Influence of altered gravity on the cytochemical localization of cytochrome oxidase activity in central and peripheral gravisensory systems in developing cichlid fish.

Cichlid fish larvae were reared from hatching to active free swimming under different gravity conditions: natural environment, increased acceleration in a centrifuge, simulated weightlessness in a clinostat and near weightlessness during space flight. Cytochrome oxidase activity was analyzed semiquantitatively on the ultrastructural level as a marker of regional neuronal activity in a primary, vestibular brainstem nucleus and in gravity receptive epithelia in the inner ear. Our results show, that gravity seems to be positively correlated with cytochrome oxidase activity in the magnocellular nucleus of developing fish brain. In the inner ear the energy metabolism is decreased under microgravity concerning utricle but not saccule. Hypergravity has no effect on cytochrome oxidase activity in sensory inner ear epithelia.     

Influence of proprioceptive information on space orientation on the ground and in orbital weightlessness.

Conscious space orientation depends on afferent information from different sense organs including the labyrinth, the eyes, tactile cues from the skin, joint receptors, muscle spindles, tendon organs and possibly viscera. An important role is played by impulses from the cervical position receptors in interaction with concomitant information from the otolith system. In order to isolate the effect of cervical position receptors from that of the otolith system, space experiments in orbital weightlessness and in parabolic aircraft flight were performed. It was found that stimulation of the neck receptors in weightlessness markedly influences the perception of the subjective vertical and horizontal and in addition has a weak effect on ocular torsion.     

Calcium influx through stretch-activated channels mediates microfilament reorganization in osteoblasts under simulated weightlessness

We have explored the role of Ca²⁺ signaling in microfilament reorganization of osteoblasts induced by simulated weightlessness using a random positioning machine (RPM). The RPM-induced alterations of cell morphology, microfilament distribution, cell proliferation, cell migration, cytosol free calcium concentration ([Ca²⁺]_i), and protein expression in MG63 osteoblasts were investigated. Simulated weightlessness reduced cell size, disrupted microfilament, inhibited cellular proliferation and migration, and induced an increase in [Ca²⁺]_i in MG63 human osteosarcoma cells. Gadolinium chloride (Gd), an inhibitor for stretch-activated channels, attenuated the increase in [Ca²⁺]_i and microfilament disruption. Further, the expression of calmodulin was significantly increased by simulated weightlessness, and an inhibitor of calmodulin, W-7, aggravated microfilament disruption. Our findings demonstrate that simulated weightlessness induces Ca²⁺ influx through stretch-activated channels, then results in microfilament disruption.     

Early development in aquatic vertebrates in near weightlessness during the D-2 Mission STATEX project.

Aboard the German-Spacelab-Mission D-2 the project "Gravity Perception and Neuronal Plasticity (STATEX II)" was performed. STATEX is for STATolith EXperiment. Objects were growing tadpoles of the South African Toad (Xenopus laevis D.) and a juvenile cichlid fish (Oreochromis mossambicus). The results give a broader base for the understanding of how environmental stimuli (e.g. linear accelerations) affect the development and function of the gravity perceiving systems in these two vertebrates. These systems are accepted as models for the human vestibulum. Results of experiments in hyper-g (up to 5 g), simulated weightlessness (Fast-rotating-clinostat) and parabolic flights are compared and discussed.     

Protozoa as model systems for the study of cellular responses to altered gravity conditions.

The orientation behavior of Paramecium changed in a similar way after transition to conditions of free-fall in a sounding rocket and after transition to conditions of simulated weightlessness on a fast rotating clinostat. After a period of residual orientation, Paramecium cells distributed themselves randomly 80 s (120 s) after onset of free-fall (simulated weightlessness). Swimming velocity increased significantly; however, the increase was transient and subsided after 3 min in the rocket experiments, while the velocity remained enhanced even during 2 h of rotation on a fast clinostat. Trichocysts were present and without morphological changes in Paramecium cells which had been exposed to a rocket flight, as well as to fast or slow rotation on a clinostat. Regeneration of the oral apparatus of Stentor and morphogenesis of Eufolliculina proceeded normally on the clinostat. The results demonstrate that the clinostat is a useful tool to simulate the conditions of weightlessness on earth and to detect gravisensitive cellular functions.     

