Differential regulation of glucose transporter activity and expression in red and white skeletal muscle.

Insulin-stimulated glucose transport activity and GLUT4 glucose transporter protein expression in rat soleus, red-enriched, and white-enriched skeletal muscle were examined in streptozotocin (STZ)-induced insulin-deficient diabetes. Six days of STZ-diabetes resulted in a nearly complete inhibition of insulin-stimulated glucose transport activity in perfused soleus, red, and white muscle which recovered following insulin therapy. A specific decrease in the GLUT4 glucose transporter protein was observed in soleus (3-fold) and red (2-fold) muscle which also recovered to control values with insulin therapy. Similarly, cardiac muscle displayed a marked STZ-induced decrease in GLUT4 protein that was normalized by insulin therapy. White muscle displayed a small but statistically significant decrease in GLUT4 protein (23%), but this could not account for the marked inhibition of insulin-stimulated glucose transport activity observed in this tissue. In addition, GLUT4 mRNA was found to decrease in red muscle (2-fold) with no significant alteration in white muscle. The effect of STZ-induced diabetes was time-dependent with maximal inhibition of insulin-stimulated glucose transport activity at 24 h in both red and white skeletal muscle and half-maximal inhibition at approximately 8 h. In contrast, GLUT4 protein in red and white muscle remained unchanged until 4 and 7 days following STZ treatment, respectively. These data demonstrate that red skeletal muscle displays a more rapid hormonal/metabolic-dependent regulation of GLUT4 glucose transporter protein and mRNA expression than white skeletal muscle. In addition, the inhibition of insulin-stimulated glucose transport activity in both red and white muscle precedes the decrease in GLUT4 protein and mRNA levels. Thus, STZ treatment initially results in a rapid uncoupling of the insulin-mediated signaling of glucose transport activity which is independent of GLUT4 protein and mRNA levels.     

Sarcolemmal carbonic anhydrase in red and white rabbit skeletal muscle

Sarcolemmal vesicles of white and red skeletal muscles of the rabbit were prepared by consecutive density gradient centrifugations in sucrose and dextran according to Seiler and Fleischer (1982, J. Biol. Chem. 257, 13,862-13,871). White and red muscle membrane fractions enriched in sarcolemma were characterized by high ouabain-sensitive Na+, K(+)-ATPase, by high Mg2(+)-ATPase activity, and by a high cholesterol content. Ca2(+)-ATPase activity, a marker enzyme for sarcoplasmic reticulum, was not detectable in the highly purified white and red muscle sarcolemmal fractions. White and red muscle sarcolemmal fractions exhibited no significant differences with regard to Na+, K(+)-ATPase, Mg2(+)-ATPase, and cholesterol. Specific activity of carbonic anhydrase in white muscle sarcolemmal fractions was 38 U.ml/mg and was 17.6 U.ml/mg in red muscle sarcolemma. Inhibition properties of sarcolemmal carbonic anhydrase were analyzed for acetazolamide, chlorzolamide, and cyanate. White muscle sarcolemmal carbonic anhydrase is characterized by inhibition constants, KI, toward acetazolamide of 4.6 X 10(-8) M, toward chlorzolamide of 0.75 X 10(-8) M, and toward cyanate of 1.3 X 10(-4) M. Red muscle sarcolemmal carbonic anhydrase is characterized by KI values toward acetazolamide of 8.1 X 10(-8) M, toward chlorzolamide of 6.3 X 10(-8) M, and toward cyanate of 0.81 X 10(-4) M. In contrast to the high specific carbonic anhydrase activities in sarcolemma, carbonic anhydrase activity in sarcoplasmic reticulum from white muscle varied between values of only 0.7 and 3.3 U.ml/mg. Carbonic anhydrase of red muscle sarcoplasmic reticulum ranged from 2.4 to 3.7 U.ml/mg.     

Respiratory capacity of developing chick red and white skeletal muscle

Oxygen consumption, cytochrome oxidase and succinoxidase activity was measured in samples of leg and breast muscle from chick embryos ranging in age from 11 to 19 days. Respiratory parameters increased significantly in both muscle groups during embryonic life. By the later stages of incubation, leg and breast muscles differed significantly in cytochrome and succinoxidase activity. Oxygen uptake between leg and breast muscles did not differ significantly during later development. The results suggest at least a partial pre-natal differentiation of skeletal muscle in the domestic fowl.     

