Daily short-period gravitation can prevent functional and structural changes in arteries of simulated microgravity rats.

This study was designed to clarify whether simulated microgravity-induced differential adaptational changes in cerebral and hindlimb arteries could be prevented by daily short-period restoration of the normal distribution of transmural pressure across arterial vasculature by either dorsoventral or footward gravitational loading. Tail suspension (Sus) for 28 days was used to simulate cardiovascular deconditioning due to microgravity. Daily standing (STD) for 1, 2, or 4 h, or +45 degrees head-up tilt (HUT) for 2 or 4 h was used to provide short-period dorsoventral or footward gravitational loading as countermeasure. Functional studies showed that Sus alone induced an enhancement and depression in vasoconstrictor responsiveness of basilar and femoral arterial rings, respectively, as previously reported. These differential functional alterations can be prevented by either of the two kinds of daily gravitational loading treatments. Surprisingly, daily STD for as short as 1 h was sufficient to prevent the differential functional changes that might occur due to Sus alone. In morphological studies, the effectiveness of daily 4-h HUT or 1-h STD in preventing the differential remodeling changes in the structure of basilar and anterior tibial arteries induced by Sus alone was examined by histomorphometry. The results showed that both the hypertrophic and atrophic changes that might occur, respectively, in cerebral and hindlimb arteries due to Sus alone were prevented not only by daily HUT for 4 h but also by daily STD even for 1 h. These data indicate that daily gravitational loading by STD for as short as 1 h is sufficient to prevent differential adaptational changes in function and structure of vessels in different anatomic regions induced by a medium-term simulated microgravity.     

Effectiveness of intermittent -Gx gravitation in preventing deconditioning due to simulated microgravity.

This study was designed to compare the effectiveness of daily short-duration -Gx gravity exposure in preventing adverse changes in skeletal and cardiac muscles and bone due to simulated microgravity. Tail suspension for 28 days was used to simulate microgravity-induced deconditioning effects. Daily standing (STD) at 1 G for 1, 2, or 4 h/day or centrifugation (CEN) at 1.5 or 2.6 G for 1 h/day was used to provide -Gx gravitation as a countermeasure. The results indicate that the minimum gravity exposure requirements vary greatly in different systems. Cardiac muscle is most responsive to such treatment: 1 h/day of -Gx gravitation by STD was sufficient to prevent adverse changes in myocardial contractility; bone is most resistant: 4 h/day of -Gx gravitation only partially alleviated the adverse changes in physical and mechanical properties of the femur. The responsiveness of skeletal muscle is moderate: 4 h/day of -Gx gravitation prevented mass reduction and histomorphometric changes in the soleus muscle during a 28-day simulation period. Increasing gravitational intensity to 2.6 G showed less benefit or no additional benefit in preventing adverse changes in muscle and bone. The present work suggests that system specificity in responsiveness to intermittent gravity exposure should be considered one of the prerequisites in proposing intermittent artificial gravity as a potential countermeasure.     

间断性人工重力对模拟失重大鼠颈总动脉平滑肌细胞凋亡的影响

目的:研究3周模拟失重大鼠颈总动脉平滑肌细胞凋亡的变化及间断性人工重力对其的影响。方法:以尾部悬吊大鼠(SUS)模拟失重,同期每天悬吊23h、站立1h(STD)模拟间断性人工重力的对抗效果,用M30染色及Tunel染色方法观察3周SUS组、同步对照(CON)组及STD组颈总动脉平滑肌细胞早期和中晚期的凋亡情况,并用免疫组织化学方法及Western blot印迹方法观察各组大鼠颈总动脉组织Caspase-3的蛋白表达变化。结果:与CON组比较,SUS组大鼠颈总动脉平滑肌细胞M30染色阳性细胞明显减少,STD组M30染色阳性细胞较CON组及SUS组显著增加;SUS组Tunel染色阳性细胞较CON组及STD组显著减少,STD组Tunel染色阳性细胞较CON组及SUS组显著增加;SUS组Caspase-3的表达较CON组显著降低(P<0.05),STD组Caspase-3的表达较CON组及SUS组显著增高(P<0.01)。结论:模拟失重可引起大鼠颈总动脉平滑肌细胞凋亡减少,每日1 h的-Gx对抗使颈总动脉的凋亡增加。Caspase-3可能在调控模拟失重所致血管组织平滑肌细胞的凋亡中发挥作用。     

