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Hypergravity-induced changes in gene expression in Arabidopsis hypocotyls.

Under hypergravity conditions, the cell wall of stem organs becomes mechanically rigid and elongation growth is suppressed, which can be recognized as the mechanism for plants to resist gravitational force. The changes in gene expression by hypergravity treatment were analyzed in Arabidopsis hypocotyls by the differential display method, for identifying genes involved in hypergravity-induced growth suppression. Sixty-two cDNA clones were expressed differentially between the control and 300 g conditions: the expression levels of 39 clones increased, whereas those of 23 clones decreased under hypergravity conditions. Sequence analysis and database searching revealed that 12 clones, 9 up-regulated and 3 down-regulated, have homology to known proteins. The expression of these genes was further analyzed using RT-PCR. Finally, six genes were confirmed to be up-regulated by hypergravity. One of such genes encoded 3-hydroxy-3-methylglutaryl-Coenzyme A reductase (HMGR), which catalyzes a reaction producing mevalonic acid, a key precursor of terpenoids such as membrane sterols and several types of hormones. The expression of HMGR gene increased within several hours after hypergravity treatment. Also, compactin, an inhibitor of HMGR, prevented hypergravity-induced growth suppression, suggesting that HMGR is involved in suppression of Arabidopsis hypocotyl growth by hypergravity. In addition, hypergravity increased the expression levels of genes encoding CCR1 and ERD15, which were shown to take part in the signaling pathway of environmental stimuli such as temperature and water, and those of the alpha-tubulin gene. These genes may be involved in a series of cellular events leading to growth suppression of stem organs under hypergravity conditions.

Arabidopsis↗

A mechanism of adaptation to hypergravity in the statocyst of Aplysia californica.

The gravity-sensing organ of Aplysia californica consists of bilaterally paired statocysts containing statoconia, which are granules composed of calcium carbonate crystals in an organic matrix. In early embryonic development, Aplysia contain a single granule called a statolith, and as the animal matures, statoconia production takes place. The objective of this study was to determine the effect of hypergravity on statoconia production and homeostasis and explore a possible physiologic mechanism for regulating this process. Embryonic Aplysia were exposed to normogravity or 3 x g or 5.7 x g and each day samples were analyzed for changes in statocyst, statolith, and body dimensions until they hatched. In addition, early metamorphosed Aplysia (developmental stages 7-10) were exposed to hypergravity (2 x g) for 3 weeks, and statoconia number and statocyst and statoconia volumes were determined. We also determined the effects of hypergravity on statoconia production and homeostasis in statocysts isolated from developmental stage 10 Aplysia. Since prior studies demonstrated that urease was important in the regulation of statocyst pH and statoconia formation, we also evaluated the effect of hypergravity on urease activity. The results show that hypergravity decreased statolith and body diameter in embryonic Aplysia in a magnitude-dependent fashion. In early metamorphosed Aplysia, hypergravity decreased statoconia number and volume. Similarly, there was an inhibition of statoconia production and a decrease in statoconia volume in isolated statocysts exposed to hypergravity in culture. Urease activity in statocysts decreased after exposure to hypergravity and was correlated with the decrease in statoconia production observed. In short, there was a decrease in statoconia production with exposure to hypergravity both in vivo and in vitro and a decrease in urease activity. It is concluded that exposure to hypergravity downregulates urease activity, resulting in a significant decrease in the formation of statoconia.

Analysis of Variance↗

Graviperception in growth inhibition of plant shoots under hypergravity conditions produced by centrifugation is independent of that in gravitropism and may involve mechanoreceptors.

