Effects of hypergravity on the elongation growth in radish and cucumber hypocotyls

The elongation growth of the hypocotyls of radish and cucumber seedlings was examined under hypergravity in a newly developed centrifuge (Kasahara et al. 1995). The effects of hypergravity on elongation growth differed between the two species. The rate of elongation of radish hypocotyls was reduced under basipetal hypergravity (H+2O g) but not under acropetal hypergravity (H-13 g), as compared to growth under the control conditions (C+1 g and C-1 g). In cucumber hypocotyls, elongation growth was inhibited not only by basipetal but also by acropetal hypergravity. Under these conditions, the reduction in the elongation growth of both radish and cucumber hypocotyls was accompanied by an increase in their thickness. Although no distinct differences in relative composition of neutral sugars were found, the amounts of cell-wall components (pectic substances, hemicelluloses and cellulose) per unit length of hypocotyls were increased by exposure to hypergravity.     

Effects of hypergravity on the elongation growth in radish and cucumber hypocotyls

The elongation growth of the hypocotyls of radish and cucumber seedlings was examined under hypergravity in a newly developed centrifuge (Kasaharaet al. 1995). The effects of hypergravity on elongation growth differed between the two species. The rate of elongation of radish hypocotyls was reduced under basipetal hypergravity (H+20g) but not under acropetal hypergravity (H-13g), as compared to growth under the control conditions (C+1g and C-1g). In cucumber hypocotyls, elongation growth was inhibited not only by basipetal but also by acropetal hypergravity. Under these conditions, the reduction in the elongation growth of both radish and cucumber hypocotyls was accompanied by an increase in their thickness. Although no distinct differences in relative composition of neutral sugars were found, the amounts of cell-wall components (pectic substances, hemicelluloses and cellulose) per unit length of hypocotyls were increased by exposure to hypergravity.     

Effects of hypergravity on the elongation and morphology of protonemata of Adiantum capillus-veneris

Elongation growth of protonemata of Adiantum capillus-veneris , which can be controlled by light irradiation, was examined under acropetal and basipetal hypergravity conditions (from -13 to +20 g ) using a newly developed centrifugation equipment. Elongation of the protonemata under red light was inhibited by basipetal hypergravity at more than +15 g but was promoted by acropetal hypergravity from -5 to -8 g . Division of the protonemal cells that was induced by white light was inhibited under basipetal hypergravity at +20 g but was unaffected under acropetal hypergravity at -15 g . Upon exposure to continuous red light for 7 to 8 days, most of the protonemata grew as filamentous cells in the absence of a change in the normal gravitational force (control), but more than half of the protonemal cells were abnormal in terms of shape when maintained under hypergravity at +20 g .     

Responses to hypergravity in proliferation of Paramecium tetraurelia

It has been reported that Paramecium proliferates faster when cultured under microgravity in orbit, and slower when cultured under hypergravity. This shows that the proliferation rate of Paramecium affected by gravity. The effect of gravity on Paramecium proliferation has been argued to be direct in a paper with an axenic culture under hypergravity. To clear up uncertainties with regard to the effect of gravity, Paramecium tetraurelia was cultured axenically under hypergravity (20 x g) and the time course of the proliferation was investigated quantitatively by a new non-invasive method, laser-beam optical slice, for measuring the cell density. This method includes optical slicing a part of the culture and computer-aided counting of cells in the sliced volume. The effects of hypergravity were assessed by comparing the kinetic parameters of proliferation that were obtained through a numerical analysis based on the logistic growth equation. Cells grown under 20 x g conditions had a significantly lower proliferation rate, and had a lower population density at the stationary phase. The lowered proliferation rate continued as long as cells were exposed to hypergravity (> one month). Hypergravity reduced the cell size of Paramecium. The long and short axes of the cell became shorter at 20 x g than those of control cells, which indicates a decrease in volume of the cell grown under hypergravity and is consistent with the reported increase in cell volume under microgravity. The reduced proliferation rate implies changes in biological time defined by fission age. In fact the length of autogamy immaturity decreased by measure of clock time, whereas it remained unchanged by measure of fission age.     

Hypergravity increases the molecular mass of xyloglucans by decreasing xyloglucan-degrading activity in azuki bean epicotyls.

