Effect of hydrocortisone and dexamethasone on xylose uptake by isolated rat soleus muscle

To determine the effects of glucocorticoids on sugar uptake, xylose uptake by isolated rat soleus muscle of bilaterally adrenalectomized animals was studied. The results indicate that in vitro addition of 10⁻⁴ M hydrocortusine, dexamethasone or hydrocortisone sodium succinate had no inhibitory effect on basal xylose uptake. In the presence of both low and high medium insulin, the above steroids failed to inhibit insulin-stimulated uptake. When the concentration of hydrocortisone sodium succinate was increased to 10⁻² M, insulin-stimulated uptake was decreased. The results thus indicate that glucocorticoids at concentrations observed under physiological or pathological conditions do not inihibit basal or insulin-stimulated sugar uptake.     

Palmitic acid uptake by the rat soleus muscle in vitro.

Abstract: The rate of fatty acid uptake, oxidation, and deposition in skeletal muscles in relation to total and unbound to albumin fatty acids concentration in the medium were investigated in the incubated rat soleus muscle. An immunohistochemical technique was applied to demonstrate whether the albumin-bound fatty acid complex from the medium penetrates well within all areas of the muscle strips. It was found that the percentage of incorporation of palmitic acid into intramuscular lipids was fairly constant, independently of the fatty acid concentration in the medium, and amounted to 63-72% for triacylglycerols, 7-12% for diacylglycerols-monoacylglycerols, and 19-26% for phospholipids. Both palmitic acid incorporation into the muscle triacylglycerol stores and its oxidation to CO2 closely correlated with an increase in both total and unbound to albumin fatty acid concentrations in the incubation medium. Under conditions of increased total but constant unbound to albumin palmitic acid concentrations, the incorporation of palmitic acid into triacylglycerols and its oxidation to CO2 were also increased, but to a lower extent. This supports the hypothesis that the cellular fatty acid metabolism depends not only on the availability of fatty acids unbound to albumin, but also on the availability of fatty acids complexed to albumin.     

Beta-adrenergic effects on carbohydrate metabolism in the unweighted rat soleus muscle

The effects of insulin on carbohydrate metabolism in atrophied rat soleus muscle are increased after unweighting by tail-cast suspension. This work has been extended by testing the effect of unweighting on the response of carbohydrate metabolism to isoproterenol, a beta-adrenergic agonist. Isoproterenol promoted glycogen degradation more in the unweighted than in the weight-bearing soleus but showed no differences in the extensor digitorum longus, which is unresponsive to hindlimb unweighting. In soleus muscles depleted of glycogen, to avoid varied inhibitory effects of glycogen on glycogen synthesis, isoproterenol inhibited this process more in the unweighted muscle. Isoproterenol did not have a greater inhibitory effect on net uptake of 2-deoxy-D[1,2-3H]glucose by the unweighted muscle. Measurements of intracellular 2-deoxy-[3H]glucose 6-phosphate and 3-O-methyl-D-[1-3H]glucose, which cannot be phosphorylated, showed that isoproterenol inhibited glucose phosphorylation but not transport. This effect could be explained by an increase of glucose 6-phosphate, an inhibitor of hexokinase. At 100 microU insulin/ml but not at a lower amount (10 microU/ml), isoproterenol inhibited hexose phosphorylation more in the control than in the unweighted muscle. This result may be explained by greater insulin antagonism in the unweighted muscle owing to increased insulin sensitivity. However, insulin antagonism of isoproterenol stimulation of glycogenolysis or inhibition of glycogenesis was not altered by unweighting. Therefore, for some aspects of carbohydrate metabolism, the unweighted muscle has an increased response to beta-adrenergic activation, just as this muscle shows increased responses to insulin.     

Effect of hydrocortisone and dexamethasone on xylose uptake by isolated rat soleus muscle.