Hypergravity caused by centrifugation inhibits elongation growth of shoots by decreasing the cell wall extensibility via suppression of xyloglucan breakdown as well as by the thickening of cell walls. The mechanism of graviperception in hypergravity-induced growth inhibition was investigated in Arabidopsis [A. thaliana (L.) Heynh.] hypocotyls and azuki bean (Vigna angularis Ohwi et Ohashi) epicotyls. Hypergravity caused growth suppression in both sgr1-1 and pgm1, which are Arabidopsis mutants deprived of gravitropism, as in wild-type plants, suggesting that the graviperception in hypergravity-induced growth inhibition of shoots is independent of that in gravitropism. Hypergravity had no effects on growth of azuki bean epicotyls or Arabidopsis hypocotyls in the presence of lanthanum or gadolinium, which are blockers of mechanoreceptors. Moreover, lanthanum or gadolinium at the same concentration had no influence on gravitropism of azuki bean epicotyls and Arabidopsis hypocotyls. Hypergravity had no effects on cell wall extensibility and affected neither xyloglucan metabolism nor the thickness of cell walls in the lanthanum- or gadolinium-treated azuki bean epicotyls. Lanthanum or gadolinium inhibited the hypergravity-induced increase in the pH of the apoplastic fluid in the epicotyls, which is involved in the processes of the suppression of xyloglucan breakdown due to hypergravity. These findings suggest that plants perceive the hypergravity stimuli by mechanoreceptors in the plasma membrane, and utilize the perceived signal to regulate the growth rate of their shoots.

Arabidopsis↗

Male Drosophila melanogaster flies exposed to hypergravity at young age are protected against a non-lethal heat shock at middle age but not against behavioral impairments due to this shock.

Previous studies have shown that exposing flies to hypergravity (3 or 5 g) for two weeks at young age slightly increases longevity of male flies and survival time at 37 degrees C of both sexes, and delays an age-linked behavioral change. The present experiments tested whether hypergravity could also protect flies from a non-lethal 37 degrees C heat shock applied at young, middle or old age (2, 4 or 6 weeks of age). Various durations of exposure at 37 degrees C had similar deleterious effects on climbing activity, spontaneous locomotor activity and learning in flies that lived or not in hypergravity at young age. Therefore, hypergravity does not protect the behavior of flies from a deleterious non-lethal heat shock. Hypergravity increased longevity of virgin males and decreased that of mated ones; it also increased longevity of virgins at 25 degrees C, the usual rearing temperature, but not at 30 degrees C. Thus, the positive effect of hypergravity on longevity is observed only if flies are not subjected to living conditions decreasing longevity, like mating and high temperature. Finally, 4 weeks-old males that lived in hypergravity at young age lived slightly longer (+ 15%) after a non-lethal heat shock (60 or 90 min at 37 degrees C) than flies that always lived at 1 g, but this positive effect of hypergravity was not observed in females or in older males. Therefore, all these results show that hypergravity exposure can help male middle-aged flies recovering from a heat shock, but does not protect them from behavioral impairments linked to this shock: a mild stress occurring at young age can partially protect from a moderate stress at middle age.

Aging↗

Growth restoration in azuki bean and maize seedlings by removal of hypergravity stimuli.

Hypergravity stimuli, gravitational acceleration of more than 1 x g, decrease the growth rate of azuki bean epicotyls and maize coleoptiles and mesocotyls by decreasing the cell wall extensibility via an increase in the molecular mass of matrix polysaccharides. An increase in the pH in the apoplastic fluid is hypothesized to be involved in the processes of the increase in the molecular mass of matrix polysaccharides due to hypergravity. However, whether such physiological changes by hypergravity are induced by normal physiological responses or caused by physiological damages have not been elucidated. In the present study, we examined the effects of the removal of hypergravity stimuli on growth and the cell wall properties of azuki bean and maize seedlings to clarify whether the effects of hypergravity stimuli on growth and the cell wall properties are reversible or irreversible. When the seedlings grown under hypergravity conditions at 300 x g for several hours were transferred to 1 x g conditions, the growth rate of azuki bean epicotyls and maize coleoptiles and mesocotyls greatly increased within a few hours. The recovery of growth rate of these organs was accompanied by an immediate increase in the cell wall extensibility, a decrease in the molecular mass of matrix polysaccharides, and an increase in matrix polysaccharide-degrading activities. The apoplastic pH also decreased promptly upon the removal of hypergravity stimuli. These results suggest that plants regulate the growth rate of shoots reversibly in response to hypergravity stimuli by changing the cell wall properties, by which they adapt themselves to different gravity conditions. This study also revealed that changes in growth and the cell wall properties under hypergravity conditions could be recognized as normal physiological responses of plants. In addition, the results suggest that the effects of microgravity on plant growth and cell wall properties should be reversible and could disappear promptly when plants are transferred from microgravity to 1 x g. Therefore, plant materials should be fixed or frozen on orbit for detecting microgravity-induced changes in physiological parameters after recovering the materials to earth in space experiments.

