Thyroid hormone response of slow and fast sarcoplasmic reticulum Ca2+ ATPase mRNA in striated muscle.

The thyroid status markedly influences the contractile function of muscle, and changes in the activity of the Ca2+ ATPase of the sarcoplasmic reticulum (SR) contribute to these alterations. Two separate genes encode the major isoforms of SR Ca2+ ATPase. In fast skeletal muscle, sarcoplasmic endoplasmic reticulum Ca2+ ATPase type 1 (SERCa1) presents the major isoform, whereas in slow skeletal muscle SERCa type 2 (SERCa2) predominates. Cardiac muscle contains only SERCa2. To examine the mechanisms responsible for changes in contractile function, we quantitated SERCa1 and SERCa2 mRNA levels in fast extensor digitorum longus muscle (EDL), slow soleus muscle, and cardiac muscle in rats of different thyroid status. Hypothyroidism led in soleus to a marked decrease in SERCa1 mRNA and SERCa2 mRNA levels, in cardiac muscle SERCa2 mRNA decreased markedly, as previously shown by us, and in EDL SERCa1 mRNA decreased. These findings are compatible with a hypothyroidism induced decrease in SR Ca2+ ATPase activity and a delay in muscle relaxation. In contrast, SERCa2 mRNA of EDL, representing only a small percent of total SERCa mRNA in this muscle, increased to 175% of control values. Muscle specific and SERCa gene specific changes also occur after acute triiodothyronine (T3) administration to hypothyroid rats. T3 does not induce a significant change in SERCa1 or SERCa2 mRNA levels in soleus, but in the heart SERCa2 mRNA increases about 3-fold. In EDL, T3 increases SERCa1 mRNA from a hypothyroid level of 59 +/- 6% to 138 +/- 4% of control values but SERCa2 mRNA is decreased to 75 +/- 5% of control levels.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of heart failure on skeletal muscle myofibrillar protein content, isoform expression and calcium sensitivity

BACKGROUND: Alterations in skeletal muscle with heart failure contribute to exercise intolerance and physical disability. The majority of studies to date have examined abnormalities in skeletal muscle oxidative capacity and mitochondrial function. In contrast, less information is available regarding the effect of heart failure on myofibrillar protein metabolism and function. To address this issue, we examined the effect of heart failure on skeletal muscle myofibrillar protein content, isoform distribution and Ca2+ sensitivity. METHODS: We measured skeletal muscle myosin heavy chain (MHC) and actin protein content and MHC isoform distribution in soleus (SOL), extensor digitorum longus (EDL), plantaris (PL) and diaphragm (DIA) muscles and myofibrillar Ca2+ sensitivity in EDL muscles from Dahl salt-sensitive rats with (high-salt fed: HS; n=10) or without heart failure (low-salt fed: LS; n=8) and assessed the relationship of these variables to markers of disease severity. RESULTS: No differences in muscle mass were found. Similarly, no differences in MHC (mean+/-SE; SOL: 1353+/-29 vs. 1247+/-52; EDL: 1471+/-31 vs. 1441+/-31; PL: 1207+/-66 vs. 1286+/-36; DIA: 1166+/-42 vs. 1239+/-26 AU/microg protein) or actin (EDL: 348+/-13 vs. 358+/-19; PL: 245+/-20 vs. 242+/-9; DIA: 383+/-9 vs. 376+/-17 AU/microg protein) protein content or the actin-to-MHC ratio were observed, with the exception of lower (P<0.01) actin content in the soleus of LS rats (352+/-7 vs. 310+/-8 AU/microg protein). MHC isoform expression (I, IIa, IIx, IIb) did not differ between groups in SOL (I: 89+/-1% vs. 85+/-2%; IIa: 11+/-1% vs. 15+/-2%), EDL (IIx: 43+/-10% vs. 38+/-10%; IIb: 57+/-10% vs. 62+/-10%), PL (I: 6+/-4% vs. 3+/-3%; IIa: 1+/-1% vs. 1+/-1%; IIx: 31+/-3% vs. 26+/-4%; IIb: 62+/-5% vs. 71+/-6%) or DIA (I: 43+/-6% vs. 36+/-6 %; IIa: 9+/-1% vs. 7+/-1%; IIx: 47+/-6% vs. 56+/-7%; IIb: 2+/-1% vs. 1+/-0.5%) muscles. Moreover, heart failure did not affect the Ca2+ sensitivity (i.e., pCa50) of extensor digitorum longus myofilaments (5.68+/-0.11 vs. 5.65+/-0.09). Finally, MHC and actin content, MHC isoform distribution and myofibrillar Ca2+ sensitivity were not related to markers of disease severity. CONCLUSIONS: Our results show that this animal model of heart failure is not characterized by alterations in the quantity or isoform distribution of key skeletal muscle myofibrillar proteins or the Ca2+ sensitivity of isometric force production. These findings suggest that alterations in skeletal muscle myofibrillar protein metabolism do not develop in parallel with myocardial failure in the Dahl salt-sensitive rat.     

