Search

Two species of the clawed frog family, Xenopus laevis and X. tropicalis, are widely used as tools to investigate both normal and disease-state biochemistry, genetics, cell biology, and developmental biology. To support both frog specialist and non-specialist scientists needing access to these models for their research, a number of centralized resources exist around the world. These include centers that hold live and frozen stocks of transgenic, inbred and mutant animals and centers that hold molecular resources. This infrastructure is supported by a model organism database. Here, we describe much of this infrastructure and encourage the community to make the best use of it and to guide the resource centers in developing new lines and libraries.

Mitochondrial membrane permeability transition (MPT) plays a crucial role in apoptotic tail shortening during anuran metamor phosis. L-carnitine is known to shuttle free fatty acids (FFAs) from the cytosol into mitochondria matrix for -oxidation and energy production, and in a previous study we found that treatment with L-carnitine suppresses 3, 3', 5-triiodothyronine (T3) and FFA-induced MPT by reducing the level of FFAs. In the present study we focus on acetyl-L-carnitine, which is also involved in fatty acid oxidation, to determine its effect on T3-induced tail regression in Rana rugosa tadpoles and spontaneous tail regression in Xenopus laevis tadpoles. The ladder-like DNA profile and increases in caspase-3 and caspase-9 indicative of apoptosis in the tails of T3-treated tadpoles were found to be suppressed by the addition of acetyl-L-carnitine. Likewise, acetyl-L-carnitine was found to inhibit thyroid hormone regulated spontaneous metamorphosis in X. laevis tadpoles, accompanied by decreases in caspase and phospholipase A2 activity, as well as non-ladder-like DNA profiles. These findings support our previous conclusion that elevated levels of FFAs initiate MPT and activate the signaling pathway controlling apoptotic cell death in tadpole tails during anuran metamorphosis.

第28回宇宙利用シンポジウム (2012年1月23-24日. 日本学術会議), 港区, 東京都, Space Utilization Research (January 23-24, 2012. Science Council of Japan), Minato-ku, Tokyo Japan, 著者人数: 13名, 資料番号: SA6000032059, レポート番号: L19

第28回宇宙利用シンポジウム (2012年1月23-24日. 日本学術会議), 港区, 東京都, Space Utilization Research (January 23-24, 2012. Science Council of Japan), Minato-ku, Tokyo Japan, 著者人数: 13名, 資料番号: SA6000032060, レポート番号: L20

To investigate the relationship between gravity and the thyroid hormone on the pituitary-thyroid axis of amphibians, we raised frogs, Xenopus tropicana under hypergravity (5G) environment and gave thyroxin from St58 to St 59-60. The development of the frog was delayed under hypergravity condition. The total volume, cell size and cell number were reduced in pituitary, thyroid and cerebellum of hypergravity treated frogs. On the other hand, the morphological malfunctions of the treated frogs were recovered by the administration of thyroxin. These changes indicate that the drug recovers the declined functions. These results suggest that the thyroxin therapy rescues the hypofunction and dysfunction of the pituitary-thyroid axis from hypergravity., 資料番号: AA0063706064

To investigate the effects of hypergravity on the hypothalamus of amphibians, we raised frogs, Xenopus laevis under hypergravity environment (2 and 5 G) environments from hatch to the beginning of metamorphosis (St 57) and completion of metamorphosis (St 66). The frogs developed at 2 G and 5 G, but they were smaller than those raised at 1 G. In the hypergravity-treated hypothalamus, the total volume was reduced. The length of median eminence of treated frogs was also reduced. These morphological dwarves of the hypothalamus and median eminence indicate that hypergravity alters their endocrinological functions. These results suggest that gravity different from the terrestrial environment causes dysfunction and hypofunction in the endocrine systems, and that the organisms may change their inherent life cycles under such conditions., 資料番号: AA0063349090

