means of increasing eccentric work and RFD, but not as a means of potentiating concentric output. Finally, interrepetition rest seems to have the largest influence on concentric power output and RFD.

Preliminary Investigation Into the Effect of ACTN3 and ACE Polymorphisms on Muscle and Performance Characteristics.

ABSTRACT: Wagle, JP, Carroll, KM, Cunanan, AJ, Wetmore, A, Taber, CB, DeWeese, BH, Sato, K, Stuart, CA, and Stone, MH. Preliminary investigation into the effect of ACTN3 and ACE polymorphisms on muscle and performance characteristics. J Strength Cond Res 35(3): 688-694, 2021-The purpose of this investigation was to explore the phenotypic and performance outcomes associated with ACTN3 and ACE polymorphisms. Ten trained men (age = 25.8 ± 3.0 years, height = 183.3 ± 4.1 cm, body mass = 92.3 ± 9.3 kg, and back squat to body mass ratio = 1.8 ± 0.3) participated. Blood samples were analyzed to determine ACTN3 and ACE polymorphisms. Standing ultrasonography images of the vastus lateralis (VL) were collected to determine whole muscle cross-sectional area (CSA-M), and a percutaneous muscle biopsy of the VL was collected to determine type I-specific CSA (CSA-T1), type II-specific CSA (CSA-T2), and type II to type I CSA ratio (CSA-R). Isometric squats were performed on force platforms with data used to determine peak force (IPF), allometrically scaled peak force (IPFa), and rate of force development (RFD) at various timepoints. One repetition maximum back squats were performed, whereby allometrically scaled dynamic strength (DSa) was determined. Cohen's d effect sizes revealed ACTN3 RR and ACE DD tended to result in greater CSA-M but differ in how they contribute to performance. ACTN3 RR's influence seems to be in the type II fibers, altering maximal strength, and ACE DD may influence RFD capabilities through a favorable CSA-R. Although the findings of the current investigation are limited by the sample size, the findings demonstrate the potential influence of ACTN3 and ACE polymorphisms on isometric and dynamic strength testing. This study may serve as a framework to generate hypotheses regarding the effect of genetics on performance.

Skeletal Muscle Fiber Adaptations Following Resistance Training Using Repetition Maximums or Relative Intensity.

The purpose of the study was to compare the physiological responses of skeletal muscle to a resistance training (RT) program using repetition maximum (RM) or relative intensity (RISR). Fifteen well-trained males underwent RT 3 d·wk-1 for 10 weeks in either an RM group (n = 8) or RISR group (n = 7). The RM group achieved a relative maximum each day, while the RISR group trained based on percentages. The RM group exercised until muscular failure on each exercise, while the RISR group did not reach muscular failure throughout the intervention. Percutaneous needle biopsies of the vastus lateralis were obtained pre-post the training intervention, along with ultrasonography measures. Dependent variables were: Fiber type-specific cross-sectional area (CSA); anatomical CSA (ACSA); muscle thickness (MT); mammalian target of rapamycin (mTOR); adenosine monophosphate protein kinase (AMPK); and myosin heavy chains (MHC) specific for type I (MHC1), type IIA (MHC2A), and type IIX (MHC2X). Mixed-design analysis of variance and effect size using Hedge's g were used to assess within- and between-group alterations. RISR statistically increased type I CSA (p = 0.018, g = 0.56), type II CSA (p = 0.012, g = 0.81), ACSA (p = 0.002, g = 0.53), and MT (p < 0.001, g = 1.47). RISR also yielded a significant mTOR reduction (p = 0.031, g = -1.40). Conversely, RM statistically increased only MT (p = 0.003, g = 0.80). Between-group effect sizes supported RISR for type I CSA (g = 0.48), type II CSA (g = 0.50), ACSA (g = 1.03), MT (g = 0.72), MHC2X (g = 0.31), MHC2A (g = 0.87), and MHC1 (g = 0.59); with all other effects being of trivial magnitude (g < 0.20). Our results demonstrated greater adaptations in fiber size, whole-muscle size, and several key contractile proteins when using RISR compared to RM loading paradigms.