Protein turnover in different types of skeletal muscle during experimental hyperthyroidism in rats

Experimental hyperthyroidism was induced in rats by daily ip injection of triiodothyronine (T3; 100 micrograms/100 g body weight) during 3 or 10 days. Protein synthesis and degradation were measured in incubated soleus and extensor digitorum longus (EDL) muscles by determining rate of tyrosine incorporation into protein and release of tyrosine to the incubation medium respectively. Protein synthesis was unaffected by T3 administration during 3 or 10 days. Protein breakdown was significantly increased in soleus but unchanged in EDL in the 3-days experiment. Following administration of T3 for 10 days proteolysis was increased in both muscles. Weight of the soleus muscle was reduced after T3 for 3 days. After 10 days weight and protein content were reduced in both muscles. The study demonstrated that reduced muscle protein content following administration of T3 was the result of increased proteolysis, not decreased protein synthesis. The results further indicate that slow muscle (soleus) is more sensitive to the effects of thyroid hormone than fast muscle (EDL).     

Effect of corticosteroids on respiratory muscle histopathology

Pathologic alterations induced by corticosteroid administration were evaluated in the respiratory muscles and compared to those in the peripheral skeletal muscles of the rabbit. Alterations in gross and microscopic pathology as well as histochemistry were determined in the diaphragm, intercostal, sternocleidomastoid, extensor digitorum longus, and soleus muscles following 3 wk of intramuscular cortisone injections. Corticosteroid administration induced significant pathologic changes in all the muscles except the soleus. Although gross pathologic changes were greatest in the extensor digitorum longus, microscopic changes were greatest in the diaphragm. Reductions in total muscle fiber volume were similar in the diaphragm, sternocleidomastoid, and extensor digitorum longus muscles. The composition of muscle fiber types and the number of fibers in a muscle were not altered in any of the muscles studied following corticosteroids. There was significant atrophy of individual muscle fibers in all the muscles except the soleus. In the diaphragm, corticosteroids induced atrophy of all fiber types, including type I fibers. Atrophy of type I fibers was not present in the peripheral skeletal muscles or the other respiratory muscles. On the other hand, corticosteroids induced selective atrophy of type IIb muscle fibers in the intercostal and sternocleidomastoid muscles similar to that in the peripheral skeletal muscles. These findings suggest that the effect of corticosteroids on the diaphragm is unique, and one cannot extrapolate form the effect of corticosteroids on peripheral skeletal muscles to that in the diaphragm. In addition, these pathologic changes may have functional relevance, since atrophy of type I fibers may result in a reduction in muscle endurance.     

Protein degradation in skeletal muscle during experimental hyperthyroidism in rats and the effect of beta-blocking agents

beta-Blocking agents are increasingly used in the management of hyperthyroid patients. The effect of this treatment on increased muscle protein breakdown in the hyperthyroid state is not known. In the present study, experimental hyperthyroidism was induced in rats by daily ip injections of T3 (100 micrograms/100 g BW) during a 10-day period. Control animals received corresponding volumes of solvent. In groups of rats the selective beta-1-blocking agent metoprolol or the nonselective beta-blocker propranolol was infused by miniosmotic pumps implanted sc on the backs of the animals. Protein degradation was measured in incubated intact soleus and extensor digitorum longus muscles by determining tyrosine release into the incubation medium. The protein degradation rate in incubated extensor digitorum longus and soleus muscles was increased by 50-60% during T3 treatment. Metoprolol or propranolol did not influence muscle protein breakdown in either T3-treated or control animals. The results suggest that T3-induced increased muscle proteolysis is not mediated by beta-receptors, and muscle weakness and wasting in hyperthyroidism might not be affected by beta-blockers.     

