Some hormonal factors (hypophysectomy, castration and testosterone administration) modifying the course of "compensatory" muscle hypertrophy in the rat.

1. Maximum compensatory hypertrophy of the soleus and plantaris muscle in male rats is attained seven days after tenotomy of the gastrocnemius muscle (39% and 9% respectively). When tenotomy of the gastrocnemius was performed seven days ater hypophysectomy, hypertrophy in these two muscles was aproximately half that found in control animals. 2. After 81-day castration of young male rats the weight of the saleus and plantaris was reduced and hypertrophy following tenotomy of the gastrocneumius muscle did not develop. 3. Chronically castrated rats received testosterone two weeks prior to tenotomy of the gastrocnemius and a week during the muscle hypertrophy phase. Hypertrophy of the soleus in castrated rats which had received testosterone seven days after tenotomy of the gastrocnemius was 25% as compared with muscles of castrated animals. The corresponding value in the plantaris muscle was 10%. 4. These results indicate that even calf muscles of the rat, namely the soleus and plantaris muscles, are significantly affected by testosterone under these conditions, although it is not, as yet, clear whether its action is direct or indirect.     

Adaptation in synergistic muscles to soleus and plantaris muscle removal in the rat hindlimb.

Although the soleus muscle comprises only 6% of the ankle plantar flexor mass in the rat, a major role in stance and walking has been ascribed to it. The purpose of this study was to determine if removal of the soleus muscle would result in adaptations in the remaining gastrocnemius and plantaris muscles due to the new demands for force production imposed on them during stance or walking. A second purpose was to determine whether the mass or the fiber type of the muscle(s) removed was a more important determinant of compensatory adaptations. Male Sprague-Dawley rats underwent bilateral removal of soleus muscle, plantaris muscle, or both muscles. For comparison, compensatory hypertrophy was induced in soleus and plantaris muscles by gastrocnemius muscle ablation. After forty days, synergist muscles remaining intact were removed. Mass, and oxidative, glycolytic, and contractile enzyme activities were determined. Despite its role in stance and slow walking, removal of the soleus muscle did not elicit a measurable alteration in muscle mass, or in citrate synthase, lactate dehydrogenase, or myofibrillar ATPase activity in gastrocnemius or plantaris muscles. Similarly, removal of the plantaris muscle, or soleus and plantaris muscles, had no effect on the gastrocnemius muscle, suggesting that this muscle was able to easily meet the new demands placed on it. These results suggest that amount of muscle mass removed, rather than fiber type, is the most important stimulus for compensatory hypertrophy. They also suggest that slow-twitch motor units in the gastrocnemius muscle play an important role during stance and locomotion in the intact animal.     

Antioxidant capacity in rat skeletal muscle tissues determined by electron spin resonance

The amount of radical scavenging activity in muscle is unknown. The present study examines whether electron spin resonance (ESR) could measure and distinguish antioxidant capacity in muscle with different contractile and metabolic characteristics. Specimens of the soleus, plantaris, gastrocnemius (deep/surface portions), heart and diaphragm were obtained from female Wistar rats (n=7; 12 weeks old). Scavenging activity against superoxide anions in these specimens were determined by ESR using a spin-trapping chemical (5,5-dimethyl-1-pyrroline-N-oxide). The ESR signal intensity of reaction mixtures containing muscle tissues was significantly lower in the heart, soleus, diaphragm and deep portion of the gastrocnemius than in the plataris and surface portion of the gastrocnemius. Thus, the amount of scavenging activity converted into superoxide dismutase activity was the highest in the heart, and higher in the soleus, diaphragm and deep portion of the gastrocnemius than in other muscles (ANOVA, P<0.01). In addition, scavenging activity significantly correlated with citrate synthase activity (r=0.72, P<0.01, n=42) and myoglobin content (r=0.63, P<0.01, n=42). These findings suggested that ESR and spin-trapping can be detect differences in free radical scavenging activity among muscle tissues with different metabolic characteristics.     

