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The amphibian egg as a model system for analyzing gravity effects.

Amphibian eggs provide several advantageous features as a model system for analyzing the effects of gravity on single cells. Those features include large size, readily tracked intracellular inclusions, and ease of experimental manipulation. Employing novel gravity orientation as a tool, a substantial data base is being developed. That information is being used to construct a 3-D model of the frog (Xenopus laevis) egg. Internal cytoplasmic organization (rather than surface features) are being emphasized. Several cytoplasmic compartments (domains) have been elucidated, and their behavior in inverted eggs monitored. They have been incorporated into the model, and serve as a point of departure for further inquiry and speculation.

Animals↗

Biological role of gravity: hypotheses and results of experiments on "Cosmos" biosatellites.

In order to reveal the biological significance of gravity, microgravity effects have been studied at the cellular, organism and population levels. The following questions arise. Do any gravity-dependent processes exist in a cell? Is cell adaptation to weightlessness possible; if so, what role may cytoskeleton, the genetic apparatus play in it? What are the consequences of the lack of convection in weightlessness for the performance of morphogenesis? Do the integral characteristics of living beings change in weightlessness? Is there any change in "biological capacity" of space, its resistance to expansion of life? What are the direction and intensity of microgravity action as a factor of natural selection, the driving force of evolution? These problems are discussed from a theoretical point of view, and in the light of results obtained in experiments from aboard biosatellites "Cosmos".

Adaptation, Physiological↗

Gravity effects on biological systems.

Theoretical investigations of the membrane-solution interface predict different effects of gravity on vertically and horizontally oriented planar membranes. Single channel events of gramicidin incorporated into phosphatidylserine planar bilayer membranes were measured in 0.1 M KCl solution, pH 7, at room temperature. The potential difference across the membrane was set to +/- 70 mV. The mean channel current was observed to be about 20% higher in horizontally oriented membranes compared to vertical membranes. This is in good agreement with the theoretical considerations and demonstrates that gravity does affect membrane processes by interaction with the membrane-solution interface which is a ubiquitous structure in biological systems.

Animals↗

Human postural reflexes and gravity--an under water simulation.

This study represents the first attempt to investigate the influence of gravity on postural adjustments. Subjects were displaced while standing under water on a movable platform, while the buoyancy of the body was adjusted by using a variety of lead vests. Under water, an approximately linear relationship was found between body weight and impulse directed electromyographic response amplitudes in the leg and thigh muscles. Loading of the subjects out of water resulted in a saturation of the response amplitude. The biomechanical signals recorded during the displacements indicated that neither vestibulospinal nor muscle proprioceptive reflex mechanisms can account for the effect observed under water. It is suggested that the EMG responses are mediated by reflexes which are activated by pressure receptors within the body in order to hold the centre of gravity over the feet.

Adult↗

Gravity, weightlessness and the genetic structures of organisms.

The whole evolution of life on earth has proceeded under the action of earth's gravity which must have influenced the structure and function of organisms. During space flights organisms are exposed to the entirely new condition of weightlessness, and to variations in gravity that produce various changes. Current flight data suggest that organisms most often respond to weightlessness by disturbances in physiological function, which are reversible after the brief exposures that have at present been possible. Only longer space flights will show whether these changes will be compensated, or will lead to alterations in the more important systems of organisms including their hereditary properties. There is evidence that weightlessness has a direct effect on genetic properties. Thus, in microspores of Tradescantia abnormal mitoses (that were not observed in the ground-based control) were recorded to an extent of 3%. The numerous changes in various vital systems suggest that for terrestrial organisms weightlessness is a factor which, if administered for a long time, may bring about serious disturbances in their activities and heredity. The higher the evolutionary position of the organism, the more pronounced the effect is likely to be.

Adaptation, Physiological↗

Gravity as a biochemical determinant.