Daily short-term head-up tilt can prevent structural changes in arteries of tail-suspended rats

The aim of the present study was to investigate the effectiveness of daily 4-h head up tilt (HUT) in preventing the structural changes in arteries from rats induced by tail-suspension (TS). TS rat model was used to simulate the physiological effects of microgravity. Daily 4-h HUT was used to simulate the effect of intermittent artificial gravity (IAG). The results showed that TS alone induced an increase in the vessel media layer cross-sectional area (CSA), media wall thickness (T), mean number of smooth muscle cell layers (NCL) in basilar artery, and a decrease in these parameters in anterior tibial artery. Both the hypertrophic and atrophic changes in these arteries that might occur due to TS alone can be prevented by a 4h/d HUT treatment.     

Daily head-up tilt,standing or centrifugation can prevent vasoreactivity changes in arteries of simulated weightless rats

Artificial gravity will be considered for long-duration spaceflight missions. Recent studies have shown that continuous exposure to gravity does not appear necessary to prevent the adverse effects of weightlessness, instead intermittent exposure may suffice. Vernikos et al reported that 1 G intermittent exposures with and without walking exercise was effective in preventing physiologic deconditioning with 4-d, -6 degrees bedrest, and standing at 1 G was most effective in preventing orthostatic intolerance. Our previous work has demonstrated differential adaptational changes in structure, function, and innervation state of arterial vasculature in different body regions of rat during simulated weightlessness and further suggested that these changes might be one of the most important mechanisms accounting for postflight orthostatic intolerance. We therefore designed the present study involving a comprehensive evaluation of the effect of intermittent G exposure in preventing the differential functional changes of the arteries at the end of 3-wk head-down tail suspension in rats to answer the following questions: (1) do intermittent G exposure have counteracting effect in preventing differential functional changes in arterial vasculature with tail-suspension? (2) among the treatments used in the present study, i.e., head-up tilt (HUT), standing (STD), and centrifugation (CEN), what kind of exposure is more effective? (3) how much time in daily 1 G exposure is needed to maintain the normal (1 G) vascular responsiveness?     

Intermittent acceleration as a countermeasure to soleus muscle atrophy.

The centrifuge proposed for the Space Station will most likely be used, in part, for countermeasure studies. At present, there is a paucity of information concerning the duration and frequency of acceleration necessary to counteract the atrophy process associated with microgravity. The present study was designed to investigate intermittent acceleration during non-weight bearing of the soleus muscle and its resultant effects on muscular atrophy. Each day rats were removed from hindlimbs suspension and accelerated to 1.2 g for four 15-min periods evenly spaced over a 12-h interval. The soleus muscle experienced non-weight bearing the remaining 23 h each day. This paradigm, when repeated for 7 days, did not completely maintain the mass of soleus muscle, which was 84% of control. Interestingly, the identical protocol utilizing ground support in lieu of acceleration successfully maintained the soleus muscle mass. The failure of the centrifugation protocol to adequately maintain soleus muscle mass might be due to an undefined stress placed on the animals inherent in centrifugation itself. This stress may also explain the transient decline in food intake of the intermittent acceleration group on the 2nd and 3rd days of treatment. Also, these data support the concept that the frequency of exposure, as opposed to the duration of exposure, to weight bearing during hindlimb unweighting seems to be the more important determinant of maintaining postural muscle mass.     

Skeletal muscle proteolysis in response to short-term unloading in humans.

Skeletal muscle atrophy is evident after muscle disuse, unloading, or spaceflight and results from decreased protein content as a consequence of decreased protein synthesis, increased protein breakdown or both. At this time, there are essentially no human data describing proteolysis in skeletal muscle undergoing atrophy on Earth or in space, primarily due to lack of valid and accurate methodology. This particular study aimed at assessing the effects of short-term unloading on the muscle contractile proteolysis rate. Eight men were subjected to 72-h unilateral lower limb suspension (ULLS) and intramuscular interstitial levels of the naturally occurring proteolytic tracer 3-methylhistidine (3MH) were measured by means of microdialysis before and on completion of this intervention. The 3MH concentration following 72-h ULLS (2.01 +/- 0.22 nmol/ml) was 44% higher (P < 0.05) than before ULLS (1.56 +/- 0.20 nmol/ml). The present experimental model and the employed method determining 3MH in microdialysates present a promising tool for monitoring skeletal muscle proteolysis or metabolism of specific muscles during conditions resulting in atrophy caused by, e.g., disuse and real or simulated microgravity. This study provides evidence that the atrophic processes are evoked rapidly and within 72 h of unloading and suggests that countermeasures should be employed in the early stages of space missions to offset or prevent muscle loss during the period when the rate of muscle atrophy is the highest.     