To determine the effects of glucocorticoids on sugar uptake, xylose uptake by isolated rat soleus muscle of bilaterally adrenalectomized animals was studied. The results indicate that in vitro addition of 10-4 M hydrocortisone, dexamethasone or hydrocortisone sodium succinate had no inhibitory effect on basal xylose uptake. In the presence of both low and high medium insulin, the above steroids failed to inhibit insulin-stimulated uptake. When the concentration of hydrocortisone sodium succinate was increased to 10-2 M, insulinstimulated uptake was decreased. The results thus indicate that glucocorticoids at concentrations observed under physiological or pathological conditions do not inhibit basal or insulin-stimulated sugar uptake.     

Myotoxic effects of clenbuterol in the rat heart and soleus muscle.

Myocyte-specific necrosis in the heart and soleus muscle of adult male Wistar rats was investigated in response to a single subcutaneous injection of the anabolic beta(2)-adrenergic receptor agonist clenbuterol. Necrosis was immunohistochemically detected by administration of a myosin antibody 1 h before the clenbuterol challenge and quantified by using image analysis. Clenbuterol-induced myocyte necrosis occurred against a background of zero damage in control muscles. In the heart, the clenbuterol-induced necrosis was not uniform, being more abundant in the left subendocardium and peaking 2.4 mm from the apex. After position (2.4 mm from the apex), dose (5 mg clenbuterol/kg), and sampling time (12 h) were optimized, maximum cardiomyocyte necrosis was found to be 1.0 +/- 0.2%. In response to the same parameters (i.e., 5 mg of clenbuterol and sampled at 12 h), skeletal myocyte necrosis was 4.4 +/- 0.8% in the soleus. These data show significant myocyte-specific necrosis in the heart and skeletal muscle of the rat. Such irreversible damage in the heart suggests that clenbuterol may be damaging to long-term health.     

Stimulatory and inhibitory effects of EDTA on sugar transport by rat soleus muscle

The uptake of D-[¹⁴C]xylose by rat soleus muscle was stimulated rapidly and transiently by brief exposure to EDTA (0.1–20 mM). EDTA also stimulated xylose uptake in the presence of insulin (0.1 U/ml). Prolonged exposure to EDTA (60 min) inhibited insulin-stimulated xylose uptake and depressed ¹²⁵I-insulin binding; these effects were associated with the lowering of muscle ATP. The stimulatory effect was abolished by the substitution of Ca-EDTA (or Mg-EDTA) for EDTA; Ca-EDTA did not eliminate the inhibitory effect. There was no inhibitory effect when Ca²⁺ (5 mM) was added along with Ca-EDTA, or when Zn-EDTA was used instead. There was no effect of EGTA (5 mM) on xylose uptake measured in the presence or absence of insulin. It is concluded (1) that the stimulatory effect of EDTA is most likely due to the chelation of Mg²⁺, (2) that the inhibitory effects of EDTA are due to the chelation of some metal ion whith a higher affinity for the chelator than either Ca²⁺ or Mg²⁺.     

Multiple effects of sulphydryl reagents on sugar transport by rat soleus muscle

Iodoacetate, over the range 0.2-2 mM, stimulated the uptake of D-xylose by rat soleus muscle and inhibited anaerobic lactate production by soleus muscle. Stimulation of sugar transport is considered to be due to the resultant fall in ATP. p-Chloromercuribenzene sulphonate (0.5-2 mM) stimulated xylose uptake to a lesser extent than iodoacetate and induced a proportionately smaller fall in ATP, consistent with the inhibitory effect of p-chloromercuribenzene sulphonate on lactate production. Under certain conditions, p-chloromercuribenzene sulphonate stimulated sugar transport without affecting the ATP level. This suggests that whereas p-chloromercuribenzene sulphonate can be expected to stimulate sugar transport through the lowering of muscle ATP, it may also act through some other mechanism. No stimulatory effect on xylose uptake was observed when muscles were exposed to N-ethylmaleimide (0.02-2 mM) either for brief (1 min) or more prolonged (30 min) periods. Because N-ethylmaleimide induced a marked fall in muscle ATP, it is surprising that N-ethylmaleimide did not stimulate sugar transport; in most experiments this inhibitor actually inhibited sugar transport. N-Ethylmaleimide inhibited the stimulation of sugar transport by 2,4-dinitrophenol and anoxia; this inhibitory effect appears to explain why N-ethylmaleimide itself did not stimulate sugar transport. p-Chloromercuribenzene sulphonate also inhibited 2,4-dinitrophenol-stimulated xylose uptake by a mechanism which seems similar to that of N-ethylmaleimide; this could explain in part the modest stimulatory effect of this inhibitor on muscle sugar transport.     

