Interorgan signaling between adipose tissue metabolism and skeletal muscle uncoupling protein homologs: is there a role for circulating free fatty acids?

Uncoupling proteins 3 and 2 (UCP3 and UCP2) are two newly cloned genes that have been implicated in the regulation of lipids as fuel substrate in skeletal muscle on the basis that their mRNA expressions are upregulated during starvation (when fat stores are being rapidly mobilized) and downregulated during the early phase of refeeding (when fat stores are being rapidly replenished). To test the hypothesis that circulating free fatty acids (FFAs) may have a physiological role as an interorgan signal linking these dynamic changes in the fat stores to skeletal muscle expression of UCP3 and UCP2, the mRNA levels of these UCP homologs were examined in fed and fasted rats treated with the antilipolytic agent nicotinic acid. In 46-h fasted rats, we observed a threefold increase in serum FFA levels and increases in UCP3 and UCP2 mRNA levels that were more marked in the gastrocnemius and tibialis anterior muscles (predominantly fast-twitch fibers) than in the soleus muscle (predominantly slow-twitch fibers). Treatment with nicotinic acid blunted the fasting-induced increase in serum FFA levels and prevented the increase in mRNA levels of UCP3 and UCP2 in the soleus muscle, but had little or no effect on the elevated mRNA levels of these UCP homologs in the gastrocnemius and tibialis anterior muscles. Furthermore, treatment of ad libitum-fed animals with nicotinic acid resulted in a twofold reduction in serum FFA levels (i.e., by a magnitude similar to that observed during early refeeding) and significant reductions in UCP3 and UCP2 mRNA levels in the soleus muscle, but not in the gastrocnemius or tibialis anterior muscles. These results revealed a muscle-type dependency in the way UCP2 and UCP3 gene expression in skeletal muscle is regulated, and suggest that the hypothesis that circulating FFAs function as an interorgan signal between fat stores and skeletal muscle UCP3 and UCP2 gene expression is adequate only for slow-twitch (oxidative) muscles. Consequently, a signal(s) other than circulating FFAs must be implicated in the link between dynamic changes in body fat stores and UCP expression in predominantly fast-twitch (glycolytic/oxidative-glycolytic) muscles, which constitute the major fiber type of the total skeletal muscle mass and which have high susceptibility to developing insulin resistance and impairment in substrate utilization in metabolic diseases.     

The preferential mobilisation of C20 and C22 polyunsaturated fatty acids from the adipose tissue of the chick embryo: potential implications regarding the provision of essential fatty acids for neural development

The effects of 10 day clenbuterol administration on cardiac and skeletal muscle capillarities were studied, particularly in terms of the distribution of arteriolar and venular capillaries and their capillary density, in young (10-week-old) and middle-aged (37-week-old) male Wistar rats. Rats of the treated groups were fed a diet containing 2 mg kg-1 clenbuterol hydrochloride. In both young and middle aged rats, clenbuterol treatment increased the body wt and the weights of the heart and hindlimb muscles. The mean fibre cross-sectional area was significantly increased after the treatment in the left ventricle, soleus, plantaris and both deep and superficial portions of gastrocnemius (P < 0.01). In the left ventricle, the total capillary density and the density of venular capillaries were decreased after the treatment in both young (9 and 13%, respectively) and middle-aged rats (10 and 11%, respectively). A decrease in total capillary density was also observed in all skeletal muscles examined. In both young and middle-aged rats, the capillary-to-fibre (C:F) ratio and the proportion of each capillary did not change after the treatment in both the left ventricle and skeletal muscles. Clenbuterol significantly decreased the activity of succinate dehydrogenase in all skeletal muscles examined (P < 0.01). These results suggest that clenbuterol increased the diffusion distance for oxygen in the left ventricle and skeletal muscles. These changes may reduce the oxygen supply to tissues and increase muscle fatigability.     