Effects of exercise in normoxia and acute hypoxia on respiratory muscle metabolites

We determined changes in rat plantaris, diaphragm, and intercostal muscle metabolites following exercise of various intensities and durations, in normoxia and hypoxia (FIO2 = 0.12). Marked alveolar hyperventilation occurred during all exercise conditions, suggesting that respiratory muscle motor activity was high. [ATP] was maintained at rest levels in all muscles during all normoxic and hypoxic exercise bouts, but at the expense of creatine phosphate (CP) in plantaris muscle and diaphragm muscle following brief exercise at maximum O2 uptake (VO2max) in normoxia. In normoxic exercise plantaris [glycogen] fell as exercise exceeded 60% VO2max, and was reduced to less than 50% control during exhaustive endurance exercise (68% VO2max for 54 min and 84% for 38 min). Respiratory muscle [glycogen] was unchanged at VO2max as well as during either type of endurance exercise. Glucose 6-phosphate (G6P) rose consistently during heavy exercise in diaphragm but not in plantaris. With all types of exercise greater than 84% VO2max, lactate concentration ([LA]) in all three muscles rose to the same extent as arterial [LA], except at VO2max, where respiratory muscle [LA] rose to less than half that in arterial blood or plantaris. Exhaustive exercise in hypoxia caused marked hyperventilation and reduced arterial O2 content; glycogen fell in plantaris (20% of control) and in diaphragm (58%) and intercostals (44%). We conclude that respiratory muscle glycogen stores are spared during exhaustive exercise in the face of substantial glycogen utilization in plantaris, even under conditions of extreme hyperventilation and reduced O2 transport. This sparing effect is due primarily to G6P inhibition of glycogen phosphorylase in diaphragm muscle. The presence of elevated [LA] in the absence of glycogen utilization suggests that increased lactate uptake, rather than lactate production, occurred in the respiratory muscles during exhaustive exercise.     

Skeletal muscle hypertrophy in rats having growth hormone-secreting tumor

Plantaris muscle hypertrophy resulting from surgical ablation of the synergistic gastrocnemius muscle was compared between nontumor- and GH3 tumor-bearing rat groups (n = 8-10). GH3 cells (10(6)) were subcutaneously injected into 150-g female Wistar-Furth rats to initiate the tumor. After 17 days, the tumor-bearing rats gained 5.7 g body wt/day compared with 2.0 for the nontumor-bearing rats. The left gastrocnemius muscle was surgically removed from both nontumor and tumor groups. The gastrocnemius was removed from the tumor group after an increased growth rate was achieved. Seven days after surgery, the animals were killed and plantaris muscles were removed. The wet weight of the left plantaris muscle increased 45.6 and 44.0% over the unoperated contralateral control (right side) in the nontumor and tumor groups, respectively. The right control plantaris muscle in the tumor group was 63% heavier than the right control plantaris from the nontumor group; however, the proportion of body weight for plantaris was similar between the two groups. The effect of gastrocnemius ablation and tumor treatment on plantaris weight was additive, and the percent increase over the unoperated contralateral control side was similar between the two groups. These data demonstrate that skeletal muscle hypertrophy occurs in adult animals in which growth has been stimulated by a growth hormone-secreting tumor and could suggest that the muscle growth response caused by the tumor is operating by a mechanism different than work-induced hypertrophy.     

Metabolic alterations in skeletal muscle of chronically streptozotocin-diabetic rats

This study was accomplished to determine the effects of chronic streptozotocin diabetes and insulin treatment on selected enzymes and substrates used in energy transduction in muscles composed of different muscle fiber types. Triglyceride concentration in all the muscles of diabetic rats was significantly elevated. Glycogen and protein concentrations were unchanged. The enzyme activities of hexokinase and alanine aminotransferase were significantly reduced and 3-hydroxyacyl-CoA dehydrogenase increased in all the muscles. Declines in phosphofructokinase, lactate dehydrogenase, citrate synthase, and succinate dehydrogenase activities were found in the red gastrocnemius and plantaris. Glycerol-3-phosphate dehydrogenase activity was lower than normal in the red gastrocnemius. Insulin treatment to the diabetic rats returned the altered triglyceride content and enzyme activities to normal, with exception of the lower alanine aminotransferase activity in the red gastrocnemius and plantaris. However, this enzyme was significantly ameliorated when compared with the untreated diabetic rats. The findings show that hypoinsulinism has a differential effect on the enzymatic profile of the different skeletal muscle fiber types, with those of the red gastrocnemius being most severely affected. Insulin treatment returned the enzymatic profile of the fiber types in diabetic rats to essentially normal.     