Divergent Performance Outcomes Following Resistance Training Using Repetition Maximums or Relative Intensity.

PURPOSE: To compare repetition maximum (RM) to relative intensity using sets and repetitions (RISR) resistance training on measures of training load, vertical jump, and force production in well-trained lifters. METHODS: Fifteen well-trained (isometric peak force = 4403.61 [664.69] N, mean [SD]) males underwent resistance training 3 d/wk for 10 wk in either an RM group (n = 8) or RISR group (n = 7). Weeks 8 to 10 consisted of a tapering period for both groups. The RM group achieved a relative maximum each day, whereas the RISR group trained based on percentages. Testing at 5 time points included unweighted (<1 kg) and 20-kg squat jumps, countermovement jumps, and isometric midthigh pulls. Mixed-design analyses of variance and effect size using Hedge's g were used to assess within- and between-groups alterations. RESULTS: Moderate between-groups effect sizes were observed for all squat-jump and countermovement-jump conditions supporting the RISR group (g = 0.76-1.07). A small between-groups effect size supported RISR for allometrically scaled isometric peak force (g = 0.20). Large and moderate between-groups effect sizes supported RISR for rate of force development from 0 to 50 ms (g = 1.25) and 0 to 100 ms (g = 0.89). Weekly volume load displacement was not different between groups (P > .05); however, training strain was statistically greater in the RM group (P < .05). CONCLUSIONS: Overall, this study demonstrated that RISR training yielded greater improvements in vertical jump, rate of force development, and maximal strength compared with RM training, which may be explained partly by differences in the imposed training stress and the use of failure/nonfailure training in a well-trained population.

Repetition-to-Repetition Differences Using Cluster and Accentuated Eccentric Loading in the Back Squat.

The current investigation was an examination of the repetition-to-repetition magnitudes and changes in kinetic and kinematic characteristics of the back squat using accentuated eccentric loading (AEL) and cluster sets. Trained male subjects (age = 26.1 ± 4.1 years, height = 183.5 ± 4.3 cm, body mass = 92.5 ± 10.5 kg, back squat to body mass ratio = 1.8 ± 0.3) completed four load condition sessions, each consisting of three sets of five repetitions of either traditionally loaded straight sets (TL), traditionally loaded cluster sets (TLC), AEL cluster sets (AEC), and AEL straight sets where only the initial repetition had eccentric overload (AEL1). Eccentric overload was applied using weight releasers, creating a total eccentric load equivalent to 105% of concentric one repetition maximum (1RM). Concentric load was 80% 1RM for all load conditions. Using straight sets (TL and AEL1) tended to decrease peak power (PP) (d = −1.90 to −0.76), concentric rate of force development (RFDCON) (d = −1.59 to −0.27), and average velocity (MV) (d = −3.91 to −1.29), with moderate decreases in MV using cluster sets (d = −0.81 to −0.62). Greater magnitude eccentric rate of force development (RFDECC) was observed using AEC at repetition three (R3) and five (R5) compared to all load conditions (d = 0.21⁻0.65). Large within-condition changes in RFDECC from repetition one to repetition three (∆REP1⁻3) were present using AEL1 (d = 1.51), demonstrating that RFDECC remained elevated for at least three repetitions despite overload only present on the initial repetition. Overall, cluster sets appear to permit higher magnitude and improved maintenance of concentric outputs throughout a set. Eccentric overload with the loading protocol used in the current study does not appear to potentiate concentric output regardless of set configuration but may cause greater RFDECC compared to traditional loading.

Muscle hypertrophy in prediabetic men after 16 wk of resistance training.

