No change in myonuclear number during muscle unloading and reloading.

Muscle fibers are the cells in the body with the largest volume, and they have multiple nuclei serving different domains of cytoplasm. A large body of previous literature has suggested that atrophy induced by hindlimb suspension leads to a loss of "excessive" myonuclei by apoptosis. We demonstrate here that atrophy induced by hindlimb suspension does not lead to loss of myonuclei despite a strong increase in apoptotic activity of other types of nuclei within the muscle tissue. Thus hindlimb suspension turns out to be similar to other atrophy models such as denervation, nerve impulse block, and antagonist ablation. We discuss how the different outcome of various studies can be attributed to difficulties in separating myonuclei from other nuclei, and to systematic differences in passive properties between normal and unloaded muscles. During reload, after hindlimb suspension, a radial regrowth is observed, which has been believed to be accompanied by recruitment of new myonuclei from satellite cells. The lack of nuclear loss during unloading, however, puts these findings into question. We observed that reload led to an increase in cross sectional area of 59%, and fiber size was completely restored to the presuspension levels. Despite this notable growth there was no increase in the number of myonuclei. Thus radial regrowth seems to differ from de novo hypertrophy in that nuclei are only added during the latter. We speculate that the number of myonuclei might reflect the largest size the muscle fibers have had in its previous history.

Mechanotransduction in striated muscle via focal adhesion kinase.

Contractile tissues demonstrate a pronounced capacity to remodel their composition in response to mechanical challenges. Descriptive evidence suggests the upstream involvement of the phosphotransfer enzyme FAK (focal adhesion kinase) in the molecular control of load-dependent muscle plasticity. Thereby FAK evolves as a myocellular transducer of mechanical signals towards downstream transcript expression in myofibres. Recent advances in somatic gene therapy now allow the exploration of the functional involvement of this enzyme in mechanotransduction in intact muscle.

Double gene deletion reveals the lack of cooperation between PPARalpha and PPARbeta in skeletal muscle.

The peroxisome proliferator-activated receptors (PPARs) are involved in the regulation of most of the pathways linked to lipid metabolism. PPARalpha and PPARbeta isotypes are known to regulate muscle fatty acid oxidation and a reciprocal compensation of their function has been proposed. Herein, we investigated muscle contractile and metabolic phenotypes in PPARalpha-/-, PPARbeta-/-, and double PPARalpha-/- beta-/- mice. Heart and soleus muscle analyses show that the deletion of PPARalpha induces a decrease of the HAD activity (beta-oxidation) while soleus contractile phenotype remains unchanged. A PPARbeta deletion alone has no effect. However, these mild phenotypes are not due to a reciprocal compensation of PPARbeta and PPARalpha functions since double gene deletion PPARalpha-PPARbeta mostly reproduces the null PPARalpha-mediated reduced beta-oxidation, in addition to a shift from fast to slow fibers. In conclusion, PPARbeta is not required for maintaining skeletal muscle metabolic activity and does not compensate the lack of PPARalpha in PPARalpha null mice.

Chronic electrostimulation after nerve repair by self-anastomosis: effects on the size, the mechanical, histochemical and biochemical muscle properties.

This study tests the effects of chronic electrostimulation on denervated/reinnervated skeletal muscle in producing an optimal restoration of size and mechanical and histochemical properties. We compared tibialis anterior muscles in four groups of rats: in unoperated control (C) and 10 weeks following nerve lesion with suture (LS) in the absence of electrostimulation and in the presence of muscle stimulation with either a monophasic rectangular current (LSEm) or a biphasic modulated current (LSEb). The main results were (1) muscle atrophy was reduced in LSEm (-26%) while it was absent in LSEb groups (-8%); (2) the peak twitch amplitude decreased in LS and LSEm but not in LSEb groups, whereas the contraction time was shorter; (3) muscle reinnervation was associated with the emergence of type IIC fibers and proportions of types I, IIA and IIB fibers recovered in the superficial portion of LSEb muscles; (4) the ratio of oxidative to glycolytic activities decreased in the three groups with nerve injury and repair; however, this decrease was more accentuated in LSEm groups. We conclude that muscle electrostimulation following denervation and reinnervation tends to restore size and functional and histochemical properties during reinnervation better than is seen in unstimulated muscle.