Changes in transcriptional activity of chronically stimulated fast twitch muscle

mRNAs extracted from rabbit soleus, normal and 28-day, indirectly stimulated tibialis anterior muscles were translated in an in vitro system. Analysis for translation products by 2-dimensional electrophoresis showed fast myosin light chains in tibialis anterior, and slow myosin light chains in soleus muscle. The stoichiometry of the in vitro translated light chains varies from that seen in normal fast and slow twitch muscles. The stimulated muscle contained mRNA coding, both for fast and slow myosin light chains, although the pattern of slow myosin light chains appears not to be complete at this point of time of the transformation process.     

Acute effects of the beta 3-adrenoceptor agonist, BRL 35135, on tissue glucose utilisation

1. The acute effects of BRL 35135 (BRL) on tissue glucose utilisation index (GUI) in vivo were investigated in anaesthetized rats by use of 2-deoxy-[3H]-glucose. 2. Intravenous injection of BRL caused a dose-dependent increase in GUI in skeletal muscle, and white and brown adipose tissue; plasma insulin and fatty acid concentrations were also increased. Chronic treatment with BRL added to the diet caused a 34 fold increase in basal GUI of brown adipose tissue (BAT), but had no effect on GUI in other tissues. After chronic treatment, the acute tissue response to an intravenous maximal dose of BRL had disappeared completely in all tissues apart from the soleus muscle. 3. A high dose (20 mg kg-1) of the non-selective beta-antagonist, propranolol, inhibited the acute effect of BRL on GUI in BAT, but failed to affect GUI in muscle. A lower dose (1 mg kg-1) of the antagonist also inhibited the BAT response, but had little or no effect on the response in Type I (working) muscles such as soleus and adductor longus (ADL), and potentiated the response in Type II (non-working) muscles such as tibialis and extensor digitorium longus (EDL). 4. A low dose (1 mg kg-1) of the selective beta 1-antagonist, atenolol, had no effect on the BRL response but the same dose of the selective beta 2-antagonist, ICI 118551, potentiated significantly the effect of BRL on GUI in most muscles without altering plasma insulin levels. 5. It is concluded that: (i) the heterogeneous tissue responses of different muscle fibre types in the presence of P-antagonists indicates that BRL affects muscle GUI directly, in addition to effects mediated by increases in plasma insulin concentration; (ii) the resistance of the BRL response to conventional P-adrenoceptor antagonists implicates an atypical adrenoceptor mediating the GUI response in skeletal muscle, but this may not be identical to the adipose tissue P3-adrenoceptor; (iii) the potentiation of BRL responses by ICI 118551 indicates an inhibitory P2-adrenoceptor-mediated component in the muscle GUI response to BRL.     

Effects of surface electrostimulation on the structure and metabolic properties in monkey skeletal muscle

Adaptative changes in skeletal muscle following surface electrical stimulation (SES) were investigated in rhesus monkeys. SES was performed on the triceps brachialis muscle (TB) according to an intermittent pattern. The procedure was carried out for 3 wk, using a current with a medium frequency of 60 Hz normally observed in fast motor axons. The histochemical assays performed on biopsies taken from proximal and distal parts of the TB muscle, before and after the SES program, showed that the distribution of fibers typed by ATPase was unaffected. On the other hand, SES led to an overall increase in the mean fiber cross-sectional area (FCSA); P < 0.01 (+13.7%, NS, in proximal portion, vs +31%, P < 0.01 in distal portion). This increase in size occurred in all fiber types. SES was shown to induce an overall increase in capillary to fiber ratio (C/F; +11.06%, NS, in proximal portion, vs +25.93%, P < 0.05 in distal portion). The number of capillaries surrounding fiber Type II (CAFII) was significantly increased by SES (P < 0.05): +3.21%, NS, in proximal portion, versus +21.47%, P < 0.05 in distal portion. Moreover, the number of capillaries surrounding fiber Type I (CAFI) was statistically unaffected by SES. These results suggest that a stimulation of capillary growth may occur following SES-training. Citrate synthase activity was significantly increased after SES. This enhancement in oxidative potential was shown to occur in all fiber types (NADH-diaphorase staining).(ABSTRACT TRUNCATED AT 250 WORDS)     

Contractile studies of single human skeletal muscle fibers: a comparison of different muscles, permeabilization procedures, and storage techniques