Differential regulation of L-type Ca2+ channels in cerebral and mesenteric arteries after simulated microgravity in rats and its intervention by standing

This study was designed to clarify whether simulated microgravity can induce differential changes in the current and protein expression of the L-type Ca(2+) channel (Ca(L)) in cerebral and mesenteric arteries and whether these changes can be prevented by daily short-duration -G(x) exposure. Tail suspension [hindlimb unloading (HU)] for 3 and 28 days was used to simulate short- and medium-term microgravity-induced deconditioning effects. Standing (STD) for 1 h/day was used to provide -G(x) as a countermeasure. Whole cell patch-clamp experiments revealed an increase in current density of Ca(L) of vascular smooth muscle cells (VSMCs) isolated from cerebral arteries of rats subjected to HU and a decrease in VSMCs from mesenteric arteries. Western blot analysis revealed a significant increase and decrease of Ca(L) channel protein expression in cerebral and small mesenteric arterial VSMCs, respectively, only after 28 days of HU. STD for 1 h/day did not prevent the increase of Ca(L) current density in cerebral arterial VSMCs, but it prevented completely (within 3 days) and partially (28 days) the decrease of Ca(L) current density in small mesenteric arterial VSMCs. Consistent with the changes in Ca(L) current, STD for 1 h/day did not prevent the increase of Ca(L) expression in cerebrovascular myocytes but did prevent the reduction of Ca(L) expression in mesenteric arterial VSMCs subjected to 28 days of HU. These data indicate that simulated microgravity up- and downregulates the current and expression of Ca(L) in cerebral and hindquarter VSMCs, respectively. STD for 1 h/day differentially counteracted the changes of Ca(L) function and expression in cerebral and hindquarter arterial VSMCs of HU rats, suggesting the complexity of the underlying mechanisms in the effectiveness of intermittent artificial gravity for prevention of postflight cardiovascular deconditioning, which needs further clarification.     

失重状态下肌萎缩研究进展

失重条件下人和动物生理状态会发生一系列的变化,其中骨骼肌萎缩和力量下降较为显著,目前其发生的机制仍不明确且缺少特效的干预措施。本文从肌肉湿重及肌纤维横截面积的变化、肌纤维类型的变化、肌纤维超微结构的变化、肌梭的适应性变化四个方面进行简要阐述,探讨肌肉萎缩的可能发生机制。     

Effects of gravitational unloading on back muscles tone

The study involved 12 healthy volunteers who were exposed to 6-h or 3-day dry immersion (DI). The back muscle tone was recorded by resonance vibrography using parameters of transverse stiffness of the muscles under study. The measurements in 3-day DI were performed twice before DI, daily in the course of DI, and twice after its completion; in the short-term (6-h) DI, the testing was carried out twice before DI, 1 and 4 h after the beginning of DI, and during the first hour after DI completion. It has been shown that the absence of support is followed by a sharp decrease in the back extensor muscles tone, which has the maximal values during the first hours and days of DI. The possible role of back muscle atony in the development of well-known phenomena of the spine length increase and back pain appearance observed at the beginning of space flights and in the first days after landing, as well as under the conditions of simulated microgravity (DI and head-down bed rest), is discussed.     