Skeletal muscle HSP72 response to mechanical unloading: influence of endurance training.

AIMS: It has been shown that increased contractile activity results in heat shock protein 72 (HSP72) accumulation in various skeletal muscles. By contrast, there is no consensus for muscle HSP72 response to muscle disuse for short duration (5-8 days). On the basis of a greater constitutive HSP72 expression in slow-twitch muscles we tested the hypothesis that mechanical unloading for a longer period (2 weeks) would affect this phenotype to a greater extent. Secondly, we evaluated the effects of a physiological muscle heat shock protein (HSP) enhancer (endurance training) on HSP response to unloading and muscle remodelling. METHODS: Adult male Wistar rats were assigned randomly to four groups: (1) sedentary weight-bearing; (2) hindlimb-unloaded (HU) via tail suspension for 2 week; (3) trained on a treadmill (6 week) and (4) trained 6 week and then HU for 2 week. RESULTS: Unloading resulted in a preferential atrophy of slow muscles [soleus (SOL), adductor longus (AL)] and a slow-to-fast fibre transition with no change in HSP72 level. HSP72 levels were significantly lower in fast muscles [extensor digitorum longus (EDL) and plantaris (PLA)], and did not change with mechanical unloading. Endurance training was accompanied by a small (SOL) or a large (EDL, PLA) increase in HSP72 level with no change in AL. Training-induced accumulation of HSP72 disappeared with subsequent unloading in the SOL and PLA whereas HSP72 content remained elevated in EDL. CONCLUSION: The results of this study indicate that (1) after 2 weeks of unloading no change occurred in HSP72 protein levels of slow-twitch muscles despite a slow-to-fast fibre transition; and (2) the training-induced increase of HSP72 content in skeletal muscles did not attenuate fibre transition.

Effect of voluntary exercise on H2O2 release by subsarcolemmal and intermyofibrillar mitochondria.

Previous data have demonstrated that, to handle the oxidative stress encountered with training at high intensity, skeletal muscle relies on an increase in mitochondrial biogenesis, a reduced H(2)O(2) production, and an enhancement of antioxidant enzymes. In the present study, we evaluated the influence of voluntary running on mitochondrial O(2) consumption and H(2)O(2) production by intermyofibrillar mitochondria (IFM) and subsarcolemmal mitochondria (SSM) isolated from oxidative muscles in conjunction with the determination of antioxidant capacities. When mitochondria are incubated with succinate as substrate, both maximal (state 3) and resting (state 4) O(2) consumption were significantly lower in SSM than in IFM populations. Mitochondrial H(2)O(2) release per unit of O(2) consumed was 2-fold higher in SSM than in IFM. Inhibition of H(2)O(2) formation by rotenone suggests that complex I of the electron transport chain is likely the major physiological H(2)O(2)-generating system. In Lou/C rats (an inbred strain of rats of Wistar origin), neither O(2) consumption nor H(2)O(2) release by IFM and SSM were affected by long-term, voluntary wheel training. In contrast, glutathione peroxidase and catalase activity were significantly increased despite no change in oxidative capacities with long-term, voluntary exercise. Furthermore, chronic exercise enhanced heat shock protein 72 accumulation within skeletal muscle. It is concluded that the antioxidant status of muscle can be significantly improved by prolonged wheel exercise without necessitating an increase in mitochondrial oxidative capacities.

Spaceflight effects on single skeletal muscle fiber function in the rhesus monkey.