Resistance training of healthy young men typically results in muscle hypertrophy and a shift in vastus lateralis composition away from type IIx fibers to an increase in IIa fiber content. Our previous studies of 8 wk of resistance training found that many metabolic syndrome men and women paradoxically increased IIx fibers with a decrease in IIa fibers. To confirm the hypothesis that obese subjects might have muscle remodeling after resistance training very different from healthy lean subjects, we subjected a group of nine obese male volunteers to progressive resistance training for a total of 16 wk. In these studies, weight loss was discouraged so that muscle changes would be attributed to the training alone. Detailed assessments included comparisons of histological examinations of needle biopsies of vastus lateralis muscle pretraining and at 8 and 16 wk. Prolonging the training from 8 to 16 wk resulted in increased strength, improved body composition, and more muscle fiber hypertrophy, but euglycemic clamp-quantified insulin responsiveness did not improve. Similar to prior studies, muscle fiber composition shifted toward more fast-twitch type IIx fibers (23 to 42%). Eight weeks of resistance training increased the muscle expression of phosphorylated Akt2 and mTOR. Muscle GLUT4 expression increased, although insulin receptor and IRS-1 expression did not change. We conclude that resistance training of prediabetic obese subjects is effective at changing muscle, resulting in fiber hypertrophy and increased type IIx fiber content, and these changes continue up to 16 wk of training.NEW & NOTEWORTHY Obese, insulin-resistant men responded to 16 wk of progressive resistance training with muscle hypertrophy and increased strength and a shift in muscle fiber composition toward fast-twitch, type IIx fibers. Activation of muscle mTOR was increased by 8 wk but did not increase further at 16 wk despite continued augmentation of peak power and rate of force generation.

Pre-Training Muscle Characteristics of Subjects Who Are Obese Determine How Well Exercise Training Will Improve Their Insulin Responsiveness.

Stuart, CA, Lee, ML, South, MA, Howell, MEA, Cartwright, BM, Ramsey, MW, and Stone, MH. Pre-training muscle characteristics of subjects who are obese determine how well exercise training will improve their insulin responsiveness. J Strength Cond Res 31(3): 798-808, 2017-Only half of prediabetic subjects who are obese who underwent exercise training without weight loss increased their insulin responsiveness. We hypothesized that those who improved their insulin responsiveness might have pretraining characteristics favoring a positive response to exercise training. Thirty nondiabetic subjects who were obese volunteered for 8 weeks of either strength training or endurance training. During training, subjects increased their caloric intake to prevent weight loss. Insulin responsiveness by euglycemic clamps and muscle fiber composition, and expression of muscle key biochemical pathways were quantified. Positive responders initially had 52% higher intermediate muscle fibers (fiber type IIa) with 27% lower slow-twitch fibers (type I) and 23% lower expression of muscle insulin receptors. Whether after weight training or stationary bike training, positive responders' fiber type shifted away from type I and type IIa fibers to an increased proportion of type IIx fibers (fast twitch). Muscle insulin receptor expression and glucose transporter type 4 (GLUT4) expression increased in all trained subjects, but these moderate changes did not consistently translate to improvement in whole-body insulin responsiveness. Exercise training of previously sedentary subjects who are obese can result in muscle remodeling and increased expression of key elements of the insulin pathway, but in the absence of weight loss, insulin sensitivity improvement was modest and limited to about half of the participants. Our data suggest rather than responders being more fit, they may have been less fit, only catching up to the other half of subjects who are obese whose insulin responsiveness did not increase beyond their pretraining baseline.

Comparison of the Relationship between Lying and Standing Ultrasonography Measures of Muscle Morphology with Isometric and Dynamic Force Production Capabilities.