The study of single muscle fibers has improved our understanding of muscle physiology and pathology. To compare three techniques for fiber preparation and storage, biopsies were obtained from the tibialis anterior and vastus lateralis muscles of a hemiparetic patient and a control subject. Single fibers were prepared with: (1) chemical skinning (CS) and storage at -20 degrees C; (2) chemical skinning followed by sucrose (SU) incubation and storage at -80 degrees C; or (3) freeze-drying (FD) and -80 degrees C storage. Cross-sectional area (CSA), resting, maximal (P0), and specific tension (P0/CSA), and maximum shortening velocity (V0) were determined in 189 cells. CSA was similar in all groups. Resting tension was higher and P0 and P0/CSA lower after FD. In general, V0 was the same in all groups. Our data suggest that CS and SU preserve the properties of single muscle fibers better than FD. SU may allow longer storage of fibers.     

A novel approach for comparative study of periosteum, muscle, subcutis, and skin microcirculation by intravital fluorescence microscopy

Herein, we report a new model, which allows comparative study of the microcirculation of different peripheral tissues, i.e., periosteum, skeletal muscle, subcutis, and skin. Using dextran-insensitive Wistar rats gracilis and semitendinosus muscles of the left hindlimb were prepared in association with their appertaining tibial fragments, subcutis, and skin. Blood supply was guaranteed by the femoral artery via the saphenous vessels. High-resolution intravital epi-illumination microscopy of the two muscles displayed the typical microvascular architecture with the capillaries running in parallel to each other (capillary density (CD) 128.4 +/- 4.5 cm-1). In subcutis and skin, capillaries were found arranged as interconnecting mesh-like networks with a density, which was significantly higher (P < 0.05) in subcutis (191.0 +/- 5.5 cm-1) compared with skin (108.9 +/- 3.3 cm-1). Analysis of periosteal tissue revealed two distinct types of arrangements of microvascular architecture. Adjacent to the major feeding and draining vessels of the periosteum, capillaries were organized in densely meshed shunt-like networks, revealing the highest capillary density (242.7 +/- 13.2 cm-1; P < 0.05) of all tissues studied. Periosteal capillaries distant from the major feeding and draining vessels were arranged in parallel to the longitudinal axis of the tibial bone and presented with a density similar to that of the skeletal muscle (128. 6 +/- 9.4 cm-1). Topical application of acetylcholine for analysis of physiological reactivity of the microvasculature showed dose-dependent arteriolar dilation. Moreover, a 3-min upstream femoral artery occlusion demonstrated an appropriate hyperemic response in all tissues studied, indicating intact myogenic control. A prolonged period of ischemia (120 min) followed by reperfusion (60 min) caused massive (P < 0.05) leukocyte-endothelial cell interaction in postcapillary venules, similarly as reported in other microvascular tissue preparations. We propose that the model presented provides a good approach to all peripheral tissues for both the analysis of the physiology of tissue-confined microvascular control and the development of novel therapeutic strategies to counteract manifestation of nutritional dysfunction and inflammatory response in disease.     

Remarks on the structure and course of renal efferent glomerular arterioles

Patients with chronic heart failure have structural and metabolic changes in skeletal muscle, which may be of importance for symptomatology. The origin of these changes are still unknown. The relationship between fiber composition and capillarization in skeletal muscle with exercise capacity and central hemodynamic variables was examined. Biopsies from the lateral vastus muscle were taken in 12 patients with chronic heart failure. Samples from eight normal subjects served as control samples. All patients underwent maximal exercise tests. Central hemodynamic variables were measured during exercise in five patients. The patients had a higher percentage of type II B fibers (P = .03) and fewer capillaries per fiber (P = .02) than the controls subjects. VO2 max correlated with the percentage of type I fibers, whereas the correlation with the type II A fibers was inverse. Cardiac index and pulmonary capillary wedge pressure at submaximal and maximal exercise were related to fiber type composition and relative fiber areas. Skeletal muscle fiber type composition and capillarization was changed in patients with chronic heart failure. These changes might influence exercise capacity. There were relationships between central hemodynamic variables and skeletal muscle changes. What the cause and effects were need further investigation.     