The purpose of this investigation was to understand how 14 days of weightlessness alters the cellular properties of individual slow- and fast-twitch muscle fibers in the rhesus monkey. The diameter of the soleus (Sol) type I, medial gastrocnemius (MG) type I, and MG type II fibers from the vivarium controls averaged 60 +/- 1, 46 +/- 2, and 59 +/- 2 microm, respectively. Both a control 1-G capsule sit (CS) and spaceflight (SF) significantly reduced the Sol type I fiber diameter (20 and 13%, respectively) and peak force, with the latter declining from 0.48 +/- 0.01 to 0.31 +/- 0.02 (CS group) and 0.32 +/- 0.01 mN (SF group). When the peak force was expressed as kiloNewtons per square meter (kN/m(2)), only the SF group showed a significant decline. This group also showed a significant 15% drop in peak fiber stiffness that suggests that fewer cross bridges were contracting in parallel. In the MG, SF but not CS depressed the type I fiber diameter and force. Additionally, SF significantly depressed absolute (mN) and relative (kN/m(2)) force in the fast-twitch MG fibers by 30% and 28%, respectively. The Ca(2+) sensitivity of the type I fiber (Sol and MG) was significantly reduced by growth but unaltered by SF. Flight had no significant effect on the mean maximal fiber shortening velocity in any fiber type or muscle. The post-SF Sol type I fibers showed a reduced peak power and, at peak power, an elevated velocity and decreased force. In conclusion, CS and SF caused atrophy and a reduced force and power in the Sol type I fiber. However, only SF elicited atrophy and reduced force (mN) in the MG type I fiber and a decline in relative force (kN/m(2)) in the Sol type I and MG type II fibers.

Physical exercise, aortic blood pressure, and aortic wall elasticity and composition in rats.

With a training schedule (8 weeks' treadmill running at 30 m/min up a 10% incline 5 d/wk for 90 min/day), we investigated whether exercise modifies aortic wall dimensions, composition (calcium and elastin content), or stiffness in normotensive 6-month-old male Wistar WAG/Rij rats. Maximal oxygen uptake was measured in half of the rats (n=10 per group). Wall stiffness was evaluated in the other half (9 trained and 10 untrained) on the basis of changes in thoracoabdominal pressure pulse wave velocity and differences in amplitude between the peripheral and central aortic pressure signals. Experiments were performed in nonanesthetized, unrestrained rats and then after pithing. The impact of exercise on the oxidative capacity of the plantaris muscles was evaluated with the measurement of citrate synthase activity. Training increased maximal oxygen uptake by 34% and citrate synthase activity by 40%. Mean peripheral aortic pressure increased by 6% and 19% in trained rats, under awake and pithed conditions, whereas mean central aortic pressure increased by 16%, after pithing only. All indexes of aortic stiffness were similar in trained and control rats, as were aortic wall dimensions, composition, cardiac mass, and heart rate. In conclusion, physical exercise in young rats appears to have no effect on aortic stiffness.

Effect of spaceflight on the maximal shortening velocity, morphology, and enzyme profile of fast- and slow-twitch skeletal muscle fibers in rhesus monkeys.

Weightlessness has been shown to cause limb muscle wasting and a reduced peak force and power in the antigravity soleus muscle. Despite a reduced peak power, Caiozzo et al. observed an increased maximal shortening velocity in the rat soleus muscle following a 14-day space flight. The major purpose of the present investigation was to determine if weightlessness induced an elevated velocity in the antigravity slow type I fibers of the rhesus monkey (Macaca mulatta), as well as to establish a cellular mechanism for the effect. Spaceflight or models of weightlessness have been shown to increase glucose uptake, elevate muscle glycogen content, and increase fatigability of the soleus muscle. The latter appears to be in part caused by a reduced ability of the slow oxidative fibers to oxidize fats. A second goal of this study was to establish the extent to which weightlessness altered the substrate profile and glycolytic and oxidative enzyme capacity of individual slow- and fast-twitch fibers.

Structural changes in arm muscles after microgravity.

Disuse muscle atrophy is a well-known consequence of spaceflight. However, most of the available muscle data concern lower limb muscles of rats and primates exposed to microgravity aboard Russian Cosmos biosatellites and American Space Shuttles. The purpose of our study was, therefore, to provide information concerning the effects of a 14-day spaceflight on two upper limb muscles of rhesus monkeys (Macaca mulatta). Our objective was to compare structural adaptations after 14 days of microgravity in a slow-twitch extensor muscle, i.e., the triceps, with a fast-twitch flexor muscle, i.e., the biceps. We hypothesize that muscle responses will be muscle specific, i.e., slow will differ from fast muscles, flexors will differ from extensors, and arms will differ from legs.