This study attempts to investigate abnormal Programmed Cell Death (PCD) and abnormal gene expression induced under high gravity conditions. Uncleaved Xenopus laevis eggs at 10 and 20 min after insemination, and embryos at 2 cell stage were raised at 2 G and 5 G. Controls were raised in normal gravity. PCD for body surface and inner structures of hatched tadpoles was scored using the TUNEL staining method, and gene expression was analyzed by whole-mount in situ hybridization when eggs and embryos reached tail-bud embryos. The results showed that: (1) High gravity retarded the development of eggs and embryos and induced microcephaly and microphthalmia. (2) The number of dying cells in the skin of tadpoles developed from treated eggs and cell 2 stage embryos was greater at 2 G, but less at 5 G, compared to controls. The number of PCD in head epidermis of tadpoles developed from treated eggs and embryos was higher at 2 G than in controls, but the same as controls at 5 G. Brain PCD was found to be the same at 2 G, but higher at 5 G than for controls. (3) High gravity strongly suppressed the expression of the two genes, XAG1 (regulating cement gland formation) and Xotx2 (involved in forebrain and eye development). These findings suggest that high gravity induces abnormal PCD and suppresses gene functions essential for head and brain development, leading to retardation and various morphological abnormalities., 資料番号: AA0048095063

Previously it was reported that high gravity retarded the development of amphibian eggs and embryos, and induced two heads, microcephally and multiple malformation. Programmed cell death (PCD), or apoptosis, is an essential process for normal development, tissue homeostasis and harmful cell elimination in multicellular organisms. In order to further clarify the effects of high gravity environments on amphibian development, the present study investigates the induction of abnormal PCD by high gravity. Uncleaved Xenopus laevis eggs at 20 min after insemination, and embryos at cell stage 2 were raised in 5 G for 4 days. Controls were raised in normal gravity. PCD was analyzed for body surface and inner structures of developing embryos using more sensitive in situ endolabelling technique that identifies fragmented nuclear DNA in dying cells. The results showed that: 1) Treated eggs and embryos showed an initial delay of development at day 2. 2) The number of dying cells in the surface of embryos developed from treated eggs and cell stage 2 embryos was less compared to controls. 3) Autopsy revealed that control embryos had high numbers of PCDs in the skin, brain, eyes and forehead, while embryos developed from treated eggs and stage 2 embryos showed a random distribution of PCD in the brain, eyes and forehead. These observations suggest that high gravity induces suppression of PCD, which probably results in retarded development and various abnormalities., 資料番号: AA0046917019

In order to clarify the possible effects of high gravity environments on eggs and developing embryos, Rana rugosa unfertilized eggs, fertilized eggs and embryos at cell stages 2, 4, 8, 16, 32 and 64, the blastula, gastrula, neurula and tail- bud stages were raised in 2 G, 5 G, 7 G and 10 G up to premetamorphic Taylor and Kollros' stage 1. The results showed that: 1) High gravity strikingly retarded the development of eggs and embryos beginning treatment before the blastula stage and induce microcephally and multiple malformation, suggesting that high gravity is apt to disrupt the animal-vegetal axis. On the other hand, embryos beginning treatment after the gastrula stage showed a striking increase in the number of normal stage 1 tadpoles. 2) Autopsy revealed that brains, notochords and muscles were reduced in development and differentiation for embryos and tadpoles developed in high gravity., 資料番号: AA0045438015

第27回宇宙利用シンポジウム (2011年1月24日-25日, 宇宙航空研究開発機構宇宙科学研究所相模原キャンパス), 相模原市, 神奈川県, The Twenty-seventh Space Utilization Symposium (January 24-25, 2011. Institute of Space and Astronautical Science, Japan Aerospace Exploration Agency (JAXA)(ISAS)), Sagamihara, Kanagawa Japan, Previously we showed that amphibian embryos raised under hypergravity conditions develop the various abnormalities, including head-defects such as microcephaly and cyclopia as the most common. We also showed that the expression of Xag (cement grand marker), and Xotx2 (involved in fore- and midbrain and eye development) are suppressed in embryos under hypergravity. However, it is not known which region of the head is most sensitive to hypergravity, and if all regions of the head are affected by hypergravity. In the present study we investigated the effects of hypergravity on gene expression of Xag, Xotx2 and En2 (expressed in the region between midbrain and hindbrain) in tail-bud embryos of Xenopua laevis by wholemount in situ hybridization (WISH). Double staining with Xag and Xotx2 revealed that Xotx2 expression was suppressed in embryos exposed to 5G, although the region expressed Xag was not changed. Furthermore, En2 expression was not affected. These results strongly suggest that development of eye and fore- and midbrain is most sensitive to hypergavity and hypergravity does not affect the region posterior to the hindbrain., 著者人数: 13人, Number of authors: 13, 資料番号: AA0065129074