Morphometry, ultrastructure, myosin isoforms, and metabolic capacities of the "mini muscles" favoured by selection for high activity in house mice

Prolonged selective breeding of mice (Mus musculus) for high levels of voluntary wheel running has favoured an unusual phenotype ("mini muscles"), apparently caused by a single Mendelian recessive allele, in which most hind-limb muscles are markedly reduced in mass, but have increased mass-specific activities of mitochondrial enzymes. We examined whether these changes reflect changes in fibre size, number or ultrastructure in normal and "mini-muscle" mice within the two (of four) selectively bred lines (lab designations L3 and L6) that exhibit the phenotype at generations 26 and 27. In both lines, the gastrocnemius and plantaris muscles are smaller in mass (by >50% and 20%, respectively) in affected individuals. The mass-specific activities of mitochondrial enzymes in the gastrocnemius and plantaris muscles were increased in the mini phenotype in both lines, with stronger effects in the gastrocnemius muscle. In the gastrocnemius, the % myosin heavy chain (MHC) IIb was reduced by 50% in L3 and by 30% in L6, whereas the % MHC IIa and I were higher, particularly in L3. Fibre number in the plantaris muscle did not significantly differ between mini and normal muscles, although muscle mass was a significant positive correlate of fibre number. Small fibres were more abundant in mini than normal muscles in L3. Mitochondrial volume density was significantly higher in mini than normal muscle fibres in L3, but not in L6. Microscopy revealed a surprising attribute of the mini muscles: an abundance of small, minimally differentiated, myofibril-containing cells positioned in a disorderly fashion, particularly in the surface layer. We hypothesise that these unusual cells may be satellite cells or type IIb fibres that did not complete their differentiation. Together, these observations suggest that mice with the mini phenotype have reduced numbers of type IIb fibres in many of their hind-limb muscles, leading to a decrease in mass and an increase in mass-specific aerobic capacity in muscles that typically have a high proportion of type IIb fibres. Moreover, the several statistically significant interactions between muscle phenotype and line indicate that the effect of the underlying allele is altered by genetic background.     

Diaphragm muscle and its feed artery after chronic respiratory airway obstruction in rats

A chronic respiratory load was produced in Wistar rats by tracheal binding to produce a twofold increase of pleural pressure oscillation amplitude during respiration. Eight weeks after the surgery, a higher proportion of type-I muscle fibers (MFI) in the costal diaphragm along with a greater MFI cross-section area and a higher succinate dehydrogenase activity in MFII in the lumbar diaphragm were observed. During recording the mechanical activity of ring preparations of diaphragm arteries under isometric conditions, an increase in endothelium-dependent relaxation was found, whereas endothelium-independent relaxation and arterial reactivity to noradrenaline did not change. Tracheal binding did not produce any changes of MF in the gastrocnemius muscle, but endothelium-dependent relaxation of gastrocnemius feed arteries was reduced. We conclude that chronic respiratory load affects the endothelial function in diaphragm arteries in a manner favorable for blood flow control in the diaphragm. Functional alterations in gastrocnemius arteries may be associated with the reduced locomotor activity of operated rats.     