The purpose of the current study was (1) to examine the differences between standing and lying measures of vastus lateralis (VL), muscle thickness (MT), pennation angle (PA), and cross-sectional area (CSA) using ultrasonography; and (2) to explore the relationships between lying and standing measures with isometric and dynamic assessments of force production-specifically peak force, rate of force development (RFD), impulse, and one-repetition maximum back squat. Fourteen resistance-trained subjects (age = 26.8 ± 4.0 years, height = 181.4 ± 6.0 cm, body mass = 89.8 ± 10.7 kg, back squat to body mass ratio = 1.84 ± 0.34) agreed to participate. Lying and standing ultrasonography images of the right VL were collected following 48 hours of rest. Isometric squat assessments followed ultrasonography, and were performed on force platforms with data used to determine isometric peak force (IPF), as well as RFD and impulse at various time points. Forty-eight hours later, one-repetition maximum back squats were performed by each subject. Paired-samples t-tests revealed statistically significant differences between standing and lying measurements of MT (p < 0.001), PA (p < 0.001), and CSA (p ≤ 0.05), with standing values larger in all cases. Further, standing measures were correlated more strongly and abundantly to isometric and dynamic performance. These results suggest that if practitioners intend to gain insight into strength-power potential based on ultrasonography measurements, performing the measurement collection with the athlete in a standing posture may be preferred.

Effects of Short-Term Free-Weight and Semiblock Periodization Resistance Training on Metabolic Syndrome.

South, MA, Layne, AS, Stuart, CA, Triplett, NT, Ramsey, MW, Howell, ME, Sands, WA, Mizuguchi, S, Hornsby, WG, Kavanaugh, AA, and Stone, MH. Effects of short-term free-weight and semiblock periodization resistance training on metabolic syndrome. J Strength Cond Res 30(10): 2682-2696, 2016-The effects of short-term resistance training on performance and health variables associated with prolonged sedentary lifestyle and metabolic syndrome (MS) were investigated. Resistance training may alter a number of health-related, physiological, and performance variables. As a result, resistance training can be used as a valuable tool in ameliorating the effects of a sedentary lifestyle including those associated with MS. Nineteen previously sedentary subjects (10 with MS and 9 with nonmetabolic syndrome [NMS]) underwent 8 weeks of supervised resistance training. Maximum strength was measured using an isometric midthigh pull and resulting force-time curve. Vertical jump height (JH) and power were measured using a force plate. The muscle cross-sectional area (CSA) and type were examined using muscle biopsy and standard analysis techniques. Aerobic power was measured on a cycle ergometer using a ParvoMedics 2400 Metabolic system. Endurance was measured as time to exhaustion on a cycle ergometer. After training, maximum isometric strength, JH, jump power, and V[Combining Dot Above]O2peak increased by approximately 10% (or more) in both the metabolic and NMS groups (both male and female subjects). Over 8 weeks of training, body mass did not change statistically, but percent body fat decreased in subjects with the MS and in women, and lean body mass increased in all groups (p ≤ 0.05). Few alterations were noted in the fiber type. Men had larger CSAs compared those of with women, and there was a fiber-specific trend toward hypertrophy over time. In summary, 8 weeks of semiblock free-weight resistance training improved several performance variables and some cardiovascular factors associated with MS.

Myosin content of individual human muscle fibers isolated by laser capture microdissection.

Muscle fiber composition correlates with insulin resistance, and exercise training can increase slow-twitch (type I) fibers and, thereby, mitigate diabetes risk. Human skeletal muscle is made up of three distinct fiber types, but muscle contains many more isoforms of myosin heavy and light chains, which are coded by 15 and 11 different genes, respectively. Laser capture microdissection techniques allow assessment of mRNA and protein content in individual fibers. We found that specific human fiber types contain different mixtures of myosin heavy and light chains. Fast-twitch (type IIx) fibers consistently contained myosin heavy chains 1, 2, and 4 and myosin light chain 1. Type I fibers always contained myosin heavy chains 6 and 7 (MYH6 and MYH7) and myosin light chain 3 (MYL3), whereas MYH6, MYH7, and MYL3 were nearly absent from type IIx fibers. In contrast to cardiomyocytes, where MYH6 (also known as α-myosin heavy chain) is seen solely in fast-twitch cells, only slow-twitch fibers of skeletal muscle contained MYH6. Classical fast myosin heavy chains (MHC1, MHC2, and MHC4) were present in variable proportions in all fiber types, but significant MYH6 and MYH7 expression indicated slow-twitch phenotype, and the absence of these two isoforms determined a fast-twitch phenotype. The mixed myosin heavy and light chain content of type IIa fibers was consistent with its role as a transition between fast and slow phenotypes. These new observations suggest that the presence or absence of MYH6 and MYH7 proteins dictates the slow- or fast-twitch phenotype in skeletal muscle.