第22回宇宙利用シンポジウム(2006年1月17日-19日, 日本学術会議6階会議室 六本木、東京), The Twenty-second Space Utilization Symposium (January 17-19, 2006: Science Council of Japan, Roppongi, Tokyo, Japan), Developing oocytes arrest at the prophase of meiosis I, while the hormone progesterone restarts the process, leading to full maturation into a fertile egg. In this study we investigated the effects of hypergravity on this phase of maturation using oocytes of Xenopus. No differences were found between 2G, 5G and untreated controls regarding time to resumption of maturation, as judged by the appearance of a white spot in the animal hemisphere, whereas the white spot itself was larger and brighter in treated oocytes at time of appearance, and became even more pronounced as maturation proceeded. These findings suggest that although hypergravity does not affect the resumption of maturation, it might adversely affect the subsequent process and final results., 共催: 日本学術会議, Meeting sponsors: The Science Council of Japan, The Institute of Space and Astronautical Science, Japan Aerospace Exploration Agency (ISAS)(JAXA), 資料番号: AA0064113076

An investigation was undertaken to test the validity of the hypothesis that edema is the major cause of the low viability of haploid embryos and tadpoles. Chimeras were produced by exchanging the anterior and posterior parts of haploid embryos (H) with those of diploid embryos (D) by joining them after cutting transversely at the site immediately before and behind the pronephros (p). It was found that severe edema usually found in haploid embryos at the early stage is remarkably diminished in the Dp・H and H・pD chimeras which have diploid pronephroi in the anterior and posterior diploid halves, respectively. Especially, the Dp・H chimeras which consist of anterior diploid halves with pronephroi and posterior haploid halves are comparatively good in development. Of the Dp・H chimeras, 18 (9.7%) in Rana nigromaculata and 15 (10%) in Rana japonica attained the stage immediately before or after the completion of metamorphosis.

第26回宇宙利用シンポジウム(2010年1月25日-26日, 宇宙航空研究開発機構宇宙科学研究本部相模原キャンパス), The Twenty-sixth Space Utilization Symposium (January 25-26, 2010: ISAS/JAXA Sagamihara, Japan), In the present study, we investigated the effects of a strong static magnetic field (SMF) on oocyte maturation in Silurana tropicalis. First of all, we confirmed that oocyte maturation is triggered by progesterone, and this triggering is required for translation but not transcription. Under SMF the kinetics of initiation of oocyte maturation, as judged by the appearance of a white spot in the animal hemisphere, was found to be essentially the same as for controls. Cytological examination of treated specimens showed that the first polar body was excluded and spindle of meiosis II was successfully formed. However, as maturation proceeded, distribution of the pigment in the vegetable hemisphere became abnormal. The most of the white band characteristic of stage VI oocytes descended toward the vegetal pole and the rest of it invaded animal hemisphere. These results suggest that SMF might affect the progression of oocyte maturation., 共催: 日本学術会議, joint hosting: The Science Council of Japan, 資料番号: AA0064730084

第26回宇宙利用シンポジウム(2010年1月25日-26日, 宇宙航空研究開発機構宇宙科学研究本部相模原キャンパス), The Twenty-sixth Space Utilization Symposium (January 25-26, 2010: ISAS/JAXA Sagamihara, Japan), Life on earth is constantly exposed to a variety of natural and artificial electromagnetic and static magnetic forces. In order to help clarify the effects of such exposure on biological systems, we investigate the effects of static magnetic fields on embryos of Xenopus laevis as well as the expression of Xotx2 (an important regulator of fore- and midbrain morphogenesis) and Xag1 (essential for cement gland formation). Results showed that: (1) Embryos developed from treated eggs were non-viable and had multiple malformations including two- or microcephalic heads, deformed cement glands and abnormal body proportions. The eyes and brain were morphological deficient or entirely absent. (2) Xotx2 and Xag1 expression was suppressed. Abnormal expression of these genes together with poor viability are possibly responsible for obstruction of egg cortex rotation, incomplete formation of grey crescent and changes in spatial distribution of genes., 共催: 日本学術会議, 著者人数: 17人, joint hosting: The Science Council of Japan, 資料番号: AA0064730085