Centrifugal intensity and duration as countermeasures to soleus muscle atrophy

Mechanical acceleration is a countermeasure that may be employed to prevent atrophy of slow-twitch muscle during non-weight bearing. In the present study, daily centrifugation of rats for different durations (1 or 2 h) and at different gravitational intensities (1.5 or 2.6 G) was used to test whether mechanical acceleration could ameliorate the atrophy of the soleus muscle induced by non-weight bearing (tail-traction model). The soleus muscle atrophied 32% during 7 days of non-weight bearing without countermeasures. Centrifugation treatment did not completely prevent atrophy relative to precontrol wet weight of the soleus muscle. Non-weight-bearing groups receiving 2-h daily treatments of 1, 1.5, or 2.6 G had 48, 56, and 65%, respectively, of the atrophy observed in the non-weight-bearing-only group compared with the precontrol group. No evidence was obtained that centrifugation at 2.6 G was more effective than exposure to 1 or 1.5 G as a countermeasure to non-weight-bearing-induced atrophy of the soleus muscle.     

模拟微重力下秀丽隐杆线虫的力学感知和肌萎缩的发生机制——太空飞行线虫试验地面平行对照研究部分

目前,微重力导致肌萎缩的分子机制尚不清楚,重力感知是该事件发生的关键环节．为了回答这一问题,在此之前首先实施了太空线虫试验,这部分结果已经在本刊报道过．而本次研究主要是在地面上建立了模拟微重力环境,观察处理后秀丽隐杆线虫（C.elegans）体壁肌细胞结构和功能的变化,一方面用于验证太空试验,同时比较两种处理结果的异同,以便于评价地面模拟微重力的有效性．经过14天19．5h旋转模拟微重力处理后,对线虫生存率和运动能力进行了观察,并检测了几个重要的肌相关基因表达和蛋白质水平．模拟微重力下线虫生存率没有明显变化,但运动频率显著下降,爬行轨迹也发生了轻微改变,运动幅度降低,提示线虫运动功能出现障碍．从形态学上观察发现：肌球蛋白A（myosin A）免疫荧光染色显示模拟微重力组肌纤维面积缩小,而肌细胞致密体（dense-body）染色可见荧光亮度下降．这些结果直接提示模拟微重力使线虫出现了肌萎缩．随后Western blotting试验结果揭示,模拟微重力组线虫体壁肌的主要结构蛋白——myosin A含量减少,进一步确证了微重力性肌萎缩发生．在基因水平,旋转后抗肌萎缩蛋白基因（dys-1）表达明显上升,而hlh-1,unc-54,myo-3和egl-19的mRNA水平均下调,提示dys-1在骨骼肌感知和传导力学信息方面有重要作用,而hlh-1,unc-54,myo-3和egl-19则分别从结构和功能两个途径促进了微重力性肌萎缩的发生和发展．本次试验所得到的结果同太空飞行试验结果十分相似,一方面强化了太空试验结论,另一方面说明在地面上模拟微重力对生物体进行研究是有效可行的,将有助于提高太空试验的质量．     

Regulation of Protein Synthesis in Inactivated Skeletal Muscle: Signal Inputs,Protein Kinase Cascades,and Ribosome Biogenesis

Disuse atrophy of skeletal muscles is characterized by a significant decrease in the mass and size of muscle fibers. Disuse atrophy develops as a result of prolonged reduction in the muscle functional activity caused by bed rest, limb immobilization, and real or simulated microgravity. Disuse atrophy is associated with the downregulation of protein biosynthesis and simultaneous activation of protein degradation. This review is focused on the key molecular mechanisms regulating the rate of protein synthesis in mammalian skeletal muscles during functional unloading.     

Rat muscle plasticity in response to simulated or real microgravity

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.     

Studies on atrophic change of soleus muscle and its countermeasures in suspended rat

In the environment of microgravity, the disused atrophy of skeletal muscle, especially leg's muscle, would occur. The three purposes of this study were: 1. To observe the dynamic changes of disused atrophy of skeletal muscle under simulated weightlessness; 2. To approach the mechanism of disused atrophy of muscle; 3. To approach the countermeasures for reducing the degree of atrophy of muscle.     