The existence of obvious morphological and physiological changes in living systems exposed to altered gravity immediately informs us that prior changes have taken place in the chemistry of exposed cells, tissues and organs. These changes include transients that return more or less promptly to the norm when the system is restored to the terrestrial g-field. For example, altered serum hormone and electrolyte levels in man, which appear to reflect successful adaptation to the conditions of orbital weightlessness, disappear shortly after return to Earth. Other changes--in mineral and protein constituents of the skeletal system in man, and cell wall composition in plants--are more persistent or even permanent. Hypogravitational departures from the norm include not only "weightlessness" as achieved in orbit, but also experimental modes of compensation, on the clinostat or by flotation. These techniques are useful in the study of hypogravity but cannot replace fully the weightless environment. Plant ethylene and peroxidase both increase under orbital, clinostat and/or flotation conditions whereas 3-phosphoglyceraldehyde-dehydrogenase increases under orbital but not clinostat conditions; cytochrome reductase and malic dehydrogenase levels are affected by the clinostat, but not by actual weightless conditions. How do the altered organismal biochemistries induced by the centrifuge and the clinostat relate to one another? Does gravity operate on living systems as a continuous variable from 0 to superterrestrial values, or do deviations from g(earth) generate non-uniform, discontinuous stress responses, irrespective of sign? In plants, measurements of wall lignin content and peroxidase activity yield opposite answers. Given the limited data so far available we will consider the meaning of these contradictions.

Adaptation, Physiological↗

Is gravity a morphological determinant in plants at the cellular level?

It is not yet known whether higher plants, which have evolved in a gravity oriented environment, can reproduce normally or develop via single cells (somatic or zygotic) and pass normally through all phases of development, tissue differentiation and reproduction through consecutive generations at near-zero g. Most published work deals with directional gravitational stimuli (geotropisms) as they operate on the subsequent growth of preformed plant organs that have acquired the ability to perceive and respond to gravity during development at 1 g. But it should be established whether plant development can proceed normally in the weightless state, especially that critical stage in which single cells produce multicellular units leading to embryos with the growing regions of shoot and root which, in turn, give rise to all the tissues of the plant body. We consider here an experiment that tested whether carrot embryos, capable of developing from cultured somatic cells, could do so under conditions of near-weightlessness. Space flight seemed to have no adverse affects on the induction of such embryos or on the initiation of their organs. But there are still reservations which are discussed. Other systems adaptable to the study of embryogenesis and development in space and to resolve the current reservations are described and their place in future space research is indicated.

Arabidopsis↗

Loading effects on rat craniomandibular morphology: a system for gravity studies.

Gravity effects on muscle and bone are a major impediment to long-term space travel. We introduce a model for studying these effects, the craniomandibular system. Some advantages of this system include: (1) craniomandibular morphology is determined by epigenetic factors including gravity, (2) relatively light forces can significantly alter its morphology, and (3) soft diet and tooth loss produce effects that are similar to those produced in lower limbs by weightlessness. In the study, implants made either of gold (experimental group) or lightweight acrylic (controls) were attached to adult rats' mandibles. After 13 weeks, the animals' skulls and mandibles were dissected. Pair-wise comparisons indicated that the experimental animals showed significantly shortened and narrowed cranial bases, and significant changes in the posterior zygomatic arch region. These results indicate that simulated macrogravity influences bone remodeling in the adult craniomandibular system.

Acrylic Resins↗

Cell-to-cell interactions in changed gravity: ground-based and flight experiments.