Elongation growth of dark-grown azuki bean (Vigna angularis Ohwi et Ohashi cv. Takara) epicotyls was suppressed by hypergravity at 30 x g and above. Acceleration at 300 x g significantly decreased the mechanical extensibility of cell walls. The amounts of cell wall polysaccharides (pectin, hemicellulose-II and cellulose) per unit length of epicotyls increased under the hypergravity condition. Hypergravity also increased the amounts and the weight-average molecular mass of xyloglucans in the hemicellulose-II fraction, while decreasing the activity of xyloglucan-degrading enzymes extracted from epicotyl cell walls. These results suggest that hypergravity increases the amounts and the molecular mass of xyloglucans by decreasing xyloglucan-degrading activity. Modification of xyloglucan metabolism as well as the thickening of cell walls under hypergravity conditions seems to be involved in making the cell wall mechanically rigid, thereby inhibiting elongation growth of azuki bean epicotyls.     

Effects of hypergravity environment on lignin formation in Arabidopsis.

Lignin and the secondary wall formation are essential for evolution of land plants. In this study, effects of hypergravity environment on the morphology of the secondary wall and the lignin content were examined in Arabidopsis thaliana. Xylem vessels showed intense staining with phloroglucinol-HCl and autofluorescence under UV light at the basal region of the flower stalk when seedlings grown for 3 days after hypergravity treatment for 24 hours. And, the flower stalk exposed to hypergravity showed slight increase in the lignin content. These results suggest that the lignin formation is positively regulated under hypergravity.     

Hypergravity results in human platelet hyperactivity

Thrombotic diseases or fatalities have been reported to occasionally occur under conditions of hypergravity although the mechanism is still unclear. To investigate the effect of hypergravity on platelets that are the primary players in thrombus formation, platelet rich plasma (PRP) or washed platelets were exposed to hypergravity at 8 G for 15 minutes. No platelet aggregation was induced by 8 G alone, whereas ristocetin or collagen-induced platelet aggregation was significantly increased. The number of platelets adherent to immobilized fibrinogen and the area of platelets spreading on von Willbrand factor (VWF) matrix were increased simultaneously. Flow cytometry assay indicated that integrin αIIbß3 was partially activated in 8 Gexposed platelets, but there was no significant difference in P-selectin surface expression between platelets treated with 8 G and 1 G control. The results indicate that hypergravity leads to human platelet hyperactivity, but fails to incur essential platelet activation events, suggesting a novel mechanism for thrombotic diseases occurring under hypergravitional conditions.     

Increased molecular mass of hemicellulosic polysaccharides is involved in growth inhibition of maize coleoptiles and mesocotyls under hypergravity conditions.

Elongation growth of dark grown maize (Zea mays L cv. Cross Bantam T51) coleoptiles and mesocotyls was suppressed by hypergravity at 30 g and above. Acceleration at 300 g significantly decreased the mechanical extensibility of cell walls of both organs. Hypergravity increased the amounts of hemicellulose and cellulose per unit length in mesocotyl walls, but not in coleoptile walls. The weight average molecular masses of hemicellulosic polysaccharides were also increased by hypergravity in both organs. On the other hand, the activities of beta-glucanases extracted from coleoptile and mesocotyl cell walls were decreased by hypergravity. These results suggest that the decreased activities of beta-glucanases by hypergravity cause an increase in the molecular mass of hemicellulosic polysaccharides of both organs. The upshift of molecular mass of hemicellulosic polysaccharides as well as the thickening of cell walls under hypergravity conditions seems to be involved in making the cell wall mechanically rigid, thereby inhibiting elongation growth of maize coleoptiles and mesocotyls.     

Effect of hypergravity stimulus on XTH gene expression in Arabidopsis thaliana.

Hypergravity stimulus suppresses plant shoot growth by making the cell wall rigid. Xyloglucan endotransglucosylase/hydrolase (XTH) is involved in determining the rigidity of cell walls. We demonstrated that hypergravity influenced the expression of some XTH genes in shoots of Arabidopsis thaliana L.; in response to hypergravity stimulus of 300 g, the expression of AtXTH22 was up-regulated, while that of AtXTH15 was down-regulated. The effect of hypergravity on the expression of these genes was nullified by lanthanum chloride at 0.1 mM, suggesting that the expression of these XTH genes in Arabidopsis is under the control of the mechanoreceptor.     