Mitochondrial adaptations in skeletal muscle cells in mammals exposed to gravitational unloading

It is known that exposure to actual or simulated weightlessness is often accompanied by decreased muscle dynamic performance, and increased level of blood lactate accumulation. Decreased mitochondrial content found in fibers of the working muscles is considered to be one of the possible causes for those changes. Studies on oxidative potential of the muscle cell (i.e. capacity of the cell to oxidative energy production) under conditions of altered gravity have been carried out since late 70-ties. It was shown that the relatively short term spaceflight and hindlimb suspension induced significant decrease oxidative enzyme activities and mitochondrial volume density in rat fast muscle. However postural soleus muscle failed to exhibit similar changes, although the absolute mitochondrial content was found to be sufficiently lower after exposure to simulated microgravity. This phenomenon allowed to conclude that the pronounced soleus fiber atrophy masked the proportional absolute decrease in oxidative potential which failed to be revealed as subsequent changes in mitochondrial volume density and oxidative enzyme activity. It is also important, that biosatellite studies exposed considerable changes in mitochondria distribution pattern inside m. soleus fibers: volume density of mitochondria (and, correspondingly, activity of oxidative enzymes) increases (or does not change) in the center of fiber, and decreases at its periphery, in subsarcolemmal area. However the time course of mitochondrial alterations development (particularly during long-duration exposures to real or simulated microgravity) and some peculiarities of the mitochondria distribution were not described yet. Also, materials dealing with simultaneous time-course comparative analysis of mitochondrial characteristics and indices of physiological cost of submaximal exercise are very rare. The present paper is purposed to compare the data, obtained in several experimental studies, allowed to analyze the possible contribution of muscle mitochondria changes to changes in metabolic cost of submaximal exercise and the time-course dynamics of mitochondrial characteristics under conditions of actual or simulated gravitational unloading.     

In-vivo and ex-vivo studies on region-specific remodeling of large elastic arteries due to simulated weightlessness and its prevention by gravity-based countermeasure

The present study was designed to test the hypothesis that a medium-term simulated microgravity can induce region-specific remodeling in large elastic arteries with their innermost smooth muscle (SM) layers being most profoundly affected. The second purpose was to examine whether these changes can be prevented by a simulated intermittent artificial gravity (IAG). The third purpose was to elucidate whether vascular local renin-angiotensin system (L-RAS) plays an important role in the regional vascular remodeling and its prevention by the gravity-based countermeasure. This study consisted of two interconnected series of in-vivo and ex-vivo experiments. In the in-vivo experiments, the tail-suspended, hindlimb unloaded rat model was used to simulate microgravity-induced cardiovascular deconditioning for 28 days (SUS group); and during the simulation period, another group was subjected to daily 1-hour dorso-ventral (-G(x)) gravitation provided by restoring to normal standing posture (S + D group). The activity of vascular L-RAS was evaluated by examining the gene and protein expression of angiotensinogen (Ao) and angiotensin II receptor type 1 (AT1R) in the arterial wall tissue. The results showed that SUS induced an increase in the media thickness of the common carotid artery due to hypertrophy of the four SM layers and a decrease in the total cross-sectional area of the nine SM layers of the abdominal aorta without significant change in its media thickness. And for both arteries, the most prominent changes were in the innermost SM layers. Immunohistochemistry and in situ hybridization revealed that SUS induced an up- and down-regulation of Ao and AT1R expression in the vessel wall of common carotid artery and abdominal aorta, respectively, which was further confirmed by Western blot analysis and real time PCR analysis. Daily 1-hour restoring to normal standing posture over 28 days fully prevented these remodeling and L-RAS changes in the large elastic arteries that might occur due to SUS alone. In the ex-vivo experiments, to elucidate the important role of transmural pressure in vascular regional remodeling and differential regulation of L-RAS activity, we established an organ culture system in which rat common carotid artery, held at in-vivo length, can be perfused and pressurized at varied flow and pressure for 7 days. In arteries perfused at a flow rate of 7.9 mL/min and pressurized at 150 mmHg, but not at 0 or 80 mmHg, for 3 days led to an augmentation of c-fibronectin (c-FN) expression, which was also more markedly expressed in the innermost SM layers, and an increase in Ang II production detected in the perfusion fluid. However, the enhanced c-FN expression and increased Ang II production that might occur due to a sustained high perfusion pressure alone were fully prevented by daily restoration to 0 or 80 mmHg for a short duration. These findings from in-vivo and ex-vivo experiments have provided evidence supporting our hypothesis that redistribution of transmural pressures might be the primary factor that initiates region-specific remodeling of arteries during microgravity and the mechanism of IAG is associated with an intermittent restoration of the transmural pressures to their normal distribution. And they also provide support to the hypothesis that L-RAS plays an important role in vascular adaptation to microgravity and its prevention by the IAG countermeasure.     