Changes in the diaphragm muscle and its feed artery after chronic respiratory airway obstruction in rats

A chronic respiratory load was produced in Wistar rats by tracheal binding to produce a twofold increase of pleural pressure oscillation amplitude during respiration. Eight weeks after the surgery, a higher proportion of type-I muscle fibers (MFI) in the costal diaphragm along with a greater MFI cross-section area and a higher succinate dehydrogenase activity in MFII in the crural diaphragm were observed. During recording the mechanical activity of ring preparations of diaphragm arteries under isometric conditions, an increase in endothelium-dependent relaxation was found, whereas endothelium-independent relaxation and arterial reactivity to noradrenaline did not change. Tracheal binding did not produce any changes of MF in the gastrocnemius muscle, but endothelium-dependent relaxation of gastrocnemius feed arteries was reduced. We conclude that chronic respiratory load affects the endothelial function in diaphragm arteries in a manner favorable for blood flow control in the diaphragm. Functional alterations in gastrocnemius arteries may be associated with the reduced locomotor activity of operated rats.     

Myosin isoform expression and MAFbx mRNA levels in hibernating golden-mantled ground squirrels (Spermophilus lateralis)

Hibernating mammals present many unexplored opportunities for the study of muscle biology. The hindlimb muscles of a small rodent hibernator (Spermophilus lateralis) atrophy slightly during months of torpor, representing a reduction in the disuse atrophy commonly seen in other mammalian models. How torpor affects contractile protein expression is unclear; therefore, we examined the myosin heavy-chain (MHC) isoform profile of ground squirrel skeletal muscle before and after hibernation. Immunoblotting was performed first to identify the MHC isoforms expressed in this species. Relative percentages of MHC isoforms in individual muscles were then measured using SDS-PAGE (sodium dodecyl sulfate-polyacrylamide gel electrophoresis). The soleus and diaphragm did not display differences in isoforms following hibernation, but we found minor fast-to-slow isoform shifts in MHC protein in the gastrocnemius and plantaris. These subtle changes are contrary to those predicted by other models of inactivity but may reflect the requirement for shivering thermogenesis during arousals from torpor. We also measured mRNA expression of the Muscle Atrophy F-box (MAFbx), a ubiquitin ligase important in proteasome-mediated proteolysis. Expression was elevated in the hibernating gastrocnemius and the plantaris but was not associated with atrophy. Skeletal muscle from hibernators displays unusual plasticity, which may be a combined result of the intense activity during arousals and the reduction of metabolism during torpor.     

Work overload induced changes in fast and slow skeletal muscle myosin heavy chain gene expression

Work induced hypertrophy of the slow postural soleus and the fast phasic plantaris muscles was produced by tenotomy of the synergistic gastrocnemius muscle. Increases in weight of both muscles were associated with proportionately even larger increases in total RNA and mRNA levels. Alterations in levels of specific myosin heavy chain (MHC) isoform mRNAs were measured using the slot blot procedure with radioactively labelled oligonucleotides as probes. Type 1 MHC gene expression was unaffected in both muscles by work overload, whereas type 2a was deinduced in the soleus and type 2b was deinduced in the plantaris. The neonatal MHC gene was transiently reinduced in the plantaris.     

Morphological adaptation of capillary network in compensatory hypertrophied rat plantaris muscle

The aim of this study was to examine the morphological adaptation of the capillary network in hypertrophied plantaris muscles by examining both capillary numbers and luminal circumferences. Hypertrophy of the plantaris muscle was induced by myectomy of the gastrocnemius muscle. This hypertrophy was characterised by increases in muscle mass and fibre cross-sectional area. All capillary parameters were determined using morphometric methods in perfusion-fixed plantaris muscle. Increased capillary-to-fibre ratio was observed in the overloaded plantaris muscle while no change was observed in the capillary luminal circumference. No differences were observed in the capillary density and the capillary-to-fibre perimeter ratio of the normal and the hypertrophied plantaris muscle. These results indicated that chronic overload-induced neocapillarization, but not enlargement of capillary luminal circumference, contributed to the prevention of decreases in the capillary-to-fibre perimeter ratio in the plantaris muscle in the hypertrophied process. Accepted: 13 August 1996     

Protein synthesis in skeletal muscle of the perfused rat hemicorpus compared with rates in the intact animal.