Synaptopathy under conditions of altered gravity: Changes in synaptic vesicle fusion and glutamate release

Glutamate release and synaptic vesicle heterotypic/homotypic fusion were characterized in brain synaptosomes of rats exposed to hypergravity (10 G, 1 h). Stimulated vesicular exocytosis determined as KCl-evoked fluorescence spike of pH-sensitive dye acridine orange (AO) was decreased twice in synaptosomes under hypergravity conditions as compared to control. Sets of measurements demonstrated reduced ability of synaptic vesicles to accumulate AO (∼10% higher steady-state baseline level of AO fluorescence). Experiments with preloaded l-[¹⁴C]glutamate exhibited similar amount of total glutamate accumulated by synaptosomes, equal concentration of ambient glutamate, but the enlarged level of cytoplasmic glutamate measuring as leakage from digitonin-permeabilized synaptosomes in hypergravity. Thus, it may be suggested that +G-induced changes in stimulated vesicular exocytosis were a result of the redistribution of intracellular pool of glutamate, i.e. a decrease in glutamate content of synaptic vesicles and an enrichment of the cytoplasmic glutamate level. To investigate the effect of hypergravity on the last step of exocytosis, i.e. membrane fusion, a cell-free system consisted of synaptic vesicles, plasma membrane vesicles, cytosolic proteins isolated from rat brain synaptosomes was used. It was found that hypergravity reduced the fusion competence of synaptic vesicles and plasma membrane vesicles, whereas synaptosomal cytosolic proteins became more active to promote membrane fusion. The total rate of homo- and heterotypic fusion reaction initiated by Ca²⁺ or Mg²⁺/ATP remained unchanged under hypergravity conditions. Thus, hypergravity could induce synaptopathy that was associated with incomplete filling of synaptic vesicles with the neuromediator and changes in exocytotic release.     

Modification of cell wall architecture of wheat coleoptiles grown under hypergravity conditions.

Cell wall structure of wheat coleoptiles grown under continuous hypergravity (300 g) conditions was investigated. Length of coleoptiles exposed to hypergravity for 2-4 days from germination stage was 60-70% of that of 1 g control. The amounts of cell wall polysaccharides substantially increased during the incubation period both in 1 g control and hypergravity-treated coleoptiles. As a results, the levels of cell wall polysaccharides per unit length of coleoptile, which mean the thickness of cell walls, largely increased under hypergravity conditions. The major sugar components of the hemicellulose fraction, a polymer fraction extracted from cell walls with strong alkali, were arabinose (Ara), xylose (Xyl) and glucose (Glc). The molar ratios of Ara and Xyl to Glc in hypergravity-treated coleoptiles were higher than those in control coleoptiles. Furthermore, the fractionation of hemicellulosic polymers into the neutral and acidic polymers by the anion-exchange column showed that the levels of acidic polymers in cell walls of hypergravity-treated coleoptiles were higher than those of control coleoptiles. These results suggest that hypergravity stimuli bias the synthesis of hemicellulosic polysaccharides and increase the proportion of acidic polymers, such as arabinoxylans, in cell walls of wheat coleoptiles. These structural changes in cell walls may contribute to plant resistance to hypergravity stimuli.     

Hypergravity stimulus enhances primary xylem development and decreases mechanical properties of secondary cell walls in inflorescence stems of Arabidopsis thaliana

BACKGROUND AND AIMS: The xylem plays an important role in strengthening plant bodies. Past studies on xylem formation in tension woods in poplar and also in clinorotated Prunus tree stems lead to the suggestion that changes in the gravitational conditions affect morphology and mechanical properties of xylem vessels. The aim of this study was to examine effects of hypergravity stimulus on morphology and development of primary xylem vessels and on mechanical properties of isolated secondary wall preparations in inflorescence stems of arabidopsis. METHODS: Morphology of primary xylem was examined under a light microscope on cross-sections of inflorescence stems of arabidopsis plants, which had been grown for 3-5 d after exposure to hypergravity at 300 g for 24 h. Extensibility of secondary cell wall preparation, isolated from inflorescence stems by enzyme digestion of primary cell wall components (mainly composed of metaxylem elements), was examined. Plants were treated with gadolinium chloride, a blocker of mechanoreceptors, to test the involvement of mechanoreceptors in the responses to hypergravity. KEY RESULTS: Number of metaxylem elements per xylem, apparent thickness of the secondary thickenings, and cross-section area of metaxylem elements in inflorescence stems increased in response to hypergravity. Gadolinium chloride suppressed the effect of hypergravity on the increase both in the thickness of secondary thickenings and in the cross-section area of metaxylem elements, while it did not suppress the effect of hypergravity on the increase in the number of metaxylem elements. Extensibility of secondary cell wall preparation decreased in response to hypergravity. Gadolinium chloride suppressed the effect of hypergravity on cell wall extensibility. CONCLUSIONS: Hypergravity stimulus promotes metaxylem development and decreases extensibility of secondary cell walls, and mechanoreceptors were suggested to be involved in these processes.     