Baroreflex control of muscle sympathetic nerve activity after 120 days of 6 degrees head-down bed rest

To examine how long-lasting microgravity simulated by 6 degrees head-down bed rest (HDBR) induces changes in the baroreflex control of muscle sympathetic nerve activity (MSNA) at rest and changes in responses of MSNA to orthostasis, six healthy male volunteers (range 26-42 yr) participated in Valsalva maneuver and head-up tilt (HUT) tests before and after 120 days of HDBR. MSNA was measured directly using a microneurographic technique. After long-term HDBR, resting supine MSNA and heart rate were augmented. The baroreflex slopes for MSNA during Valsalva maneuver (in supine position) and during 60 degrees HUT test, determined by least-squares linear regression analysis, were significantly steeper after than before HDBR, whereas the baroreflex slopes for R-R interval were significantly flatter after HDBR. The increase in MSNA from supine to 60 degrees HUT was not different between before and after HDBR, but mean blood pressure decreased in 60 degrees HUT after HDBR. In conclusion, the baroreflex control of MSNA was augmented, whereas the same reflex control of R-R interval was attenuated after 120 days of HDBR.     

Muscle progenitor cell proliferation during passive stretch of unweighted soleus in dystrophin deficient mice

Dystrophin, subsarcolemmal protein communicating muscle fiber cytoskeleton to extracellular matrix, is believed to be involved in mechanical signal transduction. The experiment was carried out to assess the role of dystrophin in passive stretch-induced preventing unloaded muscle fiber atrophy and possible linkage between this protein and muscle progenitor (satellite cells) proliferation activity. The study was performed on two months old C57 black and mdx (dystrophin-deficient) mice. Passive stretch resulted in attenuating atrophy development in two fiber types of both C57 black and mdx mice. Altered dystrophin synthesis in mdx mice had virtually no effect on passive stretch preventive action. Thus the hypothesis about dystrophin key role in mediating stretch-induced hypertrophy effects didn't find its confirmation concerning gravitational unloading atrophy. Chronic hindlimb unloading downregulated SC proliferative activity in soleus muscle, passive stretch drastically increased proliferation both in C57 and mdx mice. Thus we observed no relationship between altered dystrophin synthesis and satellite cell proliferation activity in soleus muscle under conditions of simulated microgravity and concurrent passive stretch.     

Muscles in microgravity: from fibres to human motion

In simulated or actual microgravity, human and animal postural muscles undergo substantial atrophy: after about 270 days, the muscle mass attains a constant value of about 70% of the initial one. Most animal studies reported preferential atrophy of slow twitch fibres whose mechanical properties change towards the fast type. However, in humans, at the end of a 42-days bed rest study, a similar atrophy of slow and fast fibres was observed. After microgravity, the maximal force of several muscle groups showed a substantial decrease (6-25% of pre-flight values). The maximal power during very short "explosive" efforts of 0.25-0.30s showed an even greater fall, being reduced to 65% after 1 month and to 45% (of pre-flight values) after 6 months. The maximal power developed during 6-7s "all-out" bouts on an isokinetic cycloergometer was reduced to a lesser extent, attaining about 75% of pre-flight values, regardless of the flight duration. In these same subjects, the muscle mass of the lower limbs declined by only 9-13%. Thus, a substantial fraction of the observed decreases of maximal power is probably due to a deterioration of the motor co-ordination brought about by the absence of gravity. To prevent this substantial decay of maximal absolute power, we propose that explosive exercise be added to the daily in-flight training schedule. We also describe a system aimed at reducing cardiovascular deconditioning wherein gravity is simulated by the centrifugal acceleration generated by the motion of two counter rotating bicycles ridden by the astronauts on the inner wall of a cylindrical space module. Finally, cycling on circular or elliptical tracks may be useful to reduce cardiovascular deconditioning in permanently manned lunar bases. Indeed, on the curved parts of the path, a cyclist generates an outward acceleration vector (ac). To counterbalance ac, the cyclist must lean inwards, so that the vectorial sum of ac plus the lunar gravity tends to the acceleration of gravity prevailing on Earth.