Effects of simulated weightlessness on fish otolith growth: Clinostat versus Rotating-Wall Vessel

Stimulus dependence is a general feature of developing sensory systems. It has been shown earlier that the growth of inner ear heavy stones (otoliths) of late-stage Cichlid fish (Oreochromis mossambicus) and Zebrafish (Danio rerio) is slowed down by hypergravity, whereas microgravity during space flight yields an opposite effect, i.e. larger than 1 g otoliths, in Swordtail (Xiphophorus helleri) and in Cichlid fish late-stage embryos. These and related studies proposed that otolith growth is actively adjusted via a feedback mechanism to produce a test mass of the appropriate physical capacity. Using ground-based techniques to apply simulated weightlessness, long-term clinorotation (CR; exposure on a fast-rotating Clinostat with one axis of rotation) led to larger than 1 g otoliths in late-stage Cichlid fish. Larger than normal otoliths were also found in early-staged Zebrafish embryos after short-term Wall Vessel Rotation (WVR; also regarded as a method to simulate weightlessness). These results are basically in line with the results obtained on Swordtails from space flight.     

Otoliths as biomechanical gravisensors.

This paper analyzes experimental data related to the reaction of otolith afferents in response to acceleration (Fernandez and Goldberg, 1976). It considers the assumptions that were the basis of the interpretation of the stimulus-response characteristics of afferents proposed by Fernandez and Goldberg. Comparing these experimental data with the results of modeling the otolith structures of vertebrates indicates that some peculiarities of the neural responses may be explained by the spatial dependence of the material parameters of the otolithic membrane across its thickness and within the volume of the membrane corresponding to the terminal field. The importance of the spatial dependence of the material parameters of the otolithic membrane for otolith functioning is discussed.     

Calcium gradients in the fish inner ear sensory epithelium and otolithic membrane visualized by energy filtering transmission electron microscopy (EFTEM).

Inner ear otolith formation in fish is supposed to be performed by the molecular release of proteinacious precursor material from the sensory epithelia, followed by an undirected and diffuse precipitation of calcium carbonate (which is mainly responsible for the functionally important weight of otoliths). The pathway of calcium into the endolymph, however, still remains obscure. Therefore, the presence of calcium within the utricle of larval cichlid fish Oreochromis mossambicus was analyzed by means of energy filtering transmission electron microscopy (EFTEM). Electron spectroscopic imaging (ESI) and electron energy loss spectra (EELS) revealed discrete calcium precipitations, which were especially numerous in the proximal endolymph as compared to the distal endolymph. A decreasing proximo-distal gradient was also present within the proximal endolymph between the sensory epithelium and the otolith. Further calcium particles covered the peripheral proteinacious layer of the otolith. They were especially pronounced at the proximal surface of the otolith. Other calcium precipitates were found to be accumulated at the macular junctions. These results strongly suggest that the apical region of the macular epithelium is involved in the release of calcium and that calcium supply of the otoliths takes place in the proximal endolymph.     

Identification of specific gravity sensitive signal transduction pathways in human A431 carcinoma cells.

Epidermal growth factor (EGF) activates a well characterized signal transduction cascade in human A431 epidermoid carcinoma cells. The influence of gravity on EGF-induced EGF-receptor clustering and early gene expression as well as on actin polymerization and actin organization have been investigated. Different signalling pathways induced by the agents TPA, forskolin and A23187 that activate gene expression were tested for sensitivity to gravity. EGF-induced c-fos and c-jun expression were decreased in microgravity. However, constitutive beta-2 microglobulin expression remained unaltered. Under simulated weightlessness conditions EGF- and TPA-induced c-fos expression was decreased, while forskolin- and A23187-induced c-fos expression was independent of the gravity conditions. These results suggest that gravity affects specific signalling pathways. Preliminary results indicate the EGF-induced EGF-receptor clustering remained unaltered irrespective of the gravity conditions. Furthermore, the relative filamentous actin content of steady state A431 cells was enhanced under microgravity conditions and actin filament organization was altered. Under simulated weightlessness actin filament organization in steady state cells as well as in EGF-treated cells was altered as compared to the 1 G reference experiment. Interestingly the microtubule and keratin organization in untreated cells showed no difference with the normal gravity samples. This indicates that gravity may affect specific components of the signal transduction circuitry.     