Cell Wall↗

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.

Arabidopsis↗

Effects of hypergravity on ovarian-hypophyseal function in antepartum and postpartum rats.

BACKGROUND: Rats exposed to microgravity during the post-implantation phase of pregnancy had minimal alterations in ovarian and hypophyseal parameters during the antepartum and postpartum periods. In the current study, a similar parallel experimental design was employed to ascertain the effects of hypergravity on ovarian and hypophyseal function. HYPOTHESIS: We hypothesized that hypergravity exposure during the post-implantation stage of pregnancy would not alter antepartum and postpartum ovarian and hypophyseal function. METHODS: Pregnant rats were assigned to hypergravity (1.5 G, 1.75 G, or 2.0 G), rotational control, or stationary control groups (n = 10 each group) beginning on gestation day 11 and ending on day 20. Hypophyseal and ovarian analyses were conducted on 5 of the animals from each group at day 20. The remaining animals in each group were allowed to go to term and the same analyses were conducted 3 h postpartum. RESULTS: Hypergravity at all levels decreased the percent body mass gain from gestation day 11 to 20 (p < 0.05); however, the wet weight of the pituitaries and ovaries was not changed. There was no effect of hypergravity on the number of healthy or atretic antral follicles of any size at gestation day 20 or postpartum. The number of corpora lutea of pregnancy was decreased in all hypergravity groups, but the number of live fetuses at gestation day 20 or pups at term was not altered. Plasma concentrations of luteinizing hormone (LH), follicle stimulating hormone (FSH), prolactin, and progesterone were not changed at gestation day 20 or postpartum. Pituitary content of LH, FSH, and prolactin was not altered by hypergravity at gestation day 20, but LH content was significantly increased (p < 0.05) at 1.5 and 1.75 G postpartum. CONCLUSIONS: We conclude that hypergravity, up to and including 2.0 G, is compatible with maintenance of pregnancy and has minimal effects on hypophyseal parameters. Ovarian follicles are not altered by hypergravity, but corpora lutea may regress at a more rapid rate.

Animals↗

Perception mechanism of gravity stimuli in hypergravity-induced growth inhibition of azuki bean roots.

We reported that elongation growth of plant shoots and roots is suppressed by hypergravity, with the rate decreasing in proportion to logarithm of the magnitude of gravity. In hypergravity-induced growth inhibition of shoots, graviperception is supposed to be independent of that in gravitropism and to involve mechanoreceptors. However, the graviperception mechanism in the hypergravity-induced growth inhibition of roots is not known. In the present study, we compared the mechanism in the hypergravity-induced growth inhibition of roots with that in gravitropism. The removal of root cap did not influence hypergravity-induced growth inhibition of roots, although the gravitropic curvature was completely inhibited. Hypergravity had no effects on growth of azuki bean roots in the presence of lanthanum or gadolinium, which are blockers of mechanoreceptors. On the contrary, lanthanum or gadolinium at the same concentration did not influence gravitropism of roots. These results suggest that the graviperception mechanism in the hypergravity-induced growth inhibition of roots is independent of that in gravitropism. Hypergravity-induced growth inhibition of azuki bean roots was observed irrespective of the direction of stimuli, which disappeared in the presence of lanthanum or gadolinium. Thus, in the hypergravity-induced growth inhibition, roots may perceive the gravity signal by mechanoreceptors on the plasma membrane independently of the direction of stimuli, and may utilize it to regulate their growth rate.