Effect of heart failure on muscle capillary geometry: implications for 02 exchange

There is strong evidence that chronic heart chronic heart failure (CHF) impairs skeletal muscle function independent of blood flow and bulk O2 delivery. Purpose: This investigation sought to determine whether alterations in muscle capillary geometry and surface area that are thought to be primary determinants of the efficacy for blood-tissue 02 exchange might be altered in CHF and contribute to these changes. Methods: Plantaris (fast twitch) and soleus (slow twitch) muscles from control (C) and 6- to 7-wk post myocardial infarcted (CHF) rates were perfusion-fixed in situ. These muscles were analyzed using morphometric techniques that facilitated determination of muscle sarcomere length fiber cross-sectional area, capillary tortuosity and branching coefficient (c(K,0)), capillary length, volume, and surface area. Results: Normalized to a sarcomere length of 2.1 microns, plantaris fiber cross-sectional area decreased by 21% (P < 0.05), and capillary-to-fiber ratio decreased from 2.05 +2- 0.07 in C to 1.79 +2- 0.04 (P < 0.05) in CHF, but these variables were unchanged in soleus. These was no change in c(K,0) or capillary diameter in either muscle, and thus capillary length and surface area per fiber volume remained unchanged. From the measured fiber atrophy and capillary involution in plantaris reductions of total muscle capillary length, volume, and surface area of 11%, 9% and 17%, respectively, are estimated. Conclusion: These changes, coupled with reduced blood flow may impair the effective matching of muscle fiber 02 delivery to 02 requirement during repeated muscle contractions (i.e. exercise). The scenario is expected to reduce intramyocyte 02 partial pressure and thereby contribute to the greater fatigability characteristic of the CHF condition.     

Electrophysiologic evaluation of denervated muscles in incomplete paraplegia using macro electromyography

OBJECTIVE: To evaluate denervated muscles in persons with incomplete paraplegia due to thoracolumbar spinal injury (TLSI) using macro electromyography in determining indications for functional electrical stimulation (FES). DESIGN: A randomized clinical trial and a criterion standard. SETTING: A department of orthopedic surgery in a university hospital. PATIENTS AND OTHER PARTICIPANTS: Eighteen patients with incomplete paraplegia, including 11 with TSLI, and 50 healthy adults. INTERVENTION: Area and amplitude of macro motor unit potential (macro MUP) were measured at the tibialis anterior, the vastus lateralis, and the vastus medialis. The normal limits of macro MUP parameters were defined based on values from healthy subjects. Abnormal denervated muscles were detected by macro EMG and conventional EMG in paralytic patients. The correlation between macro MUP parameter values and muscle forces of the tibialis anterior and quadriceps femoris induced by electrical stimulation was analyzed. MAIN OUTCOME MEASURES: The number of abnormal muscles, parameter values, and muscle force induced by electrical stimulation. RESULTS: Abnormal muscles were found only in the TLSI patients and 13 abnormal muscles were detected by macro EMG only. The abnormal muscles defined by macro EMG showed insufficient contraction induced by electrical stimulation. The increase of parameter value negatively correlated with the muscle force (tibialis anterior area r=-.797, amplitude r=-.866; quadriceps area r=-.866, amplitude r=-.893; p < .001). CONCLUSIONS: These results suggest that macro EMG is useful in detecting denervated muscles, in determining indications for FES, and in predicting FES effects before implantation of electrodes.     

Muscle fiber length and moment arm coordination during dorsi- and plantarflexion in the mouse hindlimb

The purpose of this study was to test the hypothesis that muscle fiber length and joint moment arm are combined in such a way that maximum muscle force is produced during locomotion. Plantarflexor (soleus, SOL and medial gastrocnemius, MG) and dorsiflexor (extensor digitorum longus, EDL and tibialis anterior, TA) muscle architecture in the mouse was measured along with their associated moment arms. Fiber length varied significantly between muscles ranging from 5.7 +/- 0.2 mm (MG) to 7.6 +/- 0.2 mm (TA). Plantarflexor moment arms were over twice as large as dorsiflexor moment arms (1.88 +/- 0.06 mm vs. 0.84 +/- 0.03 mm) suggesting a greater muscle length change with joint angle for plantarflexors compared to dorsiflexors. Using a simple muscle-joint model, the active sarcomere length range in these muscle groups was calculated and proved to be quite similar between functional groups. The active range for dorsiflexors was 2.2-2.4 microns, while the active range for plantarflexors was 2.2-2.5 microns, indicating that both muscle groups operate primarily near the plateau of their length tension-relation. Finally, when calculating force produced by muscles during locomotion, the combination of moment arm and fiber length measured in all muscle groups yielded muscle-joint systems that produced near maximal forces at the velocities modeled. These data indicate that fiber length and moment arm appear to be coordinated to yield the greatest possible force production during locomotion.     