A drop-tower experiment to determine the threshold of gravity for inducing motion sickness in fish.

It has been repeatedly shown earlier that some fish of a given batch reveal motion sickness (a kinetosis) at the transition from 1 g to microgravity. In the course of parabolic aircraft flight experiments, it has been demonstrated that kinetosis susceptibility is correlated with asymmetric inner ear otoliths (i.e., differently weighed statoliths on the right and the left side of the head) or with genetically predispositioned malformed cells within the sensory epithelia of the inner ear. Hitherto, the threshold of gravity perception for inducing kinetotic behavior as well as the relative importance of asymmetric otoliths versus malformed epithelia for kinetosis susceptibility has yet not been determined. The following experiment using the ZARM drop-tower facility in Bremen, Germany, is proposed to be carried out in order to answer the aforementioned questions. Larval cichlid fish (Oreochromis mossambicus) will be kept in a camcorder-equipped centrifuge during the microgravity phases of the drops and thus receive various gravity environments ranging from 0.1 to 0.9 g. Videographed controls will be housed outside of the centrifuge receiving 0 g. Based on the video-recordings, animals will be grouped into kinetotically and normally swimming samples. Subsequently, otoliths will be dissected and their size and asymmetry will be measured. Further investigations will focus on the numerical quantification of inner ear supporting and sensory cells as well as on the quantification of inner ear carbonic anhydrase reactivity. A correlation between: (1) the results to be obtained concerning the g-loads inducing kinetosis and (2) the corresponding otolith asymmetry/morphology of sensory epithelia/carbonic anhydrase reactivity will further contribute to the understanding of the origin of kinetosis susceptibility. Besides an outline of the proposed principal experiments, the present study reports on a first series of drop-tower tests, which were undertaken to elucidate the feasibility of the proposal (especially concerning the question, if some 4.7 s of microgravity are sufficient to induce kinetotic behavior in larval fish).     

Neuroplastic reactivity of fish induced by altered gravity conditions: a review of recent results.

A review is being presented concerning behavioural, biochemical, histochemical and electronmicroscopical data on the influence of altered gravitational forces on the swimming performance and on the neuronal differentiation of the brain of cichlid fish larvae and adult swordtail fish that had been exposed to hyper-gravity (3g in laboratory centrifuges), hypo-gravity (>10(-2) g in a fast-rotating clinostat) and to near weightlessness (10(-4) g aboard the Spacelab D-2 mission). After long-term alterations of gravity (and parallel light deprivation), initial disturbances in the swimming behaviour followed by a stepwise regain of normal swimming modes are induced. Parallel, neuroplastic reactivities on different levels of investigation were found, such as adaptive alterations of activities of various enzymes in whole brain as well as in specific neuronal integration centers and an intraneuronal reactivity on ultrastructural level in individual brain parts and in the sensory epithelia of the inner ear. Taken together, these data reveal distinct adaptive neuroplastic reactions of fish to altered gravity conditions.     

基于Lagrange方法的失重人体建模与仿真

阐述了利用多刚体系统动力学进行航天员舱外活动仿真EVA(Extra Vehicular Activity)的必要性.给出了应用计算多刚体系统动力学建立的通用失重人体4关节反向运动学与反向动力学模型.选取典型的实例,在对其进行适当简化的基础上,运用通用模型对其进行仿真计算,计算时为考虑失重对人体质量、惯量与力量等参数的影响,对通用模型进行了修正.利用能量比较法对结果进行分析,得出当手部的运动轨迹半径与角速度减小、时间延长,髋关节做前驱运动时航天员工作最为节省能量.通过能量比较法计算得到了人体运动时各关节作功最为节省能量的范围.计算方法对航天员舱内外活动仿真及工效分析有一定的参考价值.      

Efficacy of an ototoxic aminoglycoside (gentamicin) on the differentiation of the inner ear of cichlid fish.