第25回宇宙利用シンポジウム(2009年1月14日-15日, 宇宙航空研究開発機構宇宙科学研究本部相模原キャンパス), The Twenty-fifth Space Utilization Symposium (January 14-15, 2009: ISAS/JAXA Sagamihara, Japan), Xenopus embryos raised under hypergravity condition develop a variety of abnormalities. Among them, head-defects such as microcephaly and cyclopia are the most common. Normal head formation is regulated by Wnt signaling. Inhibition of Wnt signaling causes head-defects in embryos, and ectopic activation of Wnt signaling induces extra body axis with head. Embryos are most sensitive to hypergravity within 30 min after fertilization, and activation of endogenous Wnt signaling occurs at almost the same time by cortical rotation which is sensitive to gravity. These facts suggest that the target of hypergravity might be Wnt signaling. In order to investigate this possibility, Wnt signaling activity in embryos raised under hypergravity conditions was analyzed by the measuring the expression of the Wnt-target genes. Quantitative RT-PCR revealed that the expression of some of Wnt-target genes was reduced in these embryos. This result strongly suggests that Wnt signaling is the target of the hypergravity derived head-defects in Xenopus embryos., 資料番号: AA0064297059

第22回宇宙利用シンポジウム(2006年1月17日-19日, 日本学術会議6階会議室 六本木、東京), The Twenty-second Space Utilization Symposium (January 17-19, 2006: Science Council of Japan, Roppongi, Tokyo, Japan), In order to analyze effects of hypergravity on the pituitary gland and the target organs of frogs, we raised Xenopus laevis in two and five gravity environments from fertilization to beginning of metamorphosis(St57) and completion of metamorphosis (st66). The frogs developed in 2G and 5G, but the body size of exposed frogs became smaller. Moreover, development of the animals was delayed in both environments. In the hypergravity treated pituitary, volume and cell size were downsized, and cell number was reduced. In the treated thyrold, volume and epithelium were also undersized. These morphological changes indicate that hypergravity alters the function of pituitary and thyroid. The results suggest that gravities unlike the terrestrial environment cause dysfunction and hypofunction in endocrine systems, and that the animals may not be able to maintain the inherent life cycles., 共催: 日本学術会議, 著者人数: 12人, Meeting sponsors: The Science Council of Japan, The Institute of Space and Astronautical Science, Japan Aerospace Exploration Agency (ISAS)(JAXA), 資料番号: AA0064113077

資料番号: SA1004097000

資料番号: SA1004098000

Gynogenetic diploids were produced from eggs of 10 female Rana rugosa by refrigeration after insemination with UV-irradiated sperm of 10 males of the same species. They were compared with the control diploids and gynogenetic haploids in developmental capacity. When refrigerated at 0∿2℃ for two hours after insemination with UV-irradiated sperm, 73.10f the eggs cleaved normally, and 65.1 0x1.5443cp+823nd 20.916032271143f these normally cleaved eggs hatched normally and became normally metamorphosed frogs, respectively. The diploidy of the individuals in the experimental series was confirmed by counting chromosomes at the tadpole stage. Of the 457 gynogenetic diploids, 418 were females and 39 (8.5%) were males. There were 931 females and 25 (2.6%) males among the 956 offspring of six gynogenetic diploid males mated with normal females. As the great majority of gynogenetic diploids and the offspring of gynogenetic males mated with normal females were females, it is assumed that the male is heterogametic in Rana rugosa and that gynogenetic males are genetic females. In the gynogenetic diploid tadpoles, there were 26 (2.4%) black-eyed and 23 (2.1%) gray-eyed mutants.

Of the 1209 eggs of Rana rugosa refrigerated after insemination with sperm of the same species, 832 (68.8%) cleaved normally, and 338 (28.0%) became normally feeding tadpoles. By counting the chromosomes of these tadpoles, it was found that 254 (82.5%) were triploids (3n=39), seven were diploids, eight were mosaics, and the remaining 39 were uncertain in ploidy. Of the triploid tadpoles, 242 became normally metamorphosed frogs and 11 of them were reared until sexual maturity. No significant differences were observed between these triploids and the control diploids in development and growth rate. All the 211 triploid frogs were males or hermaphrodites which were transforming into males. Of the 2272 eggs from four field-caught females mated with eight of the 11 triploid males, 360 (15.8%) cleaved normally and 136 (6.0%) became normally feeding tadpoles. Only one of these tadpoles completed metamorphosis, while none of the eggs of the same four females mated with the other three triploid males cleaved normally. The chromosomes were counted in 81 tadpoles produced from four diploid females mated with five triploid males. The results showed that 79 of them were aneuploids between diploid and triploid, and the remaining two were diploids.