Effect of immobilization on skeletal muscle nicotinic cholinergic receptors in the rat

The effect of 4 weeks of hind limb immobilization on nicotinic acetylcholinergic receptors (nAChRs) in the neuromuscular junction of the soleus (SOL) and tibialis anterior (TIB) muscles was studied in rats. Quantitative measurements of the receptors was performed using [3H]alpha-bungarotoxin ([3H]alpha-BTx) receptor autoradiography. Junctional and extrajunctional nAChRs were significantly increased in the SOL and TIB after 4 weeks immobilization. However, a significant decrease in fiber cross-sectional area was observed only in the SOL muscle. Remobilization for 4 weeks reversed the changes in cholinergic receptors and muscle fibers but not in bone. Our findings suggested that lack of nerve impulses are of importance for the events that take place after immobilization leading to muscle atrophy and osteoporosis.     

Molecular forms of acetylcholinesterase in dystrophic (mdx) mouse tissues

We analyzed the activity of acetylcholinesterase (AChE) and its molecular forms in the tissues of normal and dystrophic (mdx) mice, at different developmental stages. We studied the brain, the heart and the serum, in addition to four predominantly fast-twitch muscles (tibialis, plantaris, gastrocnemius and extensor digitorum longus (EDL)) and the slow-twitch, soleus muscle. We found no difference between mdx and control mice in the AChE activity of the brain and the heart. The skeletal muscles affected by the disease undergo active degeneration counterbalanced by regeneration between 3 and 14 weeks after birth. The distribution of AChE patches associated with neuromuscular junctions was abnormally scattered in mdx muscles, and in some cases (tibialis and soleus), the number of endplates was more than twice that of normal muscles. There were only minor differences in the concentration and pattern of AChE molecular forms during the acute phase of muscle degeneration and regeneration. After this period, however, we observed a marked deficit in the membrane-bound G4 molecular form of AChE in adult mdx tibialis, gastrocnemius and EDL but not in the plantaris or in the soleus, as compared with their normal counterparts. Whereas the amount of AChE markedly decreased in the serum of normal mice during the first weeks of life, it remained essentially unchanged in the serum of mdx mice. It is likely that this excess of AChE activity in serum originates from the muscles. A deficit in muscle G4 was also reported in other forms of muscular dystrophy in the mouse and chicken. Since it is not correlated to the acute phase of the disease in mdx and also occurs in genetically different dystrophies, it probably represents a secondary effect of the dystrophy.     

Fiber architecture of the extensors of the hindlimb in semiterrestrial and arboreal guenons

Fiber architecture of the extensor musculature of the knee and ankle is examined in two African gueon species--the semiterrestrial Cercopithecus aethiops, and the arboreal C. ascanius. Using histologic and microscopic techniques to measure lengths of sarcomeres, the original lengths of muscle fasciculi and angles of pinnation in quadriceps femoris and triceps surae are reconstructed from direct measurements on cadavers. Calculations of reduced physiological cross-sectional area, mass/predicted effective tetanic tension, maximum excursion, and tendon length/fasciculus+tendon lengths are correlated to preferred locomotor modalities in the wild. For both species, greater morphological differences occur among the bellies of quadriceps femoris--rectus femoris, vastus intermedius, v. lateralis, and v. medialis--than among the bellies of triceps surae--gastrocnemius lateralis, g. medialis, plantaris, and soleus. With regard to quadriceps femoris, few differences occur between species. Interspecific differences in the triceps surae indicate (1) redirection of muscle force to accommodate arboreality in which the substrate is less than body width; (2) muscles more suited for velocity in the semiterrestrial vervets; and (3) muscles used more isotonically in vervets and more isometrically in red-tailed monkeys. The inherent flexibility of muscles may be preadaptive to a primary species shift in locomotor modality until the bony morphology is able to adapt through natural selection.     