Chronic hypoxia increases insulin-stimulated glucose uptake in mouse soleus muscle

People living at high altitude appear to have lower blood glucose levels and decreased incidence of diabetes. Faster glucose uptake and increased insulin sensitivity are likely explanations for these findings: skeletal muscle is the largest glucose sink in the body, and its adaptation to the hypoxia of altitude may influence glucose uptake and insulin sensitivity. This study tested the hypothesis that chronic normobaric hypoxia increases insulin-stimulated glucose uptake in soleus muscles and decreases plasma glucose levels. Adult male C57BL/6J mice were kept in normoxia [fraction of inspired O? = 21% (Control)] or normobaric hypoxia [fraction of inspired O? = 10% (Hypoxia)] for 4 wk. Then blood glucose and insulin levels, in vitro muscle glucose uptake, and indexes of insulin signaling were measured. Chronic hypoxia lowered blood glucose and plasma insulin [glucose: 14.3 ± 0.65 mM in Control vs. 9.9 ± 0.83 mM in Hypoxia (P < 0.001); insulin: 1.2 ± 0.2 ng/ml in Control vs. 0.7 ± 0.1 ng/ml in Hypoxia (P < 0.05)] and increased insulin sensitivity determined by homeostatic model assessment 2 [21.5 ± 3.8 in Control vs. 39.3 ± 5.7 in Hypoxia (P < 0.03)]. There was no significant difference in basal glucose uptake in vitro in soleus muscle (1.59 ± 0.24 and 1.71 ± 0.15 μmol·g?1·h?1 in Control and Hypoxia, respectively). However, insulin-stimulated glucose uptake was 30% higher in the soleus after 4 wk of hypoxia than Control (6.24 ± 0.23 vs. 4.87 ± 0.37 μmol·g?1·h?1, P < 0.02). Muscle glycogen content was not significantly different between the two groups. Levels of glucose transporters 4 and 1, phosphoinositide 3-kinase, glycogen synthase kinase 3, protein kinase B/Akt, and AMP-activated protein kinase were not affected by chronic hypoxia. Akt phosphorylation following insulin stimulation in soleus muscle was significantly (25%) higher in Hypoxia than Control (P < 0.05). Neither glycogen synthase kinase 3 nor AMP-activated protein kinase phosphorylation changed after 4 wk of hypoxia. These results demonstrate that the adaptation of skeletal muscles to chronic hypoxia includes increased insulin-stimulated glucose uptake.     

Combined effects of tenotomy and denervation on the dynamic properties of soleus muscle of the rat

Isometric contraction time (CT), half relaxation time (1/2 RT), tetanus fusion frequency (TFF) and tetanus: twitch ratio (T : t ratio) were measured in the denervated (D) and tenotomized-denervated (TD) Soleus muscle of the rat. In D muscle there was an apparent speeding effect at the 2nd day after denervation, with a significant decrease of CT, which was followed by the usual slowing process of denervated muscle. In TD muscle, denervation was performed a week after tenotomy. Tenotomy "per se" was ineffective in modifying dynamic properties of muscle, but it accentuated the early shortening of CT caused by denervation, while reducing and delaying the subsequent slowing process. The results are discussed in the light of the hypothesis that muscle disuse has a speeding effect which counteracts the slowing effect of denervation, and/or that tenotomy modifies the effects of denervation by changing the pattern of fibrillation development.     