Human growth hormone treatment of hypophysectomized rats increases the proportion of type-1 fibres in skeletal muscle

The studies describe alterations after hypophysectomy in the proportion of the type-1 and type-2 fibres in rat skeletal muscles, and the effects of replacement treatment with pituitary human (h) GH. Cytochemical analysis of myosin ATPase, succinate dehydrogenase and lactate dehydrogenase activities in sections of rat hind limb muscles were used as markers of fibre type and revealed that hypophysectomy reduced the proportion of type-1 fibres by 50% in soleus and in extensor digitorum longus muscles. This reduction in the proportion of type-1 fibres was accompanied by the appearance of transitional fibres (type 2C/1B). Following seven daily injections of hGH (60 mIU/day) to hypophysectomized rats, the proportion of type-1 fibres in both soleus and in extensor digitorum longus was increased with a concomitant reduction in the number of transitional fibres. After 11 days of treatment, all these transitional fibres had reverted back to type-1 fibres. Only hGH was observed to elicit this effect; injections of other pituitary hormones had no effect on the proportions of these transitional fibres. These alterations in fibre type occurred more rapidly than the changes reported after prolonged electrical stimulation of muscle or following extended exercise. These findings suggest that hypophysectomy and GH injection can result in a rapid alteration in the fibre composition of skeletal muscle, which may have important implications in terms of the resistance to fatigue and speed of contraction of the muscle.     

Effects of hypophysectomy and growth hormone administration on the mRNA levels of collagen I, III and insulin-like growth factor-I in rat skeletal muscle.

The effect of short-term treatment of normal or hypophysectomized rats with biosynthetic growth hormone (GH) was studied in extensor digitorum longus and soleus muscles. In situ hybridization revealed that in normal rats, mRNA for collagen I, collagen III and insulin-like growth factor-I (IGF-I) are expressed by fibroblasts between the muscle fibre areas and that the specificity of this location was not altered by GH administration. Hypophysectomy appeared to cause a decrease in IGF-I and decreased collagen I and III gene expression (P < 0.001, P < 0.001, respectively). GH administration seemed to increase IGF-I mRNA levels in all the animals studied. Quantitative image analysis that GH administration to hypophysectomized rats caused an increase in collagen I gene expression after 2 days (P < 0.05) and an increase in collagen III gene expression after 4 days (P < 0.05). The results indicate that the fibroblast cells are an important target for the action of GH on skeletal muscle and that the fibroblasts respond to GH by increases in the expression of mRNA for collagen I and collagen III.     

Lipid peroxidation and free radical scavengers in thyroid dysfunction in the rat: a possible mechanism of injury to heart and skeletal muscle in hyperthyroidism

This study was designed to determine if peroxidation of biomembrane lipid and the protective system can be modified by the change in oxidative metabolism induced by thyroid dysfunction. The free radical scavengers (i.e. cuprozinc cytosolic and mangano mitochondrial superoxide dismutases, glutathione peroxidase, and catalase), mitochondrial oxidative marker enzymes (cytochrome c oxidase and fumarase), and lipid peroxide were measured in liver, heart, soleus (slow oxidative), and extensor digitorum longus (fast glycolytic) muscles. Rats were rendered hyper- or hypothyroid for 4 weeks and then killed. Superoxide dismutases were detected by specific RIAs: catalase by polarography, and lipid peroxide by fluorimetry. Hypothyroid rats failed to grow, while hyperthyroid rats had hypertrophied hearts but no growth failure. An increase in lipid peroxide was observed in the soleus and heart muscles of hyperthyroid rats. This was accompanied by an increase in mitochondrial superoxide dismutase and oxidative markers. No such change was observed in either fast glycolytic muscle or liver. Glutathione peroxidase decreased in all tissues of hyperthyroid rats, and there was a parallel decrease in catalase in most tissues. On the other hand, hypothyroidism induced a reduction in oxidative markers and mitochondrial superoxide dismutase in heart and skeletal muscles, but only a marginal change in lipid peroxidation. The cytosolic superoxide dismutase did not change in relation to either oxidative metabolism or lipid peroxidation. These results suggest that the enhanced oxidative metabolism and decreased glutathione peroxidase in hyperthyroidism result in an increase in lipid peroxidation and, in slow oxidative and heart muscle, possible organ damage. No adverse reaction mediated by active oxygen species was found in hypothyroid rat tissues.     