Protein composition and function of red and white skeletal muscle mitochondria

Red and white muscles are faced with very different energetic demands. However, it is unclear whether relative mitochondrial protein expression is different between muscle types. Mitochondria from red and white porcine skeletal muscle were isolated with a Percoll gradient. Differences in protein composition were determined using blue native (BN)-PAGE, two-dimensional differential in gel electrophoresis (2D DIGE), optical spectroscopy, and isobaric tag for relative and absolute quantitation (iTRAQ). Complex IV and V activities were compared using BN-PAGE in-gel activity assays, and maximal mitochondrial respiration rates were assessed using pyruvate (P) + malate (M), glutamate (G) + M, and palmitoyl-carnitine (PC) + M. Without the Percoll step, major cytosolic protein contamination was noted for white mitochondria. Upon removal of contamination, very few protein differences were observed between red and white mitochondria. BN-PAGE showed no differences in the subunit composition of Complexes I-V or the activities of Complexes IV and V. iTRAQ analysis detected 358 mitochondrial proteins, 69 statistically different. Physiological significance may be lower: at a 25% difference, 48 proteins were detected; at 50%, 14 proteins were detected; and 3 proteins were detected at a 100%. Thus any changes could be argued to be physiologically modest. One area of difference was fat metabolism where four β-oxidation enzymes were ～25% higher in red mitochondria. This was correlated with a 40% higher rate of PC+M oxidation in red mitochondria compared with white mitochondria with no differences in P+M and G+M oxidation. These data suggest that metabolic demand differences between red and white muscle fibers are primarily matched by the number of mitochondria and not by significant alterations in the mitochondria themselves.     

Glycolytic enzyme activity in developing red and white muscle

The activities of the constant proportion enzymes of the Embden-Meyerhof chain (triose phosphate isomerase (TIM), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), phosphoglycerate kinase (PGK), phosphoglycerate mutase (PGM) and enolase (ENOL)), and the activity of lactic dehydrogenase (LDH) were studied in developing red (trapezius) and white (longissimus) muscles of the pig from a fetal stage to 24 weeks postnatal. Both muscles were differentiated by two weeks postnatal in the sense that they had reached the adult level of enzyme activity. Enzyme activities were two- to three-fold greater in the longissimus than in the trapezius. Enzyme activity ratios based on GAPDH were not consistent in the fetal and day 1 samples but were consistent during later stages of growth. Ratios of enzyme activity based on activity at 105 days gestation revealed that TIM, PGK and PGM are grouped and follow the same pattern, but GAPDH and ENOL are quite different from each other and from the pattern shown by TIM, PGK and PGM. The constant proportion concept in developing muscle is therefore questioned.     

Effects of catecholamine treatment on UDPG levels in white and red skeletal muscle

Adrenaline causes of 5 fold increase of glucose-6-phosphate and of glucose-1-phosphate both in the white extensor digitorum longus and in the red soleus (where the levels of the two sugar phosphates are significantly lower). On the other hand, UDPG levels--which are similar in the two muscles--are significantly decreased after adrenaline. It has been concluded that the levels of glucose-1-phosphate and of UDPG in muscle are not bound to change together.     

Metabolic differentiation of red and white skeletal muscle in vivo and in monolayer culture

Cytochrome oxidase, succinate oxidase and lactate dehydrogenase were compared in: (a) leg and breast muscle from 11-19-day-old chick embryos; and (b) 2, 6, 10 and 14-day-old primary cell cultures established from myoblasts of embryonic leg and breast muscle. Cytochrome oxidase, succinate oxidase and lactate dehydrogenase activities were higher (48.8, 65.4, 277.6%, respectively) in leg muscle after 19 days in ovo. Cytochrome and succinate oxidase activities were higher (111.3, 48.1%, respectively) in leg muscle cell cultures after 14 days in vitro. The data represent evidence for intrinsic developmental patterns for certain enzymes.     

In vitro studies of skeletal muscle membranes. Effects of denervation on the macromolecular components of cation transport in red and white skeletal muscle.

The effects of denervation on the macromolecular components of active monovalent cation transport in skeletal muscle have been studied using purified sarcolemma membranes. A comparison of membrane activities of fast-twitch, slow-twitch, and mixed-fiber muscles was made to determine what role, if any, the motor nerve has in regulating this important aspect of muscle metabolism. A dramatic increase in the basal sarcolemmal Mg++ ATPase activity (three- to fourfold) was found for both major muscle types. An increase in the ouabain-inhibitable (Na+ + K+)-stimulated enzyme was also found, but the effect was substantially less (1.5- to twofold). [3H]-ouabain binding, as an index of glycoside receptor sites, also increased (two- to threefold) midway in the course of denervation. On the other hand, the phosphorylated intermediate activity, a functional component of the transport system, clearly decreased over the same time course and remained below control values for the remainder of the course. This resulted in a two- to threefold increase in the turnover number, suggesting that active transport of cations should increase dramatically with denervation. The membrane protein patterns on SDS gels were less obvious than the changes observed in the functional components. The major effects appeared after only one week and seemed to be restricted to high molecular weight membrane proteins, especially in the 100,000 to 250,000 daltons range. This effect was more prominent in slow-twitch membranes with an apparent semiquantitative decrease in stain at 240,000 daltons. In gels of membranes from fast-twitch muscles a decreased stain in the range of 100,000 to 110,000 daltons occurred, and this became more obvious with longer periods of denervation. The results suggest that considerable influence on the macromolecular components of active cation transport in skeletal muscle is exerted by the motor nerve. No appreciable difference was found in this effect when the two major types of skeletal muscle, fast-twitch and slow-twitch, were compared, suggesting that motor nerve regulation of this membrane property is qualitatively the same.     