Insulin resistance and muscle insulin receptor substrate-1 serine hyperphosphorylation.

Insulin resistance in metabolic syndrome subjects is profound in spite of muscle insulin receptor and insulin-responsive glucose transporter (GLUT4) expression being nearly normal. Insulin receptor tyrosine kinase phosphorylation of insulin receptor substrate-1 (IRS-1) at Tyr896 is a necessary step in insulin stimulation of translocation of GLUT4 to the cell surface. Serine phosphorylation of IRS-1 by some kinases diminishes insulin action in mice. We evaluated the phosphorylation status of muscle IRS-1 in 33 subjects with the metabolic syndrome and seventeen lean controls. Each underwent euglycemic insulin clamps and a thigh muscle biopsy before and after 8 weeks of either strength or endurance training. Muscle IRS-1 phosphorylation at six sites was quantified by immunoblots. Metabolic syndrome muscle IRS-1 had excess phosphorylation at Ser337 and Ser636 but not at Ser307, Ser789, or Ser1101. Ser337 is a target for phosphorylation by glycogen synthase kinase 3 (GSK3) and Ser636 is phosphorylated by c-Jun N-terminal kinase 1 (JNK1). Exercise training without weight loss did not change the IRS-1 serine phosphorylation. These data suggest that baseline hyperphosphorylation of at least two key serines within muscle IRS-1 diminishes the transmission of the insulin signal and thereby decreases the insulin-stimulated translocation of GLUT4. Excess fasting phosphorylation of muscle IRS-1 at Ser636 may be a major cause of the insulin resistance seen in obesity and might prevent improvement in insulin responsiveness when exercise training is not accompanied by weight loss.

Insulin responsiveness in metabolic syndrome after eight weeks of cycle training.

INTRODUCTION: Insulin resistance in obesity is decreased after successful diet and exercise. Aerobic exercise training alone was evaluated as an intervention in subjects with the metabolic syndrome. METHODS: Eighteen nondiabetic, sedentary subjects, 11 with the metabolic syndrome, participated in 8 wk of increasing intensity stationary cycle training. RESULTS: Cycle training without weight loss did not change insulin resistance in metabolic syndrome subjects or sedentary control subjects. Maximal oxygen consumption (V·O 2max), activated muscle AMP-dependent kinase, and muscle mitochondrial marker ATP synthase all increased. Strength, lean body mass, and fat mass did not change. The activated mammalian target of rapamycin was not different after training. Training induced a shift in muscle fiber composition in both groups but in opposite directions. The proportion of type 2× fibers decreased with a concomitant increase in type 2a mixed fibers in the control subjects, but in metabolic syndrome, type 2× fiber proportion increased and type 1 fibers decreased. Muscle fiber diameters increased in all three fiber types in metabolic syndrome subjects. Muscle insulin receptor expression increased in both groups, and GLUT4 expression increased in the metabolic syndrome subjects. The excess phosphorylation of insulin receptor substrate 1 (IRS-1) at Ser337 in metabolic syndrome muscle tended to increase further after training in spite of a decrease in total IRS-1. CONCLUSIONS: In the absence of weight loss, the cycle training of metabolic syndrome subjects resulted in enhanced mitochondrial biogenesis and increased the expression of insulin receptors and GLUT4 in muscle but did not decrease the insulin resistance. The failure for the insulin signal to proceed past IRS-1 tyrosine phosphorylation may be related to excess serine phosphorylation at IRS-1 Ser337, and this is not ameliorated by 8 wk of endurance exercise training.

Slow-twitch fiber proportion in skeletal muscle correlates with insulin responsiveness.