Cell proliferation of Paramecium tetraurelia under clinorotation.

It has been reported that Paramecium proliferates faster under microgravity in space, and slower under hypergravity (Kato et al., 2003). Effects of gravity on cell proliferation could be discussed in terms of energetics of swimming. Because of the characteristics of 'gravikinesis' as well as 'gravitaxis', Paramecium would decrease the energy expenditure under microgravity and increase it under hypergravity. The larger stock of energy would enhance the proliferation under microgravity. In order to simulate the effect of microgravity, we investigated the proliferation under clinorotation. When cells were rotated at 2.5 rpm, the proliferation rate decreased. Similar but less pronounced decrease was also found under low speed clinorotation (0.2 rpm).     

Effects of hypergravity on growth and cell wall properties of cress hypocotyls

Elongation growth of etiolated hypocotyls of cress (Lepidium sativum L.) was suppressed when they were exposed to basipetal hypergravity at 35 x g and above. Acceleration at 135 x g caused a decrease in the mechanical extensibility and an increase in the minimum stress-relaxation time of the cell wall. Such changes in the mechanical properties of the cell wall were prominent in the lower regions of hypocotyls. The amounts of cell wall polysaccharides per unit length of hypocotyls increased under the hypergravity condition and, in particular, the increase in the amount of cellulose in the lower regions was conspicuous. Hypergravity did not influence the neutral sugar composition of either the pectin or the hemicellulose fraction. The amount of lignin was also increased by hypergravity treatment, although the level was low. The data suggest that hypergravity modifies the metabolism of cell wall components and thus makes the cell wall thick and rigid, thereby inhibiting elongation growth of cress hypocotyls. These changes may contribute to the plants' ability to sustain their structures against hypergravity.     

Increased Molecular Mass of Hemicellulosic Polysaccharides is Involved in Growth Inhibition of Maize Coleoptiles and Mesocotyls under Hypergravity Conditions

Zea mays L. cv. Cross Bantam T51) coleoptiles and mesocotyls was suppressed by hypergravity at 30 g and above. Acceleration at 300 g significantly decreased the mechanical extensibility of cell walls of both organs. Hypergravity increased the amounts of hemicellulose and cellulose per unit length in mesocotyl walls, but not in coleoptile walls. The weight-average molecular masses of hemicellulosic polysaccharides were also increased by hypergravity in both organs. On the other hand, the activities of β-glucanases extracted from coleoptile and mesocotyl cell walls were decreased by hypergravity. These results suggest that the decreased activities of β-glucanases by hypergravity cause an increase in the molecular mass of hemicellulosic polysaccharides of both organs. The upshift of molecular mass of hemicellulosic polysaccharides as well as the thickening of cell walls under hypergravity conditions seems to be involved in making the cell wall mechanically rigid, thereby inhibiting elongation growth of maize coleoptiles and mesocotyls. Received 22 February 1999/ Accepted in revised form 20 April 1999     

Changes in food intake and growth rate in mice under hypergravity.

Mice exposed to hypergravity, especially soon after start of exposure, diminished the body weight and the food intake. The amount of food intake was kept less than that of the ground control during hypergravity breeding for 2 weeks. Furthermore, the weight of testis relative to the body weight increased compared to that of 1 G control although the relative weights of liver and kidney were not changed. The purpose of this study was whether the low growth rate of the body weight and the increase of the relative testis weight were induced by the decrease of the food intake under hypergravity. We divided 3 weeks old male mice to 3 groups; the 1 G (ground control), the food restricted (FR) under 1 G, and the 3 G group. The 3 G group bred for 2 weeks under the centrifuge at 3 G. The FR group was given the same amount of food as the group ate. The changes in the body weight and the relative weights of testis, spleen and seminal vesicle of the FR group were similar to those of the 3 G group. The hunger test revealed that only the FR group was hunger. Our results suggested that the decrease of the food intake both in response to hypergravity and the food restriction induced the decrease of the body weight but the increase of the relative testis weight.     