The rate of protein synthesis was measured in muscles of the perfused rat hemicorpus, and values were compared with rates obtained in whole animals. In gastrocnemius muscle of fed rats the rate of synthesis measured in the hemicorpus was the same as that in the whole animal. However, in plantaris, quadriceps and soleus muscles rates were higher in the hemicorpus than those in vivo. In the hemicorpus, starvation for 1 day decreased the rate of protein synthesis in gastrocnemius and plantaris muscles, in parallel with decreases in the RNA content, but the soleus remained unaffected. Similar effects of starvation were observed in vivo, so that the relationships between rates in vivo and in the hemicorpus were the same as those in fed rats. Proteins of quadriceps and plantaris muscles were separated into sarcoplasmic and myofibrillar fractions. The rate of synthesis in the sarcoplasmic fraction of the hemicorpus from fed rats was similar to that in vivo, but synthesis in the myofibrillar fraction was greater. In the plantaris of starved rats the rates of synthesis in both fractions were lower, but the relationships between rates measured in vivo and in the perfused hemicorpus were similar to those seen in fed rats. The addition of insulin to the perfusate of the hemicorpus prepared from 1-day-starved animals increased the rates of protein synthesis per unit of RNA in gastrocnemius and plantaris muscles to values above those seen in fed animals when measured in vivo or in the hemicorpus. Insulin had no effect on the soleus. Overall, the rates of protein synthesis in the hemicorpus differed from those in vivo. However, the effect of starvation when measured in the whole animal was very similar to that measured in the isolated rat hemicorpus when insulin was omitted from the perfusate.     

The relation between size of motoneurons and their position in the cat spinal cord

Experiments were performed to test whether motoneurons in the plantaris and medial gastrocnemius muscles of the cat are arranged in the spinal cord according to their sizes. It was found that motoneurons are randomly distributed with respect to size in their motor nuclei. Evidence is also presented that motoneuron density in these pools is irregular, and that there is considerable variability of position of medial gastrocnemius and plantaris motor pools from animal to animal.     

Force-length properties and functional demands of cat gastrocnemius, soleus and plantaris muscles.

The purpose of this study was to measure isometric force-length properties of cat soleus, gastrocnemius and plantaris muscle-tendon units, and to relate these properties to the functional demands of these muscles during everyday locomotor activities. Isometric force-length properties were determined using an in situ preparation, where forces were measured using buckle-type tendon transducers, and muscle-tendon unit lengths were quantified through ankle and knee joint configurations. Functional demands of the muscles were assessed using direct muscle force measurements in freely moving animals. Force-length properties and functional demands were determined for soleus, gastrocnemius and plantaris muscles simultaneously in each animal. The results suggest that isometric force-length properties of cat soleus, gastrocnemius and plantaris muscles, as well as the region of the force-length relation that is used during everyday locomotor tasks, match the functional demands.     

Changes in protein turnover in hypertrophying plantaris muscles of rats: effect of fenbufen--an inhibitor of prostaglandin synthesis

Plantaris muscle of the right hind limb of rats was subjected to hypertrophic stimulus by section of the tendons of the right gastrocnemius muscle. The RNA and protein content and the fractional rate of protein synthesis were elevated both 3 and 7 days after operation compared both with the unoperated left limb and with sham-operated control rats. The rate of protein degradation, calculated from the difference between the fractional rates of protein synthesis and protein gain of the muscles, was elevated in the plantaris 3-7 days after tenotomy. Dietary administration of the drug fenbufen reduced the RNA content and the ratio of RNA:protein in muscles from control animals. In one group of tenotomised rats administration of fenbufen commenced 3 days before tenotomy and resulted in a reduction in the ratio RNA:protein of the muscles of the left limb 3 days after the operation. Four days later, i.e. 7 days after tenotomy, both the ratio RNA:protein and the fractional rate of protein synthesis were significantly reduced in the fenbufen treated rats. In spite of these effects, fenbufen did not impair the ability of the plantaris to hypertrophy since the drug also reduced the rate of protein degradation.     