Controlled mechanical ventilation leads to remodeling of the rat diaphragm

Little is known about the structural response of the diaphragm to controlled mechanical ventilation. We examined effects of this intervention on muscle mass, myosin heavy chain isoforms, and contractile function in the rat diaphragm. Animals were mechanically ventilated for up to 4 days, and comparisons were made with normal control rats as well as spontaneously breathing animals anesthetized for the same duration as the mechanical ventilation group. The diaphragm-to-body weight ratio was significantly reduced in the mechanical ventilation group only. After mechanical ventilation, an increase in hybrid fibers coexpressing both type I (slow) and type II (fast) myosin isoforms was found within the diaphragm, which occurred at the expense of the pure type I fiber population. In contrast, the percentages of type I, type II, and hybrid fibers in the limb muscles (soleus and extensor digitorum longus) did not differ between experimental groups. The optimal length for force production, as well as maximal force-generating capacity of the diaphragm, was also significantly decreased in mechanically ventilated animals. We conclude that even short-term controlled mechanical ventilation produces significant remodeling and functional alterations of the diaphragm, which could impede efforts at discontinuing ventilatory support.     

Time course of the in vivo effects of thyroid hormone on cardiac gene expression.

The rate of response to thyroid hormone on cardiac growth, heart rate, and the relative changes in messenger RNA (mRNA) coding for alpha- and beta-myosin heavy chain (MHC), slow sarcoplasmic reticulum calcium-adenosine triphosphatase, and thyroid hormone receptors in ventricular tissue of hypothyroid rats was investigated. Hypothyroid rats had significantly smaller hearts, with slower heart rates and expressed no alpha-MHC mRNA as analyzed by an S1 nuclease protection assay when compared to euthyroid animals that expressed 79% alpha-MHC. Twelve hours after treating hypothyroid rats with 20 micrograms of L-T4, detectable levels of alpha-MHC mRNA were present and the shift to alpha-MHC mRNA was complete by 72 h of treatment. Northern blot analysis showed that hypothyroidism resulted in a 60% decrease in the level of sarcoplasmic reticulum calcium-adenosine triphosphatase mRNA which increased after 12 h of T4 administration and was 2.5-fold (P less than 0.05) greater than euthyroid levels after 72 h. In contrast, thyroid hormone receptor mRNA levels measured in poly(A)+ RNA were elevated in hypothyroid rats and decreased to euthyroid levels within 24 h after thyroid hormone treatment. These changes in cardiac gene expression occurred simultaneously with changes in both cardiac size and heart rate. The current studies characterize the coordinated changes and the time course for gene expression that occur in the hypothyroid heart after acute T4 administration.     

Alteration in the expression of GLUT-1 and GLUT-4 protein and messenger RNA levels in denervated rat muscles.