CONTEXT: The metabolic syndrome, characterized by central obesity with dyslipidemia, hypertension, and hyperglycemia, identifies people at high risk for type 2 diabetes. OBJECTIVE: Our objective was to determine how the insulin resistance of the metabolic syndrome is related to muscle fiber composition. DESIGN: Thirty-nine sedentary men and women (including 22 with the metabolic syndrome) had insulin responsiveness quantified using euglycemic clamps and underwent biopsies of the vastus lateralis muscle. Expression of insulin receptors, insulin receptor substrate-1, glucose transporter 4, and ATP synthase were quantified with immunoblots and immunohistochemistry. PARTICIPANTS AND SETTING: Participants were nondiabetic, metabolic syndrome volunteers and sedentary control subjects studied at an outpatient clinic. MAIN OUTCOME MEASURES: Insulin responsiveness during an insulin clamp and the fiber composition of a muscle biopsy specimen were evaluated. RESULTS: There were fewer type I fibers and more mixed (type IIa) fibers in metabolic syndrome subjects. Insulin responsiveness and maximal oxygen uptake correlated with the proportion of type I fibers. Insulin receptor, insulin receptor substrate-1, and glucose transporter 4 expression were not different in whole muscle but all were significantly less in the type I fibers of metabolic syndrome subjects when adjusted for fiber proportion and fiber size. Fat oxidation and muscle mitochondrial expression were not different in the metabolic syndrome subjects. CONCLUSION: Lower proportion of type I fibers in metabolic syndrome muscle correlated with the severity of insulin resistance. Even though whole muscle content was normal, key elements of insulin action were consistently less in type I muscle fibers, suggesting their distribution was important in mediating insulin effects.

Impaired muscle AMPK activation in the metabolic syndrome may attenuate improved insulin action after exercise training.

CONTEXT: Strength training induces muscle remodeling and may improve insulin responsiveness. OBJECTIVE: This study will quantify the impact of resistance training on insulin sensitivity in subjects with the metabolic syndrome and correlate this with activation of intramuscular pathways mediating mitochondrial biogenesis and muscle fiber hypertrophy. DESIGN: Ten subjects with the metabolic syndrome (MS) and nine sedentary controls underwent 8 wk of supervised resistance exercise training with pre- and posttraining anthropometric and muscle biochemical assessments. SETTING: Resistance exercise training took place in a sports laboratory on a college campus. MAIN OUTCOME MEASURES: Pre- and posttraining insulin responsiveness was quantified using a euglycemic clamp. Changes in expression of muscle 5-AMP-activated protein kinase (AMPK) and mammalian target of rapamycin (mTOR) pathways were quantified using immunoblots. RESULTS: Strength and stamina increased in both groups. Insulin sensitivity increased in controls (steady-state glucose infusion rate = 7.0 ± 2.0 mg/kg · min pretraining training vs. 8.7 ± 3.1 mg/kg · min posttraining; P < 0.01) but did not improve in MS subjects (3.3 ± 1.3 pre vs. 3.1 ± 1.0 post). Muscle glucose transporter 4 increased 67% in controls and 36% in the MS subjects. Control subjects increased muscle phospho-AMPK (43%), peroxisome proliferator-activated receptor γ coactivator 1α (57%), and ATP synthase (60%), more than MS subjects (8, 28, and 21%, respectively). In contrast, muscle phospho-mTOR increased most in the MS group (57 vs. 32%). CONCLUSION: Failure of resistance training to improve insulin responsiveness in MS subjects was coincident with diminished phosphorylation of muscle AMPK, but increased phosphorylation of mTOR, suggesting activation of the mTOR pathway could be involved in inhibition of exercise training-related increases in AMPK and its activation and downstream events.

Brain glucose transporter (Glut3) haploinsufficiency does not impair mouse brain glucose uptake.