Cell-to-cell interactions play an important role in all physiological processes and are mediated by humoral and mechanical factors. Mechanosensitive cells (e.g., osteocytes, chondrocytes, and fibroblasts) can be studied ex vivo to understand the effects of an altered gravity environment. In particular, cultured endothelial cells (EC) are very sensitive to a broad spectrum of mechanical and biochemical stimuli. Earlier, we demonstrated that clinorotation leads to cytoskeletal remodeling in cultured ECs. Long-term gravity vector changes also modulate the expression of surface adhesion molecules (ICAM-1, E-selectin, VCAM-1) on cultured ECs. To study the interactions of geterological cells, we cocultured endothelial monolayers and human lymphocytes, immune cells and myeloleucemic (K-560) cells. It was found that, although clinorotation did not alter the basal adhesion level of non-activated immune cells on endothelial monolayers, the adhesion of PMA-activated lymphocytes was increased. During flight experiments onboard the Russian segment of the International Space Station, we measured the cytotoxic activity of natural killer (NK) cells incubated with labeled target cells. It was found that immune cells in microgravity retained their ability to contact, recognize, and destroy oncogenic cells in vitro. Together, our data concerning the effects of simulated and real microgravity suggest that, despite changes in the cytoskeleton, cell motility, and expression of adhesion molecules, cell-cell interactions are not compromised, thus preserving the critical physiological functions of immune and endothelial cells.

Cell Adhesion↗

CNS development under altered gravity: cerebellar glial and neuronal protein expression in rat neonates exposed to hypergravity.

The future of space exploration depends on a solid understanding of the developmental process under microgravity, specifically in relation to the central nervous system (CNS). We have previously employed a hypergravity paradigm to assess the impact of altered gravity on the developing rat cerebellum. The present study addresses the molecular mechanisms involved in the cerebellar response to hypergravity. Specifically, the study focuses on the expression of selected glial and neuronal cerebellar proteins in rat neonates exposed to hypergravity (1.5 G) from embryonic day (E)11 to postnatal day (P)6 or P9 (the time of maximal cerebellar changes) comparing them against their expression in rat neonates developing under normal gravity. Proteins were analyzed by quantitative Western blots of cerebellar homogenates; RNA analysis was performed in the same samples using quantitative PCR. Densitometric analysis of Western blots suggested a reduction in glial (glial acidic protein, GFAP) and neuronal (neuronal cell adhesion molecule, NCAM-L1, synaptophysin) proteins, but the changes in individual cerebellar proteins in hypergravity-exposed neonates appeared both age- and gender-specific. RNA analysis suggested a reduction in GFAP and synaptophysin mRNAs on P6. These data suggest that exposure to hypergravity may interfere with the expression of selected cerebellar proteins. These changes in protein expression may be involved in mediating the effect of hypergravity on the developing rat cerebellum.

Animals↗

Gravity spun polycaprolactone fibres: controlling release of a hydrophilic macromolecule (ovalbumin) and a lipophilic drug (progesterone).

A hydrophilic macromolecule (ovalbumin (OVA)) and a lipophilic drug (progesterone) were incorporated in polycaprolactone (PCL) fibres by gravity spinning using particulate dispersions and co-solutions of PCL and steroid, respectively. PCL fibres loaded with 1% (w/w) OVA powder displayed a pronounced burst release phase (60% of the protein load) over 2 days in PBS at 37 degrees C. The release profile then tended to plateau. In contrast, OVA nanoparticle-loaded fibres exhibited delayed protein release initially and then a major increase at day 14. This behaviour may be useful for sequential release of polypeptide growth factors which are influential at specific time points in the wound healing process. SDS-PAGE analysis revealed that the protein molecular weight was conserved during fibre spinning. The amount of progesterone release from PCL fibres in PBS increased with drug loading but the cumulative release profiles (% w/w) were little affected by the initial drug loading of the fibres (1.5 and 3.5% w/w) or the concentration of the PCL spinning solution (12.5 and 20% w/v). Steroid delivery was rapid due to the high fibre surface area and high permeability of PCL resulting in complete drug loss over 24h. Released progesterone inhibited the growth of MCF-7 breast epithelial cells in culture, demonstrating retention of bioactivity. Gravity spinning shows potential for producing PCL fibre-based platforms for programmed delivery of bioactive molecules of utility for tissue engineering and drug delivery.

Biocompatible Materials↗

Gravity spun polycaprolactone fibres for soft tissue engineering: interaction with fibroblasts and myoblasts in cell culture.