Denervation induces insulin resistance of the glucose transport process in skeletal muscle. To determine whether this is due to alterations in the expression of muscle glucose transporters (GLUT) in different fiber types, we evaluated the amount of GLUT-1 and GLUT-4 protein and messenger RNA (mRNA) in extensor digitorum longus (EDL) and soleus at 1, 2, and 3 days after sciatotomy. Denervation elevated the basal rate of 2-[1,2-3H]deoxy-D-glucose (2-DOG) uptake in the EDL and decreased the insulin-stimulated DOG uptake in both muscles. Denervation after 1 day did not modify the GLUT-1 or the GLUT-4 protein level in either muscle. However, it increased GLUT-1 mRNA by 66% and decreased GLUT-4 mRNA by 70% in the EDL, but not in the soleus (P < 0.05). After 2 days of denervation, by which time GLUT-1 mRNA was increased 2-fold and GLUT-4 mRNA was reduced by 70%, we observed a 2-fold increase in GLUT-1 protein (P < 0.01) in the EDL and a 40-45% decrease in GLUT-4 protein in both muscles (P < 0.01). These results indicate that modifications in the expression of GLUT-1 and GLUT-4 protein cannot explain the insulin resistance of the glucose transport process in the EDL or soleus 1 day after denervation. After 2 days of denervation, however, alterations in GLUT-1 and GLUT-4 protein levels may contribute to the change in basal and insulin-stimulated DOG uptake in both the EDL and the soleus muscles.     

Role of adenosine 5'-monophosphate-activated protein kinase in interleukin-6 release from isolated mouse skeletal muscle

IL-6 is released from skeletal muscle during exercise and has consequently been implicated to mediate beneficial effects on whole-body metabolism. Using 5-aminoimidazole-4-carboxamide-1-beta-4-ribofuranoside (AICAR), a pharmacological activator of 5'-AMP-activated protein kinase (AMPK), we tested the hypothesis that AMPK modulates IL-6 release from isolated muscle. Skeletal muscle from AMPKalpha2 kinase-dead transgenic, AMPKalpha1 knockout (KO) and AMPKgamma3 KO mice and respective wild-type littermates was incubated in vitro, in the absence or presence of 2 mmol/liter AICAR. Skeletal muscle from wild-type mice was also incubated with the AMPK activator A-769662. Incubation of mouse glycolytic extensor digitorum longus and oxidative soleus muscle for 2 h was associated with profound IL-6 mRNA production and protein release, which was suppressed by AICAR (P < 0.001). Basal IL-6 release from soleus was increased between AMPKalpha2 kinase-dead and AMPKalpha1 KO and their respective wild-type littermates (P < 0.05), suggesting AMPK participates in the regulation of IL-6 release from oxidative muscle. The effect of AICAR on muscle IL-6 release was similar between AMPKalpha2 KD, AMPKalpha1 KO, and AMPKgamma3 KO mice and their respective wild-type littermates (P < 0.001), indicating AICAR-mediated suppression of IL-6 mRNA expression and protein release is independent of AMPK function. However, IL-6 release from soleus, but not extensor digitorum longus, was reduced 45% by A-769662. Our results on basal and A-769662-mediated IL-6 release provide evidence for a role of AMPK in the regulation of IL-6 release from oxidative skeletal muscle. Furthermore, in addition to activating AMPK, AICAR suppresses IL-6 release by an unknown, AMPK-independent mechanism.     

Induction of insulin-like growth factor I messenger ribonucleic acid during regeneration of rat skeletal muscle

Expression of insulin-like growth factor I (IGF-I) was studied in regenerating skeletal muscle. Irreversible damage to muscle cells was induced in the extensor digitorum longus muscle of adult rats by ischemia, preceded by glycogen depletion. IGF-I mRNA levels during the regeneration process were studied for periods up to 10 days after injury using a solution hybridization assay. Increased IGF-I mRNA levels could be demonstrated within 24 h after injury; maximum levels were achieved in 3 days and decreased to approximately normal levels by 10 days. Changes in IGF-I mRNA levels could not be seen in undamaged contralateral extensor digitorum longus muscles during the experimental period. An increase in IGF-I mRNA was also evident in injured muscles of hypophysectomized animals. In situ hybridization at the time of maximum induction showed the presence of IGF-I mRNA in proliferating myoblasts and in satellite cells. IGF-I, thus, may act as a locally produced non-GH dependent trophic factor during regeneration of skeletal muscle after injury.     