Muscle-specific up-regulation of rat UCP3 mRNA expression by long-term hindlimb unloading.

The effect of long-term hindlimb unloading (2 or 5 week) on the expression of uncoupling protein-3 (UCP3) gene was investigated in rat skeletal muscles. The interaction of hindlimb unloading and thyroid status was also investigated at 2 weeks. Whatever the duration, mechanical unloading induced a similar increase in UCP3 mRNA relative abundance in the slow-twitch soleus (SOL) muscle (+80%, P < 0.05), whereas no effect was observed in the fast-twitch extensor digitorum longus (EDL) muscle. Hypothyroidism down-regulated while hyperthyroidism up-regulated UCP3 mRNA relative abundance in both SOL and EDL muscles, but thyroid status did not prevent the up-regulation of UCP3 induced by 2 weeks of suspension. These data therefore indicate for the first time that long-term hindlimb unloading up-regulates muscle UCP3 gene expression in a muscle-specific manner which is independent of thyroid status.

Exercise training in chronic hypoxia has no effect on ventilatory muscle function in humans.

At the highest altitude, aerobic work is limited by environmental oxygen availability. We therefore reasoned that the hyperpnea associated with endurance training at altitude should provide a strong stimulus for adaptation of the ventilatory muscles. We measured peak inspiratory muscle pressure-flow characteristics (inspiring through graded resistors) and maximum sustainable ventilation capacity in ten permanent residents of La Paz, Bolivia (3600 m) prior to and immediately following 6 weeks of incremental endurance training. Additionally, eight local residents did no training and functioned as controls for the capacity test. While V(O2)max measured in hypoxia increased by 19% (Favier et al., 1995b. J. Appl Physiol. 78, 2286-2293.), none of the tested ventilatory variables showed significant changes. The values for the group mean slopes of maximum inspiratory pressure-flow pairs (- 10.5 vs. - 9.8 cm H2O x sec x L(-1), P=0.301; before versus after training, respectively), maximum inspiratory pressure (112.1+/-8.9 vs. 106.9+/-8.6 cmH2O, P=0.163), peak inspiratory flow (9.8+/-0.41 vs. 10.2+/-0.55 L x sec(-1) P=0.172) and the maximum volitional volume in 12 sec (43.9+/-2.4 vs. 45.6+/-2.4 L in 12 sec, P=0.133) were unchanged with exercise training. Likewise, maximal sustainable minute volume was not different between post-training and control subjects (177.4+/-7.9 vs. 165.4+/-8.4 L x min(-1), P=0.141). These data support the concept that endurance training fails to elicit functional adaptations in ventilatory muscles in humans, even when exercise is done in hypoxia.

Effects of bedrest on deltoideus muscle morphology and enzymes.

To examine the effects of unweighting on the structural and metabolic adaptations of a non-postural muscle, deltoideus muscle biopsies were taken in seven male healthy subjects, before and after a 37 day bedrest. Myofibrillar ATPase histochemistry demonstrated no change in fibre type distributions (I, IIA, IIB), in fibre cross-sectional areas nor in capillary supply. No difference was noted in enzyme activities of oxidative metabolism (citrate synthase, 3-hydroxy-acyl-CoA dehydrogenase), and glycolysis (hexokinase, lactate dehydrogenase). Electron microscopy showed a decrease in the volume density of lipids but no change in mitochondrial volume density and distribution. The results indicate that bedrest induces no major morphological and biochemical changes in deltoideus muscle, contrary to what was previously reported in vastus lateralis muscle. This lack of changes is probably related to an unaltered deltoideus muscle use.

Effect of endurance training on mRNA expression of uncoupling proteins 1, 2, and 3 in the rat.