Effect of acute exercise on citrate synthase activity in untrained and trained human skeletal muscle

Maximal citrate synthase activity (CS) is routinely used as a marker of aerobic capacity and mitochondrial density in skeletal muscle. However, reported CS has been notoriously variable, even with similar experimental protocols and sampling from the same muscles. Exercise training has resulted in increases in CS ranging from 0 to 100%. Previously, it has been reported that acute exercise may significantly affect CS. To investigate the hypothesis that the large variation in CS that occurs with training is influenced by alterations during the exercise itself, we studied CS in human vastus lateralis both in the rested and acutely exercised state while trained and untrained (n = 6). Tissues obtained from four biopsies (untrained rested, untrained acutely exercised, trained rested, and trained acutely exercised) were analyzed spectrophotometrically for maximal CS. Exercise training measured in a rested state resulted in an 18.2% increase in CS (12.3 +/- 0.3 to 14.5 +/- 0.3 micromol x min(-1) x g tissue(-1), P < or = 0.05). However, even greater increases were recorded 1 h after acute exercise: 49.4% in the untrained state (12.3 +/- 0.3 to 18.3 +/- 0.5 micromol x min(-1) x g tissue(-1), P < or = 0.05) and 50.8% in the trained state (14.5 +/- 0.3 to 21.8 +/- 0.4 micromol x min(-1) x g tissue(-1), P < or = 0.05). Ultrastructural analysis, by electron microscopy, supported an effect of acute exercise with the finding of numerous swollen mitochondria 1 h after exercise that may result in greater access to the CS itself in the CS assay. In conclusion, although unexplained, the increased CS with acute exercise can clearly confound training responses and artificially elevate CS values. Therefore, the timing of muscle sampling relative to the last exercise session is critical when measuring CS and offers an explanation for the large variation in CS previously reported.     

Intrinsic differences of insulin receptor kinase activity in red and white muscle

The sensitivity and responsiveness of glucose uptake and glycogen synthesis to insulin are 3-4-fold greater in red than in white skeletal muscle (James, D. E., Jenkins, A. B., and Kraegen, E. W. (1985) Am. J. Physiol. 248, E567-E574). In the present study, the insulin receptor tyrosine kinase activity has been examined in red and white muscle of rats. Partially purified insulin receptors were obtained from muscle following solubilization in detergent, ultracentrifugation, and lectin affinity chromatography. Total insulin receptor number per gram of tissue was slightly higher in red (30%) than in white muscle. In contrast, basal and insulin-stimulated autophosphorylation, normalized for receptor number, were 2.3-fold higher in red muscle. A similar difference was observed in the ability of partially purified receptors to phosphorylate the exogenous substrate polyglutamate/tyrosine. The integrity of the insulin receptor preparation in the two fiber types was identical as determined by affinity cross-linking of [125I-TyrB26]insulin to the receptor. Mixing partially purified receptors from red and white muscle resulted in an additive response for exogenous substrate phosphorylation, suggesting that the difference in tyrosine kinase activity was not due to the presence of an inhibitor or activator. The results suggest that there are differences in the insulin receptors of red and white muscles that lead to discordance in their basal and insulin-stimulated intrinsic tyrosine kinase activity. The correlation between these differences and insulin action in red and white muscle supports the concept that the insulin receptor tyrosine kinase activity is involved in the initiation of insulin action.     

Functional and structural adaptations in skeletal muscle of trained athletes

Twitch contractile and ultrastructural characteristics of the human triceps surae were determined in six male strength-trained athletes, six endurance-trained athletes, six active controls, and seven sedentary controls of similar height and age. Twitch contraction time in the triceps surae complex was 20% longer in strength-trained and sedentary groups than in endurance-trained or active control groups. In the 15 subjects peak twitch torque and one-half relation time in the triceps surae were 22.6 +/- 7.9 N.m and 91.1 +/- 18.3 ms, respectively. Mean fiber area in the gastrocnemius was approximately 1.6-, 1.7-, and 2.5-fold greater in the active control, endurance-trained, and strength-trained groups, respectively, relative to the sedentary group. Despite these large differences in fiber areas, the fiber fractional volume of the sarcoplasmic reticulum-transverse tubule network averaged 3.38 +/- 0.86% and 5.50 +/- 0.94% in type I and type II fibers, respectively, in all subjects. The fractional fiber volume of cytoplasm and lipid were similar for all four groups. However, mitochondrial volume was approximately 30% lower in both fiber types of the strength-trained group relative to the other groups. This implies that with exercise-induced hypertrophy, the sarcoplasmic reticulum, cytoplasm, and lipid components increase proportionately with contractile protein, whereas the mitochondrial fraction does not. The proportion of type I fibers in the soleus, medial gastrocnemius, and lateral gastrocnemius was 75.2 +/- 8.3, 58.5 +/- 6.1, and 52.4 +/- 4.2%, respectively, and was similar in all subject groups. The results demonstrate that twitch duration is prolonged in strength-trained athletes relative to endurance athletes.(ABSTRACT TRUNCATED AT 250 WORDS)