Skeletal muscle fiber types and myosin ATPase activity do not change with age or growth hormone administration

Fischer 344 rats at various ages throughout the life span have been treated with growth hormone, clonidine, and insulin-like growth factor-I to restore circulating somatomedin levels in old animals to levels found in younger rats. The injections were intended to reverse (2-week treatments) or prevent (6-month treatments) deleterious effects of aging on skeletal muscle--specifically the loss of fast fibers during the latter part of the life span. However, measurements of myosin ATPase (and subsequent histochemical fiber type determinations) revealed that the previously reported age-related decrease in ATPase activities and fast fiber content did not occur in barrier-protected specific pathogen free rats or mice. None of the treatments used had a significant effect on the ATPase activity or fiber composition of soleus, extensor digitorum longus, or diaphragm muscles, although parallel determinations in collaborating laboratories verified that the hormones had major effects in other systems. Previously reported decreases with age in ATPase activity of heart muscle were confirmed in our experiments, and these decreases were shown to reflect a change in myosin isozyme composition of the hearts, both at the protein and the mRNA levels of gene expression.     

Angiotensin converting enzyme inhibition prevents development of muscle and nerve dysfunction and stimulates angiogenesis in streptozotocin-diabetic rats

Summary The effects of the angiotensin converting enzyme inhibitor lisinopril on slow and fast twitch muscle contractile properties, nerve conduction and hypoxic resistance, and muscle and nerve capillary density were examined in streptozotocin-diabetic rats. Prolongation of soleus contraction and relaxation were partially prevented by treatment (p<0.01). A 22% deficit in fast twitch extensor digitorum longus tetanic tension production was also ameliorated (p<0.01). Sciatic motor and sensory conduction velocity, 25% and 12% reduced by diabetes respectively, were 75% normalized by lisinopril (p<0.01). There was a 47% increase in resistance to hypoxic conduction block with diabetes (p<0.01). Lisinopril treatment resulted in normal hypoxic resistance. Capillarisation of nerve and muscle was little affected by diabetes; however, there was a 17% increase in capillary density in sciatic nerve, and a 40% increase in extensor digitorum longus muscle with lisinopril (p<0.01). For soleus, a smaller treatment-induced increase in capillary density led to an elevated capillary/muscle fibre ratio (p<0.01). These results suggest that lisinopril promoted angiogenesis. It was concluded that the beneficial effect of preventive lisinopril treatment is likely to depend upon a reduction of peripheral vascular resistance and improvement of tissue blood flow, which implicates relative hypoxia as an important factor in the development of myopathy and neuropathy in experimental diabetes.     

Connexin40 messenger ribonucleic acid is positively regulated by thyroid hormone (TH) acting in cardiac atria via the TH receptor

Thyroid hormone (TH) regulates many cardiac genes via nuclear thyroid receptors, and hyperthyroidism is frequently associated with atrial fibrillation. Electrical activity propagation in myocardium depends on the transfer of current at gap junctions, and connexins (Cxs) 40 and 43 are the predominant junction proteins. In mice, Cx40, the main Cx involved in atrial conduction, is restricted to the atria and fibers of the conduction system, which also express Cx43. We studied cardiac expression of Cx40 and Cx43 in conjunction with electrocardiogram studies in mice overexpressing the dominant negative mutant thyroid hormone receptor-beta Delta337T exclusively in cardiomyocytes [myosin heavy chain (MHC-mutant)]. These mice develop the cardiac hypothyroid phenotype in the presence of normal serum TH. Expression was also examined in wild-type mice rendered hypothyroid or hyperthyroid by pharmacological treatment. Atrial Cx40 mRNA and protein levels were decreased (85 and 55%, respectively; P < 0.001) in MHC-mt mice. Atrial and ventricular Cx43 mRNA levels were not significantly changed. Hypothyroid and hyperthyroid animals showed a 25% decrease and 40% increase, respectively, in Cx40 mRNA abundance. However, MHC-mt mice presented very low Cx40 mRNA expression regardless of whether they were made hypothyroid or hyperthyroid. Atrial depolarization velocity, as represented by P wave duration in electrocardiograms of unanesthetized mice, was extremely reduced in MHC-mt mice, and to a lesser extent also in hypothyroid mice (90 and 30% increase in P wave duration). In contrast, this measure was increased in hyperthyroid mice (19% decrease in P wave duration). Therefore, this study reveals for the first time that Cx40 mRNA is up-regulated by TH acting in cardiac atria via the TH receptor and that this may be one of the mechanisms contributing to atrial conduction alterations in thyroid dysfunctions.     