Fabaceae↗

Effects of hypergravity on proliferation and differentiation of osteoblast-like cells.

We investigated the effects of hypergravity on DNA synthesis and alkaline phosphatase (ALP) activity in cloned osteoblast-like cells, MC3T3-E1. Hypergravity (5 x g) stimulated DNA synthesis in these cells in a time-dependent manner and increased it approximately up to 150% of that of the control (1 x g). 12-O-Tetra-decanoylphorbol-13-acetate (TPA), a protein kinase C activator, and insulin-like growth factor I (IGF-I) enhanced DNA synthesis additively with hypergravity (5 x g). An increase in ALP activity induced by 10% fetal calf serum (FCS) was suppressed by hypergravity (2 x g, 5 x g). Five x g completely suppressed the increase in ALP activity. TPA and hypergravity (2 x g) suppressed the increase in ALP activity induced by FCS additively. Hypergravity (5 x g) showed no significant effect on cAMP nor cGMP production in these cells, but increased prostaglandin E2 (PGE2) production. Exogenous PGE2 stimulated DNA synthesis in these cells but had little effect on 10% FCS-induced ALP activity. These results suggest that hypergravity stimulates proliferation but suppresses differentiation of osteoblast-like cells through a pathway independent of the activation of protein kinase C and the production of cyclic nucleotides, and that hypergravity and IGF-I stimulate proliferation of these cells through an independent signal transduction pathway. Moreover, our data strongly suggest that PGE2 mediates the signalling of hypergravity on the proliferation of osteoblast-like cells.

Alkaline Phosphatase↗

Hippocampal gene expression is modulated by hypergravity.

We used the cDNA microarray technique to monitor simultaneously possible changes induced by hypergravity in the expression level of thousands of hippocampal genes. We tested the mRNA level of about 5000 genes in the hippocampus of mice subjected to 1.09 g (1g) or to 1.85 g (2g) for five repeated 1-h daily rotations in a centrifuge (g = 9.81 m/s2). Data were compared with those obtained for mice kept stationary (C). The ratios 1g/C and 2g/C identified genes affected by rotation and rotation + hypergravity, respectively, whereas 2g/1g ratio identified those affected by hypergravity. We found that about 200 genes were affected by rotation and/or rotation + hypergravity. Almost all the genes affected by rotation + hypergravity were up-regulated, only five being down-regulated. The modulated genes code for proteins involved in a wide range of cellular functions (DNA/RNA metabolism, protein processing, intermediate metabolism, cytoskeleton and motility, cell cycle and apoptosis, signal transduction, neuronal structure/function), suggesting that rotation + hypergravity may affect several aspects of the hippocampal function in order to compensate for environmental changes. Six genes directly or indirectly involved in synaptic transmission and plasticity (proSAAS, neuroblastoma ras oncogene, ESTs moderately similar to thymosin beta-10, syndet, inhibin beta E and Ngfi-A binding protein 2) were found to be significantly modulated by hypergravity and unaffected or only slightly affected by rotation. The modulation by hypergravity of these genes suggests that this stimulus might induce plastic remodelling of the hippocampal circuits, possibly both at structural and functional level.

Animals↗

Hypergravity induces phosphorylation of p53 at serine 15, but not an expression of p53-downstream genes.