A hypergravity environment increases chloroplast size,photosynthesis, and plant growth in the moss Physcomitrella patens

The physiological and anatomical responses of bryophytes to altered gravity conditions will provide crucial information for estimating how plant physiological traits have evolved to adapt to significant increases in the effects of gravity in land plant history. We quantified changes in plant growth and photosynthesis in the model plant of mosses, Physcomitrella patens, grown under a hypergravity environment for 25 days or 8 weeks using a custom-built centrifuge equipped with a lighting system. This is the first study to examine the response of bryophytes to hypergravity conditions. Canopy-based plant growth was significantly increased at 10×g, and was strongly affected by increases in plant numbers. Rhizoid lengths for individual gametophores were significantly increased at 10×g. Chloroplast diameters (major axis) and thicknesses (minor axis) in the leaves of P. patens were also increased at 10×g. The area-based photosynthesis rate of P. patens was also enhanced at 10×g. Increases in shoot numbers and chloroplast sizes may elevate the area-based photosynthesis rate under hypergravity conditions. We observed a decrease in leaf cell wall thickness under hypergravity conditions, which is in contrast to previous findings obtained using angiosperms. Since mosses including P. patens live in dense populations, an increase in canopy-based plant numbers may be effective to enhance the toughness of the population, and, thus, represents an effective adaptation strategy to a hypergravity environment for P. patens.

     

The behavioral response of zebrafish to hypergravity conditions

Previous reports of the behavior of aquatic organisms in the microgravity environment of space (~10(-6) g) or during the brief weightless period of parabolic flight indicate that most species display a dramatic "looping" or "circling" response (De Jong et al. 1996, Anken, Ibsch and Rahmann 1998). However, the behavior of aquatic species under hypergravity conditions is less clear. Our objectives in the present study were to examine the behavioral response of adult zebrafish (Danio rerio) to hypergravity conditions (2-g), quantify changes in adult swimbladder volume, and to determine if the larvae of zebrafish are capable of accessing the air-water interface for initial swimbladder inflation under hypergravity conditions.     

Synaptopathy under conditions of altered gravity: Changes in synaptic vesicle fusion and glutamate release

Glutamate release and synaptic vesicle heterotypic/homotypic fusion were characterized in brain synaptosomes of rats exposed to hypergravity (10 G, 1 h). Stimulated vesicular exocytosis determined as KCl-evoked fluorescence spike of pH-sensitive dye acridine orange (AO) was decreased twice in synaptosomes under hypergravity conditions as compared to control. Sets of measurements demonstrated reduced ability of synaptic vesicles to accumulate AO (10% higher steady-state baseline level of AO fluorescence). Experiments with preloaded l-[¹⁴C]glutamate exhibited similar amount of total glutamate accumulated by synaptosomes, equal concentration of ambient glutamate, but the enlarged level of cytoplasmic glutamate measuring as leakage from digitonin-permeabilized synaptosomes in hypergravity. Thus, it may be suggested that +G-induced changes in stimulated vesicular exocytosis were a result of the redistribution of intracellular pool of glutamate, i.e. a decrease in glutamate content of synaptic vesicles and an enrichment of the cytoplasmic glutamate level. To investigate the effect of hypergravity on the last step of exocytosis, i.e. membrane fusion, a cell-free system consisted of synaptic vesicles, plasma membrane vesicles, cytosolic proteins isolated from rat brain synaptosomes was used. It was found that hypergravity reduced the fusion competence of synaptic vesicles and plasma membrane vesicles, whereas synaptosomal cytosolic proteins became more active to promote membrane fusion. The total rate of homo- and heterotypic fusion reaction initiated by Ca²⁺ or Mg²⁺/ATP remained unchanged under hypergravity conditions. Thus, hypergravity could induce synaptopathy that was associated with incomplete filling of synaptic vesicles with the neuromediator and changes in exocytotic release.     

Increase in expression level of alpha-tubulin gene in Arabidopsis seedlings under hypergravity conditions.

Under hypergravity conditions, elongation growth of plant shoots is suppressed. The analysis of the changes in gene expression by hypergravity treatment in Arabidopsis hypocotyls by the differential display method showed that a gene encoding alpha-tubulin, which is a component of microtubules, was up-regulated by hypergravity. In Arabidopsis six genes encoding alpha-tubulin (TUA1-TUA6) have been identified. In the present study, we examined the dose-response and the time course relations of the changes in the expression of all six alpha-tubulin genes in Arabidopsis hypocotyls grown under hypergravity conditions. The expression levels of all six alpha-tubulin genes, TUA1-TUA6, were increased by increasing gravity, although the extent was variable among genes. The increase in expression of all alpha-tubulin genes was detected within a few hours, when the seedlings grown at 1 g were transferred to 300 g condition. These results suggest that Arabidopsis hypocotyls regulate the expression level of six alpha-tubulin genes promptly in response to gravity stimuli. The increase in the amount of microtubules due to the activation of tubulin gene expression may be involved in the regulation by gravity signal of shoot growth.