Experimental muscle pain increases the human stretch reflex

In this study we investigated the effect of human experimental muscle pain on H- and stretch reflexes as indicators of changes in muscle spindle sensitivity. Fourteen healthy, male volunteers participated in the study. Muscle pain was produced by infusion of 5% hypertonic saline over a period of 10-15 min in m. soleus and in m. tibialis anterior. Reflexes were elicited in the relaxed and active soleus muscle (10-15 Nm ankle torque) before, during and after muscle pain. Control measurements were made with infusions of 0.9% isotonic saline. Surface electromyograms (EMG) were measured from the soleus muscle, and torque was measured from the ankle joint. With pain in the soleus muscle the mechanical stretch reflex response (ankle torque) increased significantly (P = 0.0007) as compared to before pain. With pain in the tibialis anterior muscle both the mechanical and EMG responses increased significantly (P = 0.001; P = 0.0003) as compared to before pain. The H-reflex showed no significant changes during the infusions in either muscles. This study has demonstrated a muscle pain-related increase in the amplitude of the stretch reflex without a corresponding increase in the H-reflex amplitude. One explanation could be an increased dynamic sensitivity of the muscle spindles during muscle pain caused by an increased firing rate in the dynamic gamma-motoneurones. However, the data could not support the vicious cycle model because the excitability of the alpha-motoneurone pool was unchanged.     

Modulation of H reflexes in human tibialis anterior muscle with passive movement

Significant movement-induced gain changes in H reflexes have been observed in soleus muscle following passive movement of the lower limb. Hypotheses from these concepts were tested on magnitudes of H reflexes in tonically contracted tibialis anterior. From eleven subjects at rates of 20 and 60 r.p.m. passive leg movement, statistically significant attenuation from controls and phasic modulation occurred. The results make more general the conclusions from soleus H reflexes. However, the functional effect should be much smaller, as tibialis anterior H reflexes are smaller compared to those in soleus.     

The effect of ischemic preconditioning on the recovery of skeletal muscle following tourniquet ischemia

It has been well documented that ischemic preconditioning limits ischemic-reperfusion injury in cardiac muscle, but the ability of ischemic preconditioning to limit skeletal muscle injury is less clear. Previous reports have emphasized the beneficial effects of ischemic preconditioning on skeletal muscle structure and capillary perfusion but have not evaluated muscle function. We investigated the morphologic and functional consequences of ischemic preconditioning, followed by a 2-hour period of tourniquet ischemia on muscles in the rat hindlimb. The 2-hour ischemia was imposed without preconditioning, or was preceded by three brief (10 minutes on/10 minutes off) preischemic conditioning intervals. We compared muscle morphology, isometric contractile function, and muscle fatigue properties in predominantly fast-twitch, tibialis anterior muscles 3 (n = 8) and 7 (n = 8) days after ischemia-reperfusion. Two hours of ischemia, followed by reperfusion, results in a 20 percent reduction of muscle mass (p < 0.05) and a 33 percent reduction in tetanic tension (p < 0.05) when compared with controls (n = 8) at 3 days. The same protocol, when preceded by ischemic preconditioning, results in similar decreases in muscle mass and contractile function. Neuromuscular transmission was also impaired in both ischemic groups 7 days after ischemia. Nerve-evoked maximum tetanic tension was 69 percent of the tension produced by direct muscle stimulation in the ischemia group and 65 percent of direct tension in the ischemic preconditioning/ischemia group. In summary, ischemic preconditioning, using the same protocol reported to be effective in limiting infarct size in porcine muscle, had no significant benefit in limiting injury or improving recovery in the ischemic rat tibialis anterior. The value of ischemic preconditioning in reducing imposed ischemic-reperfusion-induced functional deficits in skeletal muscle remains to be demonstrated.