Previous investigations revealed that the growth of fish inner ear otoliths depends on the amplitude and the direction of gravity, thus suggesting the existence of a (negative) feedback mechanism. In the course of these experiments, it was shown that altered gravity both affected otolith size (and thus the provision of the proteinacious matrix) as well as the incorporation of calcium. It is hitherto unknown, as of whether sensory hair cells are involved either in the regulation of otolith growth or in the provision of otolithic material (such as protein or inorganic components) or even both. The ototoxic aminoglycoside gentamicin (GM) damages hair cells in many vertebrates (and is therefore used for the treatment of Meniere's disease in humans). The present study was thus designed to determine as of whether vestibular sensory cells are needed for otolith growth by applying GM in order to induce a (functionally relevant) loss of these cells. Developing cichlid fish Oreochromis mossambicus were therefore immersed in 120 mg/l GM for 10 or 21 days. At the beginning and at the end of the experimental periods, the fish were incubated in the calcium-tracer alizarin complexone (AC). After the experiment, otoliths were dissected and the area grown during GM-exposure (i.e., the area enclosed by the two AC labellings) was determined planimetrically. The results showed that incubating the animals in a GM-solution had no effect on otolith growth, but the development of otolith asymmetry was affected. Ultrastructural examinations of the sensory hair cells revealed that they had obviously not been affected by GM-treatment (no degenerative morphological features observed). Overall, the present results suggest that hair cells are not affected by GM concerning their possible role in (general) otolith growth, but that these cells indeed might have transitionally been impaired by GM resulting in a decreased capacity of regulating otolith symmetry.     

Clinorotation reduces number, but not size, of cartilaginous nodules formed in micromass cultures of mouse limbbud cells.

In previous studies we used a ground based model to investigate the cellular responses to microgravity by exposing micromass cultures of embryonic limb cells to simulated weightlessness on a clinostat. Cultures set up in T-flasks and rotated at 30 rpm showed that clinostatted cultures had less chondrocyte differentiation than stationary or rotation controls, as assessed by number of nodules/culture stained with cartilage specific Alcian blue. In the current study, nodule size and shape of these nodules was assessed by interactive measurement of area, perimeter, circularity, and equivalent diameters, using the Optimas imaging software. Results show no significant difference in any of the measurements, indicating that clinorotation has no effect on expansion of the nodules either by differentiation of cells within the nodule, or by recruitment of cells into the nodule. The reduction in number of nodules without an alteration in size and shape indicates that the effect of simulated microgravity is to reduce the cell interactions required for the initial condensation of cells into a nodule, probably by interference with cell adhesion molecules.     

Weightlessness acts on human breast cancer cell line MCF-7.

Because cells are sensitive to mechanical forces, weightlessness might act on stress-dependent cell changes. Human breast cancer cells MCF-7, flown in space in a Photon capsule, were fixed after 1.5, 22 and 48 h in orbit. Cells subjected to weightlessness were compared to 1 g in-flight and ground controls. Post-flight, fluorescent labeling was performed to visualize cell proliferation (Ki-67), three cytoskeleton components and chromatin structure. Confocal microscopy and image analysis were used to quantify cycling cells and mitosis, modifications of the cytokeratin network and chromatin structure. Several main phenomena were observed in weightlessness: The perinuclear cytokeratin network and chromatin structure were looser; More cells were cycling and mitosis was prolonged. Finally, cell proliferation was reduced as a consequence of a cell-cycle blockade; Microtubules were altered in many cells. The results reported in the first point are in agreement with basic predictions of cellular tensegrity. The prolongation of mitosis can be explained by an alteration of microtubules. We discuss here the different mechanisms involved in weightlessness alteration of microtubules: i) alteration of their self-organization by reaction-diffusion processes, and a mathematical model is proposed, ii) activation or deactivation of microtubules stabilizing proteins, acting on both microtubule and microfilament networks in cell cortex.     

Gravitaxis and graviperception in Euglena gracilis.