Endurance exercise training has been shown to decrease diet-induced thermogenesis (DIT) in rats and humans. In rodents, most thermogenesis is thought to occur in brown adipose tissue via activation of the uncoupling protein-1 (UCP1) and in skeletal muscle. Since the level of UCP1 mRNA in rat BAT was reported to be unmodified by exercise training, the newly described uncoupling proteins UCP2 and UCP3 could be responsible for the decreased DIT in trained rats. UCP3 mRNA levels in endurance-trained rats were found to be reduced by 76% and 59% in tibialis anterior and soleus muscles, respectively. UCP2 mRNA levels were also decreased in tibialis anterior and in heart by 54% and 41%, respectively. Neither white adipose tissue UCP2 nor brown adipose tissue UCP1, UCP2, and UCP3 mRNA levels were modified. The results of this study show that a need for a higher metabolic efficiency is associated with decreased mRNA expression of the uncoupling proteins in skeletal and heart muscles, which would decrease energy dissipation in these tissues. The down-regulation of UCP3 and UCP2 expressions might also contribute to the rapid weight gain known to occur when exercise training ceased.

Structural and functional adaptations of skeletal muscle to weightlessness.

This brief review examines the effects of weightlessness on the structure and function of human and rat skeletal muscle. Data collected from spaceflights or Earth-based models suggest that a rapid atrophy (5-8 days) occurs in lower limb muscles associated with impairments in muscle strength, physical work capacity and locomotor coordination. The reduction in muscle cross-sectional area cannot entirely explain the loss of strength in postural muscles suggesting a reduced neural drive. Muscle atrophy is accompanied by a general shift in the contractile and enzymatic profiles of a slow-twitch oxidative muscle toward that of a fast twitch glycolytic muscle. In humans, limited structural and functional data collected in astronauts after short periods of microgravity are qualitatively similar to those observed in rats after real or simulated microgravity suggesting that animal models are relevant to muscle research in space. A preventive prescription i.e. exercise or pharmacological treatment, cannot be proposed until future research has better defined the basic mechanisms of muscle plasticity during long duration spaceflights.

The interplay of central and peripheral factors in limiting maximal O2 consumption in man after prolonged bed rest.

1. The effects of bed rest on the cardiovascular and muscular parameters which affect maximal O2 consumption (VO2,max) were studied. The fractional limitation of VO2,max imposed by these parameters after bed rest was analysed. 2. The VO2,max, by standard procedure, and the maximal cardiac output (Qmax), by the pulse contour method, were measured during graded cyclo-ergometric exercise on seven subjects before and after a 42-day head-down tilt bed rest. Blood haemoglobin concentration ([Hb]) and arterialized blood gas analysis were determined at the highest work load. 3. Muscle fibre types, oxidative enzyme activities, and capillary and mitochondrial densities were measured on biopsy samples from the vastus lateralis muscle before and at the end of bed rest. The measure of muscle cross-sectional area (CSA) by NMR imaging at the level of biopsy site allowed computation of muscle oxidative capacity and capillary length. 4. The VO2,max was reduced after bed rest (-16.6%). The concomitant decreases in Qmax (-30.8%), essentially due to a change in stroke volume, and in [Hb] led to a huge decrease in O2 delivery (-39.7%). 5. Fibre type distribution was unaffected by bed rest. The decrease in fibre area corresponded to the significant reduction in muscle CSA (-17%). The volume density of mitochondria was reduced after bed rest (-16.6%), as were the oxidative enzyme activities (-11%). The total mitochondrial volume was reduced by 28.5%. Capillary density was unchanged. Total capillary length was 22.2% lower after bed rest, due to muscle atrophy. 6. The interaction between these muscular and cardiovascular changes led to a smaller reduction in VO2,max than in cardiovascular O2 transport. Yet the latter appears to play the greatest role in limiting VO2,max after bed rest (> 70% of overall limitation), the remaining fraction being shared between peripheral O2 diffusion and utilization.

Effects of acute and chronic hindlimb suspension on sensitivity and responsiveness to insulin in the rat soleus muscle.