Satellite cell proliferation is reduced in muscles of obese Zucker rats but restored with loading

The obese Zucker rat (OZR) is a model of metabolic syndrome, which has lower skeletal muscle size than the lean Zucker rat (LZR). Because satellite cells are essential for postnatal muscle growth, this study was designed to determine whether reduced satellite cell proliferation contributes to reduced skeletal mass in OZR vs. LZR. Satellite cell proliferation was determined by a constant-release 5-bromo-2-deoxyuridine (BrdU) pellet that was placed subcutaneously in each animal. Satellite cell proliferation, as determined by BrdU incorporation, was significantly attenuated in control soleus and plantaris muscles of the OZR compared with that shown in the LZR. To determine whether this attenuation of satellite cell activity could be rescued in OZR muscles, soleus and gastrocnemius muscles were denervated, placing a compensatory load on the plantaris muscle. In the LZR and the OZR after 21 days of loading, increases of approximately 25% and approximately 30%, respectively, were shown in plantaris muscle wet weight compared with that shown in the contralateral control muscle. The number of BrdU-positive nuclei increased similarly in loaded plantaris muscles from LZR and OZR. Myogenin, MyoD, and Akt protein expressions were lower in control muscles of OZR than in those of the LZR, but they were all elevated to similar levels in the loaded plantaris muscles of OZR and LZR. These data indicate that metabolic syndrome may reduce satellite cell proliferation, and this may be a factor that contributes to the reduced mass in control muscles of OZR; however, satellite cell proliferation can be restored with compensatory loading in OZR.     

Blood flow and glycogen use in hypertrophied rat muscles during exercise

Previous findings suggest that skeletal muscle that has enlarged as a result of removal of synergistic muscles has a similar metabolic capacity and improved resistance to fatigue compared with normal muscle. The purpose of the present study was to follow blood flow and glycogen loss patterns in hypertrophied rat plantaris plantaris and soleus muscles during treadmill exercise to provide information on the adequacy of perfusion of the muscles during in vivo exercise. Thirty days following surgical removal of gastrocnemius muscle, blood flows (determined with radiolabeled microspheres) and glycogen concentrations were determined in all of the ankle extensor muscles of experimental and sham-operated control rats during preexercise and after 5-6 min of treadmill exercise at 15 m/min. There were no differences (P greater than 0.05) in blood flows per unit mass or glycogen concentrations between control and hypertrophied plantaris or soleus muscles at either time, although both muscles were larger (P less than 0.05) in the experimental group (plantaris: 95%; soleus: 40%). None of the other secondary ankle extensor muscles (tibialis posterior, flexor digitorum longus or flexor hallicus longus) hypertrophied in response to removal of gastrocnemius. These results provide indirect evidence that O2 delivery in the enlarged muscles is not compromised during low-intensity treadmill exercise due to limited perfusion.     

Control of Ser2448 phosphorylation in the mammalian target of rapamycin by insulin and skeletal muscle load

We have investigated the effects of insulin, amino acids, and the degree of muscle loading on the phosphorylation of Ser(2448), a site in the mammalian target of rapamycin (mTOR) phosphorylated by protein kinase B (PKB) in vitro. Phosphorylation was assessed by immunoblotting with a phosphospecific antibody (anti-Ser(P)(2448)) and with mTAb1, an activating antibody whose binding is inhibited by phosphorylation in the region of mTOR that contains Ser(2448). Incubating rat diaphragm muscles with insulin increased Ser(2448) phosphorylation but did not change the total amount of mTOR. Insulin, but not amino acids, activated PKB, as evidenced by increased phosphorylation of both Ser(308) and Thr(473) in the kinase. Ser(2448) phosphorylation was also modulated by muscle-loading. Overloading the rat plantaris muscle by synergist muscle ablation, which promotes hypertrophy of the plantaris muscle, increased Ser(2448) phosphorylation. In contrast, unloading the gastrocnemius muscle by hindlimb suspension, which promotes atrophy of the muscle, decreased Ser(2448) phosphorylation, an effect that was fully reversible. Neither overloading nor hindlimb suspension significantly changed the total amount of mTOR. In summary, our results demonstrate that atrophy and hypertrophy of skeletal muscle are associated with decreases and increases in Ser(2448) phosphorylation, suggesting that modulation of this site may have an important role in the control of protein synthesis.