Mouse brain expresses three principal glucose transporters. Glut1 is an endothelial marker and is the principal glucose transporter of the blood-brain barrier. Glut3 and Glut6 are expressed in glial cells and neural cells. A mouse line with a null allele for Glut3 has been developed. The Glut3(-/-) genotype is intrauterine lethal by 7days post-coitis, but the heterozygous (Glut3(+/-)) littermate survives, exhibiting rapid post-natal weight gain, but no seizures or other behavioral aberrations. At 12weeks of age, brain uptake of tail vein-injected ((3))H-2-deoxy glucose in Glut3(+/-) mice was not different from Glut3(+/+) littermates, despite 50% less Glut3 protein expression in the brain. The brain uptake of injected ((18))F-2-fluoro-2-deoxy glucose was similarly not different from Glut3(+/-) littermates in the total amount, time course, or brain imaging in the Glut3(+/-) mice. Glut1 and Glut6 protein expressions evaluated by immunoblots were not affected by the diminished Glut3 expression in the Glut3(+/-) mice. We conclude that a 50% decrease in Glut3 is not limiting for the uptake of glucose into the mouse brain, since Glut3 haploinsufficiency does not impair brain glucose uptake or utilization.

Cycle training increased GLUT4 and activation of mammalian target of rapamycin in fast twitch muscle fibers.

PURPOSE: To determine whether cycle training of sedentary subjects would increase the expression of the principle muscle glucose transporters, six volunteers completed 6 wk of progressively increasing intensity stationary cycle cycling. METHODS: In vastus lateralis muscle biopsies, changes in expression of GLUT1, GLUT4, GLUT5, and GLUT12 were compared using quantitative immunoblots with specific protein standards. Regulatory pathway components were evaluated by immunoblots of muscle homogenates and immunohistochemistry of microscopic sections. RESULTS: GLUT1 was unchanged, GLUT4 increased 66%, GLUT12 increased 104%, and GLUT5 decreased 72%. A mitochondrial marker (cytochrome c) and regulators of mitochondrial biogenesis (peroxisome proliferator-activated receptor gamma coactivator 1 alpha and phospho-5'-adenosine monophosphate-activated protein kinase) were unchanged, but the muscle hypertrophy pathway component, phospho-mammalian target of rapamycin (mTOR), increased 83% after the exercise program. In baseline biopsies, GLUT4 by immunohistochemical techniques was 37% greater in Type I (slow twitch, red) muscle fibers, but the exercise training increased GLUT4 expression in Type II (fast twitch, white) fibers by 50%, achieving parity with the Type I fibers. Baseline phospho-mTOR expression was 50% higher in Type II fibers and increased more in Type II fibers (62%) with training but also increased in Type I fibers (34%). CONCLUSION: Progressive intensity stationary cycle training of previously sedentary subjects increased muscle insulin-responsive glucose transporters (GLUT4 and GLUT12) and decreased the fructose transporter (GLUT5). The increase in GLUT4 occurred primarily in Type II muscle fibers, and this coincided with activation of the mTOR muscle hypertrophy pathway. There was little impact on Type I fiber GLUT4 expression and no evidence of change in mitochondrial biogenesis.

Insulin-stimulated translocation of glucose transporter (GLUT) 12 parallels that of GLUT4 in normal muscle.

CONTEXT: GLUT4 is the predominant glucose transporter isoform expressed in fat and muscle. In GLUT4 null mice, insulin-stimulated glucose uptake into muscle was diminished but not eliminated, suggesting that another insulin-sensitive system was present. OBJECTIVE: This study was intended to determine whether insulin caused GLUT12 translocation in muscle. DESIGN: Six normal volunteers had muscle biopsies before and after euglycemic insulin infusions. SETTING: Infusions and biopsies were performed in an outpatient clinic. PARTICIPANTS: Subjects were nonobese, young adults with no family history of diabetes. MAIN OUTCOME MEASURES: GLUT12, GLUT4, and GLUT1 proteins were quantified in muscle biopsy fractions. Cultured myoblasts were used to determine whether GLUT12 translocation was phosphatidyl inositol-3 kinase (PI3-K)-dependent. INTERVENTION: Insulin was infused at 40 mU/m(2) x min for 3 h. RESULTS: In human muscle, insulin caused a shift of a portion of GLUT12 from intracellular low-density microsomes to the plasma membrane (PM) fraction (17% in PM at baseline, 38% in PM after insulin). Insulin increased GLUT4 in PM from 13 to 42%. GLUT1 was predominantly in the PM fractions at baseline and did not change significantly after insulin. L6 myoblasts in culture also expressed and translocated GLUT12 in response to insulin, but inhibiting PI3-K prevented the translocation of GLUT12 and GLUT4. CONCLUSIONS: Insulin causes GLUT12 to translocate from an intracellular location to the plasma membrane in normal human skeletal muscle. Translocation of GLUT12 in cultured myoblasts was dependent on activation of PI3-K. GLUT12 may have evolutionarily preceded GLUT4 and now provides redundancy to the dominant GLUT4 system in muscle.