Poly(epsilon-caprolactone) (PCL) fibres were produced by wet spinning from solutions in acetone under low shear (gravity flow) conditions. As-spun PCL fibres exhibited a mean strength and stiffness of 7.9 MPa and 0.1 GPa, respectively and a rough, porous surface morphology. Cold drawing to an extension of 500% resulted in increases in fibre strength (43 MPa) and stiffness (0.3 GPa) and development of an oriented, fibrillar surface texture. The proliferation rate of Swiss 3T3 mouse fibroblasts and C2C12 mouse myoblasts on as-spun, 500% cold-drawn and gelatin-modified PCL fibres was determined in cell culture to provide a basic measure of the biocompatibility of the fibres. Proliferation of both cell types was consistently higher on gelatin-coated fibres relative to as-spun fibres at time points below 7 days. Fibroblast growth rates on cold-drawn PCL fibres exceeded those on as-spun fibres but myoblast proliferation was similar on both substrates. After 1 day in culture, both cell types had spread and coalesced on the fibres to form a cell layer, which conformed closely to the underlying topography. The high fibre compliance combined with a potential for modifying the fibre surface chemistry with cell adhesion molecules and the surface architecture by cold drawing to enhance proliferation of fibroblasts and myoblasts, recommends further investigation of gravity-spun PCL fibres for 3-D scaffold production in soft tissue engineering.

3T3 Cells↗

Effect of gravity on human spontaneous 10-Hz electroencephalographic oscillations during the arrest reaction.

Electroencephalographic oscillations at 10 Hz (alpha and mu rhythms) are the most prominent rhythms observed in awake, relaxed (eye-closed) subjects. These oscillations may be considered as a marker of cortical inactivity or an index of the active inhibition of the sensory information. Different cortical sources may participate in the 10-Hz oscillation and appear to be modulated by the sensory context and functional demands. In microgravity, the marked reduction in multimodal graviceptive inputs to cortical networks participating in the representation of space could be expected to affect the 10-Hz activity. The effect of microgravity on this basic oscillation has heretofore not been studied quantitatively. Because the alpha rhythm has a functional role in the regulation of network properties of the visual areas, we hypothesised that the absence of gravity would affect its strength. Here, we report the results of an experiment conducted over the course of 3 space flights, in which we quantified the power of the 10-Hz activity in relation to the arrest reaction (i.e., in 2 distinct physiological states: eyes open and eyes closed). We observed that the power of the spontaneous 10-Hz oscillation recorded in the eyes-closed state in the parieto-occipital (alpha rhythm) and sensorimotor areas (mu rhythm) increased in the absence of gravity. The suppression coefficient during the arrest reaction and the related spectral perturbations produced by eye-opening/closure state transition also increased in on orbit. These results are discussed in terms of current theories on the source and the importance of the alpha rhythm for cognitive function.

Adult↗

Feasibility of preparing nanodrugs by high-gravity reactive precipitation.

To study the feasibility of producing nanoparticles of organic pharmaceuticals using a novel high-gravity reactive precipitation (HGRP) technique, reactive precipitation of benzoic acid as a model compound was carried out in a rotating packed bed under high gravity. The main factors such as the rotating bed speed, concentration and volume flow rate of the reactants (sodium benzoate and HCl) affecting the particle size of the precipitate were studied. Particle size was measured by transmission electron microscopy. Benzoic acid was precipitated as nanoparticles as fine as 10nm. The particle size was decreased with increasing rotating bed speed, concentration and volume flow rate of the reactants. The formation of ultrafine particles was due to intensified micro-mixing of reactants in the rotating bed to enhance nucleation while suppressing crystal growth. The results have demonstrated the feasibility to produce nanodrugs by the principle of acid-base precipitating reaction using HGRP.

Benzoic Acid↗

The critical gas saturation in a porous medium in the presence of gravity.