The effects of acute (24 h) and chronic (5 weeks) hindlimb suspension on insulin-stimulated glucose utilization by the rat soleus muscle were studied in vitro. Hindlimb suspension resulted in an enhancement of basal glucose transport, lactate production, and glycogen synthesis. An increase in the sensitivity of these processes to insulin occurred as early as 24 h and persisted for 5 weeks of the muscle unloading. An increased responsiveness to insulin was found only for glucose transport after 24 h. The present data do not support the concept that the enhanced glucose utilization and improved muscle insulin sensitivity during hindlimb suspension are related to muscle atrophy, which is not observed in the early stage of muscle unweighting.

Lactate and epinephrine during exercise in altitude natives.

We tested the hypothesis that the reported low blood lactate accumulation ([La]) during exercise in altitude-native humans is refractory to hypoxianormoxia transitions by investigating whether acute changes in inspired O2 fraction (FIo2) affect the [La] vs. power output (W) relationship or, alternatively, as reported for lowlanders, whether changes in [La] vs. W on changes in FIo2 are related to changes in blood epinephrine concentration ([Epi]). Altitude natives [n = 8, age 24 +/- 1 (SE) yr, body mass 62 +/- 3 kg, height 167 +/- 2 cm] in La Paz, Bolivia (3,600 m) performed incremental exercise with two legs and one leg in chronic hypoxia and acute normoxia (AN). Submaximal one- and two-leg O2 uptake (Vo2) vs. W relationships were not altered by FIo2. AN increased two-leg peak Vo2 by 10% and peak W by 7%. AN paradoxically decreased one-leg peak Vo2 by 7%, whereas peak W remained the same. The [La] vs. W relationships were similar to those reported in unacclimatized lowlanders. There was a shift to the right on AN, and maximum [La] was reduced by 7 and 8% for one- and two-leg exercises, respectively. [Epi] and [La] were tightly related (mean r = 0.81) independently of FIo2. Thus normoxia attenuated the increment in both [La] and [Epi] as a function of W, whereas the correlation between [La] and [Epi] was unaffected. These data suggest loose linkage of glycolysis to oxidative phosphorylation under influence from [Epi]. In conclusion, high-altitude natives appear to be not fundamentally different from lowlanders with regard to the effect of acute changes in FIo2 on [La] during exercise.

Muscle tissue adaptations of high-altitude natives to training in chronic hypoxia or acute normoxia.

Twenty healthy high-altitude natives, residents of La Paz, Bolivia (3,600 m), participated in 6 wk of endurance exercise training on bicycle ergometers, 5 times/wk, 30 min/session, as previously described in normoxia-trained sea-level natives (H. Hoppeler, H. Howald, K. E. Conley, S. L. Lindstedt, H. Claassen, P. Vock, and E. R. Weibel. J. Appl. Physiol. 59: 320-327, 1985). A first group of 10 subjects was trained in chronic hypoxia (HT; barometric pressure = 500 mmHg; inspired O2 fraction = 0.209); a second group of 10 subjects was trained in acute normoxia (NT; barometric pressure = 500 mmHg; inspired O2 fraction = 0.314). The workloads were adjusted to approximately 70% of peak O2 consumption (VO2peak) measured either in hypoxia for the HT group or in normoxia for the NT group. VO2peak determination and biopsies of the vastus lateralis muscle were taken before and after the training program. VO2peak in the HT group was increased (14%) in a way similar to that in NT sea-level natives with the same protocol. Moreover, VO2peak in the NT group was not further increased by additional O2 delivery during the training session. HT or NT induced similar increases in muscle capillary-to-fiber ratio (26%) and capillary density (19%) as well as in the volume density of total mitochondria and citrate synthase activity (45%). It is concluded that high-altitude natives have a reduced capillarity and muscle tissue oxidative capacity; however, their training response is similar to that of sea-level residents, independent of whether training is carried out in hypobaric hypoxia or hypobaric normoxia.

Rat muscle plasticity in response to simulated or real microgravity.

NASA: Data concerning muscle plasticity in real or simulated microgravity is discussed. Possible mechanisms responsible for the muscular atrophy associated with microgravity are explored, including changes in muscle protein synthesis, fast- and slow-twitch fiber specific changes, various metabolic alterations, blood supply and other factors. The authors conclude that a combination of local and systemic factors are responsible for the observed changes in muscle physiology.^ieng