We investigated the p53 signaling pathway induced by hypergravity in the human glioblastoma cell line A172. Hypergravity (20 x g) induced the accumulation of p53 and the phosphorylation of p53 at Ser-15. The phosphorylation of p53 with hypergravity was not inhibited by wortmannin, the PI3-kinase inhibitor. This indicated that the p53 signal pathway induced by hypergravity is different from other p53 signal pathways, such as that of the DNA damage signal. Hypergravity did not induce an expression of the genes Waf-1 or Bax, located downstream from p53. We also examined the expression of genes with hypergravity by using a DNA microarray containing oligo DNA from 30,000 human genes. Hypergravity (20 x g, 6 h) did induce the expression of some genes concerned with the cell signaling pathway and cytoskeleton of the cell, but not any of the p53-downstream genes. DNA microarray revealed the induction of many genes to enable the cells to adapt to the hypergravity environment.

Amino Acid Sequence↗

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.

Arabinose↗

Quantitative changes in mRNA expression of glutamate receptors in the rat peripheral and central vestibular systems following hypergravity.

In order to investigate the mechanisms responsible for adaptation to altered gravity, we assessed the changes in mRNA expression of glutamate receptors in vestibular ganglion cells, medial vestibular nucleus, spinal vestibular nucleus/lateral vestibular nucleus, cerebellar flocculus, and uvula/nodulus from rats exposed to hypergravity for 2 h to 1 week using real-time quantitative RT-PCR methods. The mRNA expression of GluR2 and NR1 receptors in the uvula/nodulus and NR1 receptors in the medial vestibular nucleus increased in animals exposed to 2 h of hypergravity, and it decreased gradually to the control level. The mRNA expression of GluR2 receptors in vestibular ganglion cells decreased in animals exposed to 1 week of hypergravity. Neither the metabotropic glutamate receptor 1 nor delta2 glutamate receptor in flocculus and uvula/nodulus was affected by a hypergravity load for 2 h to 1 week. It is suggested that the animals adapted to the hypergravity by enhancing the cerebellar inhibition of the vestibular nucleus neurons through activation of the NR1 and GluR2 receptors on the Purkinje cells in uvula/nodulus especially at the early phase following hypergravity. In the later phase following hypergravity, the animals adapted to the hypergravity by reducing the neurotransmission between the vestibular hair cells and the primary vestibular neurons via down-regulation of the postsynaptic GluR2 receptors in the vestibular periphery.

Animals↗

Repeated acute exposures to hypergravity during early development subtly affect CD-1 mouse neurobehavioural profile.

Exposure to altered gravitational environment, especially during critical ontogenetic phases, may induce persistent nervous system modifications and behavioural anomalies. This study evaluated the effects of hypergravity exposure on the development of the nervous system and assessed the relevance of parity in the mother's responses to altered gravitational stimuli. CD-1 mouse pups of both sexes delivered by primiparous and biparous dams were exposed to 1h of 2 G rotationally induced hypergravity from PND2 to PND9. Sensorimotor responses and somatic growth were daily measured (PND2-PND15), ultrasonic vocalisations recorded on PNDs 2, 5 and 9, and homing behaviour evaluated on PND12. In addition, spatial orientation ability was assessed in a T-maze on PND18, while mice exploratory behaviour and locomotor activity were evaluated in an open-field test (PND21). Long-term effects of hypergravity exposure on both spatial learning (Morris water-maze test) and brain levels of NGF and BDNF were also investigated at adulthood. Rotation per se induced a delay in somatic growth, sensorimotor responses and ultrasonic vocalisation profile, while hypergravity highlighted sex differences in open-field behaviour. Strategies to solve a spatial learning task, rather than learning per se, were affected by early exposure to rotation, while hypergravity selectively altered behavioural profile in the reversal phase of the test. Early exposure to rotation per se also decreased hypothalamic BDNF levels, while hypergravity reduced NGF levels in the frontal cortex. Previous maternal experience did not interact with hypergravity exposure, while differences between offspring of primiparous and biparous dams were observed in sensorimotor development and exploratory behaviour.

Animals↗

Effects of amygdala or hippocampus lesion on hypergravity-induced motion sickness in rats.