The critical gas saturation, S(gc), denotes the volume fraction of the gas phase at the onset of bulk gas flow during the depressurization of a supersaturated liquid in a porous medium. In the absence of gradients due to viscous or gravity forces, S(gc) is controlled by nucleation, capillary forces, and the rate of decline of the supersaturation. In this paper we address one important additional effect, that of buoyancy. We use 2-D pore-network simulations, based on invasion percolation in a gradient (IPG), and corresponding scaling relations to obtain the dependence of S(gc) on the gravity Bond number, B, under conditions of slow growth, namely when mass transfer is sufficiently fast. The critical gas saturation approaches two plateau values at low and high Bond numbers. In the in-between region it scales as a power law of B, which for a 2-D lattice is S(gc) approximately B(-0.91).

Journal Article↗

Viscous-gravity spreading of time-varying liquid drop volumes on solid surfaces.

Viscous-gravity spreading of liquid drops of time-dependent volume over a solid surface is considered. A self-similar solution for the drop configuration is obtained, in the case the liquid drop volume varies as a power-law function of time, along with the spreading laws in both cases of cylindrical and axisymmetric geometries. Results compare favorably with published experimental results and previous theoretical work. The limitations of the model are discussed, along with a comparison with viscous gravity spreading of oil on water. The validity of using approximate spreading laws is considered, and an approximate method is suggested to provide the dynamics of spreading in the general case where the drop volume does not necessarily vary as a power-law function of time.

Journal Article↗

Defects in root development and gravity response in the aem1 mutant of rice are associated with reduced auxin efflux.

The phytohormone auxin is involved in the regulation of a variety of developmental processes. In this report, we describe how the processes of lateral root and root hair formations and root gravity response in rice are controlled by auxin. We use a rice mutant aem1 (auxin efflux mutant) because the mutant is defective in these characters. The aem1 line was originally isolated as a short lateral root mutant, but we found that the mutant has a defect in auxin efflux in roots. The acropetal and basipetal indole-3-acetic acid (IAA) transports were reduced in aem1 roots compared to wild type (WT). Furthermore, gravitropic bending as well as efflux of radioactive IAA was impaired in the mutant roots. We also propose a unique distribution of endogenous IAA in aem1 roots. An immunoassay revealed a 4-fold-endogenous IAA content in the aem1 roots compared to WT, and the application of IAA to the shoot of WT seedlings mimicked the short lateral root phenotype of aem1, suggesting that the high content of IAA in aem1 roots impaired the elongation of lateral roots. However, the high level of IAA in aem1 roots contradicts the auxin requirement for root hair formation in the epidermis of mutant roots. Since the reduced development in root hairs of aem1 roots was rescued by exogenous auxin, the auxin level in the epidermis is likely to be sub-optimum in aem1 roots. This discrepancy can be solved by the ideas that IAA level is higher in the stele and lower in the epidermis of aem1 roots compared to WT and that the unique distribution of IAA in aem1 roots is induced by the defect in auxin efflux. All these results suggest that AEM1 may encode a component of auxin efflux carrier in rice and that the defects in lateral roots, root hair formation and root gravity response in aem1 mutant are due to the altered auxin efflux in roots.

Biological Transport↗

Modelling of auxin transport affected by gravity and differential radial growth.

When a tree stem deviates from verticality, as a result of different environmental factors, patterns of differential radial growth appear. Higher rates of wood production have been observed on the lower side of the tree and lower rates in the opposite side. Biological studies on plant hormones have shown that the concentration of auxin induces radial growth. They also have demonstrated the redistribution of auxin transport in response to gravity. Auxin is then designated as a mediator for differential growth. This paper presents a model for three-dimensional (3-D) auxin transport in conifer trees, which includes gravity dependence. We obtain realistic heterogeneous patterns of auxin distribution over the tree. Then, we propose a law of growth based on auxin concentration to simulate successive differential radial growths. The predicted growths are compared with experimental results of reconstruction of 3-D annual growth of Radiata pine.

Biological Transport↗