Exercise-induced adaptations of rat soleus muscle grafts

In female Wistar rats (n = 316) under pentobarbital sodium anesthesia, the soleus muscle was autografted with its nerve reimplanted. One purpose was to characterize the chronological development of graft innervation and recruitment during locomotion. Furthermore, we tested hypotheses regarding the efficacy of run conditioning of different intensities, durations, and postgrafting initiation times to alter mass and pyruvate-malate oxidation capacity of grafts. Choline acetyltransferase activity of grafts increased from 10% of control value at 7 days postgrafting to 55 and 100% at days 28 and 56, respectively. Running-induced glycogen depletion occurred in grafts; this is consistent with graft recruitment during locomotion. There was a threshold of conditioning intensity below which no improvements occurred and above which there were improvements. Spring (50 m/min) and endurance (30 m/min) conditioning of a duration of at least 28 days that was initiated at 28 or 56 days postgrafting increased mass of grafts by 30% compared with grafts from nonconditioned rats. Easy conditioning (15 m/min) had no effect on graft mass. Changes in graft total protein content paralleled those of mass. Oxidation capacity of grafts increased significantly with some conditioning protocols, but not to the same extent as mass. The exercise-induced adaptations should improve graft function in the host organism.     

Effects of oxygen deprivation on incubated rat soleus muscle

Isolated soleus muscle deprived of oxygen produces more lactate and alanine than oxygen-supplied muscle. Oxygenated muscle synthesized glutamine, while anoxic muscle used this amino acid. Oxygen deprivation decreased adenine nucleotides leading to the efflux of nucleosides. Protein synthesis and degradation responded differently to anoxia. Synthesis almost completely ceased, while proteolysis increased. Therefore, protein degradation in soleus muscle is enhanced when energy supplies and oxygen tension are low.     

Effect of protein synthesis inhibitors on xylose uptake and 125I-insulin binding by rat soleus muscle

Prolonged exposure (90–180 min) to cycloheximide (0.2 mg/ml), puromycin (0.2 mg/ml) or chloramphenicol (0.1 mg/ml) did not affect ¹²⁵I-insulin binding by rat soleus muscle. Chloramphenicol (2 mg/ml) depressed insulin binding and insulin-stimulated xylose uptake; these effects were attributed to the “toxic” effect of chloramphenicol on muscle ATP levels. Cycloheximide and puromycin inhibited insulin-stimulated xylose uptake without affecting ATP. Puromycin and chloramphenicol, but not cycloheximide, also inhibited basal sugar transport. This difference, and the rapid onset of all these inhibitory effects, suggest that they are not due to the inhibition of protein synthesis, but rather to some more direct effect on sugar transport itself.     

Growth hormone effects on creatine uptake by muscle in the hypophysectomized rat

Summary Specific radioactive enzyme assays were developed to measure the effect of growth hormone on kidney transamidinase and liver methyltransferase in the hypophysectomized rat. In contrast to minimal changes (20%) in liver methyltransferase, kidney transamidinase was decreased threefold in the hypophysectomized rat. Enzyme activities were equal to normal values in those rats receiving growth hormone for three days. The formation of creatine from radioactive precursors and the uptake of ¹⁴C-creatine in muscle was examined under these conditions. After injection of ¹⁴C-arginine in the hypophysectomized rat, the ¹⁴C-creatine content of muscle was greatly decreased compared to sham operated controls and the ¹⁴C-creatine content was normal after growth hormone administration. After injection of ¹⁴C-guanidoacetate and of ¹⁴C-creatine, the ¹⁴C-creatine content of muscle was decreased in the hypophysectomized rat, but was equal to sham control values in rats receiving growth hormone. These studies indicate that the uptake of newly synthesized creatine by muscle is impaired in the hypophysectomized rat and that growth hormone can have a role in controlling the rate of creatine uptake by muscle in addition to its effect on kidney transamidinase and to other factors involved in creatine metabolism.