We examined the effects of amygdala lesion (AL) or hippocampal lesion (HL) on hypergravity-induced motion sickness in rats. Rats do not vomit, but the behavior known as pica, the eating of non-nutritive substances such as kaolin, can be used as an index of motion sickness. In the present study, hypergravity-induced kaolin intake and apomorphine-induced kaolin intake were measured before and after brain lesions. After AL, hypergravity-induced kaolin intake and the ratio of the hypergravity- to apomorphine-induced kaolin intakes were decreased. These results indicate that AL suppressed motion sickness more than pica behavior itself, suggesting that the amygdala plays an important role in the development of motion sickness in rats. Conversely, after HL, hypergravity-induced kaolin intake was increased, as was the ratio of the hypergravity- to apomorphine-induced kaolin intakes. These results indicate that HL aggravates motion sickness induced by hypergravity in rats, suggesting that the hippocampus counteracts motion sickness.

Amygdala↗

Stimulation of cyclic GMP efflux in human melanocytes by hypergravity generated by centrifugal acceleration.

Gravity alteration (micro- and hypergravity) is known to influence cell functions. As guanosine 3',5'-cyclic monophosphate (cGMP) plays an important role in human melanocyte functions and different guanylyl cyclase isoforms are responsible for cGMP synthesis in human non-metastatic and metastatic melanoma cells, we investigated the effects of hypergravity on the regulation of cGMP levels in cultured human melanocytes and in melanoma cell lines with different metastatic potentials. Hypergravity was produced by horizontal centrifugal acceleration. Here we report that long-term application of hypergravity (up to 5 g for 24 h) stimulated cGMP efflux in cultured melanocytes and in non-metastatic melanoma cells in the presence of 0.1 mM 3-isobutyl-1-methylxanthine (IBMX), a non-selective phosphodiesterase (PDE) inhibitor. Under these conditions, cAMP synthesis and melanin production were up-regulated in pigmented melanocytes and non-metastatic melanoma cells. Hypergravity also stimulated cGMP transport in the presence of 1 microM trequinsin, an inhibitor of cGMP-binding PDE (PDE5) and of transport by multidrug resistance proteins MRP4/5, whereas 50 microM trequinsin partially inhibited cGMP transport. Transport was further inhibited by probenecid, an inhibitor of endogenous non-selective transporters as well as of MRP4/5 and by cycloheximide as an inhibitor of de novo protein synthesis. In contrast, hypergravity did not affect cGMP efflux in metastatic melanoma cells, which might be related to an up-regulated cGMP efflux at 1 g. The results of the present study indicate that hypergravity may stimulate cGMP efflux in melanocytes and in non-metastatic melanoma cells most probably by an enhanced expression of endogenous transporters and/or MRP4/5. Thus, an altered acceleration vector may induce signaling events in melanocytic cells.

Acceleration↗

Stimulative effect of high-level hypergravity on differentiated functions of osteoblast-like cells.

The exposure of freshly isolated osteoblasts and osteoblast-like cells to high-level hypergravity caused the inhibition of cell growth, elevation of cAMP content, and the stimulation of differentiated functions such as alkaline phosphatase activity, collagen synthesis, and osteocalcin synthesis. Blockage of elevation of cAMP by SQ22536, an inhibitor of adenylate cyclase, resulted in the inhibition of the hypergravity-stimulated alkaline phosphatase activity, indicating that cAMP is the intracellular mediator of this action of hypergravity. H89, an inhibitor of cAMP-dependent protein kinase (PKA), further inhibited the cell growth that was already inhibited by the hypergravity, and further stimulated the alkaline phosphatase activity that was already stimulated by hypergravity. If cAMP acts through the PKA system, H89 should have blocked the changes in cell function effected by the exposure to hypergravity. Therefore the elevated intracellular cAMP by the exposure of hypergravity caused the changes in cell function by a PKA-independent pathway.

Adenine↗

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.

Animals↗