IGF-1 controls GLUT3 expression in muscle via the transcriptional factor Sp1.

Glucose transporter 3 (GLUT3), while first found in human fetal muscle, is predominantly expressed in brain and neural tissue. By several independent techniques we have previously shown that GLUT3 is expressed in human skeletal muscle cells. The structure of the human GLUT3 gene has not been previously reported nor has there been any evaluation of the 5'-untranslated region (UTR). To this end, we have cloned and sequenced the human GLUT3 gene. Insulin-like growth factor-1 (IGF-1) increased endogenous Glut3 protein in cultured L6 myotubes, and similarly stimulated luciferase activity in a construct of the human GLUT3 5'-UTR linked to a luciferase reporter gene. Actinomycin D, an inhibitor of mRNA synthesis, prevented IGF-1 stimulation of Glut3 protein. Transfection of L6 cells with Sp1 increased Glut3 and augmented IGF-1 stimulation of Glut3 expression. Knockdown of Glut3 expression in cultured L6 muscle cells using small interference RNA (siRNA) specific for Glut3 significantly reduced myocyte glucose uptake. DNAse footprinting and gel shift assays showed Sp1 specifically bound to the human GLUT3 5'-UTR. Substitution mutants of the human GLUT3 5'-UTR luciferase construct indicated that only one of three Sp1 site clusters was involved in IGF-1 action. These data, using both a human GLUT3 5'-UTR construct and L6 cells' endogenous promoter, suggest that IGF-1 plays a role in maintaining muscle GLUT3 expression and basal glucose uptake via the transcriptional factor Sp1.

Overexpression of GLUT5 in diabetic muscle is reversed by pioglitazone.

OBJECTIVE: This study was undertaken to quantify the expression of muscle GLUT in type 2 diabetes and to determine if treatment with an insulin-enhancing thiazolidenedione drug, pioglitazone, would alter its expression. RESEARCH DESIGN AND METHODS: Twelve patients with type 2 diabetes were randomly assigned to treatment with either pioglitazone or placebo in a double-blinded 8-week protocol. Protein and mRNA for GLUT4 and GLUT5 were quantified in muscle homogenates from biopsies of vastus lateralis before and after treatment. The five additional GLUT family isoforms expressed in muscle had mRNA quantified in these samples. RESULTS: Baseline and posttreatment repeat measurements of GLUT4 protein were not different from control measurements. Compared with normal subjects, GLUT5 protein increased 2.5-fold, and GLUT5 mRNA was 82% higher in the pretreatment samples from the diabetic subjects. Concentrations of mRNA for the six other GLUTs (GLUT1, GLUT3, GLUT4, GLUT8, GLUT11, and GLUT12) were not different from control subjects before or after treatment. The proportion of type I (red) fibers (46%) in diabetic muscle was not affected by pioglitazone treatment. Pioglitazone treatment decreased muscle GLUT5 mRNA and protein by 52 and 40%, respectively, whereas placebo did not alter GLUT5 expression. Both red and white fibers had higher GLUT5 expression in the baseline diabetic muscle samples, and a pioglitazone-related decrease in GLUT5 protein also occurred in both. CONCLUSIONS: GLUT5 was dramatically increased in diabetic muscle, and pioglitazone treatment reversed this overexpression. The role of this fructose transporter expression in the insulin-enhancing effect of pioglitazone in muscle is unclear.