Differential effects of leptin in regulation of tissue glucose utilization in vivo

We have recently shown that leptin enhances systemic insulin sensitivity and whole body glucose utilization in the rat. This study examines our hypothesis that leptin has differential effects in regulating glucose utilization among the tissues, i.e. stimulating glucose utilization in brown adipose tissue (BAT) and skeletal muscle but suppressing glucose utilization in white adipose tissue (WAT) in normal male rats (275-350 g BW). The rats were treated with s.c. infusion of recombinant murine leptin (4 mg/kg x day) or vehicle (V) with Alzet osmotic pumps or with vehicle and pair-feeding (PF) for 7 days. Leptin significantly decreased food intake (leptin, 11.5 +/- 0.4 g/day; V, 16.8 +/- 1.5 g/day; P < 0.05) and body weight (maximum change, 5.0 +/- 0.2%; P < 0.05 vs. V) and lowered plasma triglyceride, insulin, and glucose levels, but raised beta-hydroxybutyrate levels. Glucose utilization by individual tissues was determined with an i.v. bolus of [1-(14)C]2-deoxyglucose (2-DG) after a 90-min hyperinsulinemic (2 mU/kg x min) euglycemic clamp. With leptin treatment, the 2-DG-determined glucose utilization in interscapular BAT was almost 3-fold that in V-treated rats and 70% greater than that in PF rats. In contrast, in the epididymal WAT, glucose utilization was reduced by leptin treatment to only 34% that in V-treated rats and 45% that in PF rats. Leptin increased 2-DG uptake by extensor digitorum longus muscle and soleus muscle compared with that in the V and PF groups. With leptin treatment, the GLUT4 glucose transporter mRNA and protein levels were increased in BAT, but decreased in WAT (both P < 0.05). There was no significant change in GLUT4 mRNA and protein expression in extensor digitorum longus muscle and soleus muscle. Oxygen consumption was significantly increased (32.1 +/- 7.4%) in BAT (139.0 +/- 8.2 nmole O2/30 min x 10(6) cells) of leptin-treated rats vs. that in V control rats (105.3 +/- 6.7 nmole O2/30 min x 10(6) cells). In conclusion, leptin has differential, tissue-specific effects on glucose and oxygen utilization, which contribute to the reduction in whole body adiposity by enhancing energy consumption in BAT and muscle while attenuating energy storage in WAT.     

Metabolism of branched-chain amino acids in leg muscles from tail-cast suspended intact and adrenalectomized rats

Degradation of branched-chain amino acids was studied in muscles of unloaded hind limbs from rats subjected to six days of tail-cast suspension. The total production of 14CO2 from uniformly labeled 14C-leucine, isoleucine, or valine, and the fluxes through leucine aminotransferase and alpha-ketoisocaproate dehydrogenase, which were measured using L-1-14C-leucine, were generally greater in the soleus and extensor digitorum longus muscles of unloaded than of weight-bearing hind limbs. Adrenalectomy abolished any difference in flux through the aminotransferase, whereas the administration of cortisol to adrenalectomized animals restored the greater flux in the unloaded soleus muscle. Adrenalectomy partially diminished the greater flux through alpha-ketoisocaproate dehydrogenase in the unloaded soleus, whereas cortisol (2 mg/100 g body weight) treatment increased this difference. In the extensor digitorum longus, adrenalectomy abolished the differences in both enzyme fluxes due to hind limb suspension. In this muscle, cortisol treatment increased these fluxes to a similar extent in both weight-bearing and suspended, adrenalectomized animals so that the normal difference was not restored. These results suggest that leucine catabolism in hind limb muscles of suspended rats was influenced primarily by increased circulating glucocorticoid hormones, which are elevated twofold to fourfold in these animals.