Gravitactic orientation in the flagellate Euglena gracilis is mediated by an active physiological receptor rather than a passive alignment of the cells. During a recent space flight on the American shuttle Columbia the cells were subjected to different accelerations between 0 and 1.5 x g and tracked by computerized real-time image analysis. The dependence of orientation on acceleration followed a sigmoidal curve with a threshold at < or = 0.16 x g and a saturation at about 0.32 x g. No adaptation of the cells to the conditions of weightlessness was observed over the duration of the space mission (12 days). Under terrestrial conditions graviorientation was eliminated when the cells were suspended in a medium the density of which (Ficoll) equaled that of the cell body (1.04 g/ml) and was reversed at higher densities indicating that the whole cytoplasm exerts a pressure on the respective lower membrane. There it probably activates stretch-sensitive calcium specific ion channels since gravitaxis can be affected by gadolinium which is a specific inhibitor of calcium transport in these structures. The sensory transduction chain could involve modulation of the membrane potential since ion channel blockers, ionophores and ATPase inhibitors impair graviperception.     

失重因素对航天员体温调节影响的分析

系统地总结了实际载人航天及地面模拟中失重因素引起的航天员生理性体温调节功能变化的种种现象和内在联系。指出在失重环境下 ,自然对流消失 ,血液重新分布 ,排尿性失水增加 ,血浆容积变小 ,心血管系统功能下降 ,最大氧摄入量降低 ,这些都可导致体温调节能力受损 ,最终使航天员的高温耐力明显下降。文章同时指出 ,在体温调节系统中 ,行为性调节是生理性调节的补充和延伸 ,前者只有通过后者才能发挥作用 ,因此 ,在航天器座舱温控系统及航天服通风 -液冷系统的设计中 ,应充分考虑失重因素的影响。     

模拟失重时家兔耳廓微循环的变化

为了解失重时体液头向分布这一因素对微循环的影响,比较了两组兔(平卧组和头低位20°悬吊组)在实验7天中耳廓微循环和血液流变学指标的变化。结果证明体液头向分布可引起一定的微循环障碍,表现为毛细血管和细静脉管径变粗、流速减慢、血液在静脉中潴留、红细胞聚集、耳廓出血及立位时微血管调节功能发生改变,血液流变学指标也出现相应变化。本文分析了其中一些变化的起因,提出一些新的看法。      

抗磁性物质磁悬浮方法在空间生物学与生物技术中的应用

失重是特定空间运动条件下的重要环境物理特征之一, 一般以微重力环境来表示. 几十年来人类利用空间失重环境进行了多学科领域的科学研究与探索. 由于真实空间失重环境下科学实验机会稀少, 人类为研究空间失重环境或效应, 开发了多种地基的空间模拟实验技术方法. 然而, 对于空间生物学和空间生物技术研究而言, 已有的各种模拟实验技术手段在原理上和应用上均存在一定的局限性. 本文介绍了抗磁性物质在大梯度强磁场中的悬浮现象, 及将其用于模拟空间失重环境的方法与原理;简述了近年来利用抗磁性物质悬浮方法进行生物大分子晶体生长、分子细胞生物学及整体生物学等方面研究与应用的进展.      

Microgravity effects on neural retina regeneration in the newt.

Data on forelimb and eye lens regeneration in urodeles under spaceflight conditions (SFC) have been obtained in our previous studies. Today, evidence is available that SFC stimulate regeneration in experimental animals rather than inhibit it. The results of control on-ground experiments with simulated microgravity suggest that the stimulatory effect of SFC is due largely to weightlessness. An original experimental model is proposed, which is convenient for comprehensively analyzing neural regeneration under SFC. The initial results described here concern regeneration of neural retina in Pleurodeles waltl newts exposed to microgravity simulated in radial clinostat. After clinorotation for seven days (until postoperation day 16), a positive effect of altered gravity on structural restoration of detached neural retina was confirmed by a number of criteria. Specifically, an increased number of Mullerian glial cells, an increased relative volume of the plexiform layers, reduced cell death, advanced redifferentiation of retinal pigment epithelium, and extended areas of neural retina reattachment were detected in experimental newts. Moreover, cell proliferation in the inner nuclear layer of neural retina increased as compared with control. Thus, low gravity appears to intensify natural cytological and molecular mechanisms of neural retina regeneration in lower vertebrates.