Hyla Arborea Japonica Research Articles

Prolactin, an Activator of Epithelial Na+Channel, Inhibits Basolateral K+Channels in Adult Tree Frog Skin

Adult amphibian skin actively transports Na+ from its apical to basolateral side while in turn, K+ is recycled through Na+, K+-ATPase and K+ channels located in the basolateral membrane. We previously found that PRL stimulates Na+ transport in the skin of the adult tree frog (Hyla arborea japonica) via an increase in the open-channel density of the epithelial Na+ channel (ENaC). If PRL also activates basolateral K+ channels, this activation would help to stimulate Na+ transport, too. Whether PRL does indeed stimulate basolateral K+ channels in the adult tree frog was examined by measuring the short-circuit current across nystatin-treated skin. Both tolbutamide, a K(ATP) channel blocker, and tetrapentylammonium (TPA), a KCa channel blocker, blocked the current, the effect of TPA being more powerful than that of tolbutamide. Contrary to expectation, PRL inhibited the basolateral K+ channels in this skin. In the presence of basolateral amiloride, PRL still inhibited the basolateral K+ current, suggesting that the (Na+)-H+ exchanger located in the basolateral membrane does not mediate the inhibitory effect of PRL on the basolateral K+ channels in Hyla.

Prolactin increases open-channel density of epithelial Na+ channel in adult frog skin.

The short-term effect of prolactin on the skin of the adult tree frog Hyla arborea japonica was investigated using current-fluctuation analysis. Basolateral application of ovine prolactin (10 microg ml(-1)) (1) increased the amiloride-blockable short-circuit current (SCC) across the skin 2.6+/-0.4-fold and (2) increased the open-channel density (M) of the epithelial Na(+) channel 6.1+/-1.2-fold but decreased the single-channel current i to 0.4+/-0.1 times the control value (N=9). The increase in SCC induced by prolactin was thus due to an increase in M, not i. Apparently, in amphibians prolactin has not only a counteracting effect on metamorphosis but also a stimulatory effect on the development of adult-type features, such as this amiloride-blockable SCC.

Immunohistochemistry of endocrine cells in the alimentary tract of the tree frog,hyla arborea japonica

The regional distribution and relative frequencies of endocrine cells were studied immunohistochemically (PAP methods) in the alimentary tract of the tree frog, Hyla arborea japonica, using specific antisera against serotonin, somatostatin, bovine Sp‐1/chromogranin (BCG), cholecystokinin (CCK)‐8, vasoactive intestinal polypeptide (VIP), gastrin, bombesin, secret in and pancreatic polypeptide (PP). Six kinds of endocrine cells were identified in this study. These immunoreactive cells were located in the gastric glands of stomach regions and in the basal portion of the epithelium of the intestinal tract or esophagus with variable frequencies. They were spherical or spindle‐shaped. Serotonin‐immunoreactive cells were observed in the whole alimentary tract including the esophagus. Somatostatin‐immunoreactive cells were also detected throughout the alimentary tract except the rectum. CCK‐8‐immunoreactive cells were observed from the pylorus to ileum. VIP‐immunoreactive cells were restricted to the rectum. Bomb...

Immunohistochemical localization of endocrine cells in the alimentary tracts of six frog species

A peroxidase‐antiperoxidase method was used to detect the cells showing immunoreactivities to six hormone antibodies in the alimentary tracts of six frog species, Rana nigromaculata, R. rugosa, R. amurensis coreana, R. catesbeiana, Bombina orientalis, and Hyla arborea japonica, inhabiting Korea. The cells immunoreactive to gastrin and cholecystokinin‐8 were observed in the pylorus of the stomachs and in the small intestines of all frog species examined. In contrast, these somatostatin‐immunoreactive cells were identified in the esophagus and the whole gastrointestinal tracts, but were absent from the large intestines in R. rugosa, R. catesbeiana, B. orientalis, and H. arborea japonica. The pancreatic polypeptide (PP)‐immunoreactive cells represented their distribution limited to the small intestines of R. amurensis coreana and H. arborea japonica, and they were additionally identified in the pylorus of the stomachs in the other four species. Serotonin‐ and glucagon‐ immunoreactive cells revealed different regional distributions in which the former were observed throughout the whole alimentary tracts in all frog species investigated, whereas the latter were not found in these regions at all. Endocrine cells were relatively abundant in the pyloric portion of the stomach compared to other organs. The present study showed that all endocrine cells except for PP had a similar distribution in the alimentary tracts of all frog species used

Angiotensin II suppresses water absorption through the ventral skin of Japanese tree-frogs in vitro.

We previously described two different water absorption systems in the ventral skin of the Japanese tree-frog, Hyla arborea japonica: i.e., a rapid enhanced flow, which is observed in dehydrated tree-frogs or those stimulated by adrenaline beta-agonists or vasotocin, and a slow basal flow, which is observed in normally hydrated frogs during the non-breeding season. The rapid flow is completely blocked by ouabain, which has no effects on the slow basal flow. In the present experiment, we show that the vaso-constrictive hormone angiotensin II completely inhibits basal water absorption, but has no effect on rapid water absorption. These results confirm our previous finding that the two water absorption systems in the ventral skin of the Japanese tree-frog are independent of each other.

Elemental composition of mast cell granules of the rat and tree frog: X-ray Microanalysis on fresh frozen dried ultrathin sections

Electron probe x-ray microanalysis using fresh air-dried spreads revealed electrolyte elements in the granules of platelets, mast cells, pancreatic acinar cells and melanocytes. Mast cell granules of the rat and tree frog are quantitatively compared by an energy dispersive spectrometry (EDS) using fresh frozen dried ultrathin sections.Adult Wistar rats (ca. 250 g) and tree frogs (Hyla arborea japonica) of both sexes were used. Fresh frozen dried smears of rat peritoneal mast cells were prepared on the collodion-membrane covered titanium grid. Fresh frozen dried ultrathin sections of the tongue of the rat and tree frog were made by the metal contact method employing the rapid freezing apparatus (RF-2) cooled with liquid nitrogen. Ultrathin cryosections (60 nmj were cut at 163 K, transferred to the frozen specimen treating apparatus (FD-2) for freeze-drying in high vacuum at 173 K for 74-96 h. The specimens were observed first by 200kV transmission electron microscopy (TEM) and then under the scanning analytical electron microscope (X-650) for scanning transmission electron microscopy (STEM) images at an acceleration voltage of 40 kV and a specimen current of 0.2 nA.

Fine structural changes in the endolymphatic sac induced by calcium loading in the tree frog, Hyla arborea japonica.

The growth rate of calcium carbonate (CaCO3) crystals in the endolymphatic sac was modulated, and morphological changes in this organ were observed by light and electron microscopy. When calcium chloride (CaCl2) was given to the three frog for a short period (3 days to 2 weeks), CaCO3 crystal production was accelerated. Epithelial cells enlarged, their rough endoplasmic reticulum (rER) and Golgi apparatus developed, and dense material increased around CaCO3 crystals and/or in the endolymphatic lumen. In addition, multiluminal endolymphatic chambers appeared in some frogs. On the other hand, as the CaCl2 loading period lengthened and CaCO3 crystal formation decreased or stopped, the epithelial cells became flat and extended with scanty cytoplasm, and the rER and Golgi apparatus decreased in number and size. Furthermore, the amount of dense material around CaCO3 crystals and in the lumen decreased. These findings suggest that the rER, Golgi apparatus and dense material have key roles in the production of CaCO3 crystals.

Light- and electron-microscopic study of the endolymphatic sac of the tree frog, Hyla arborea japonica.

The endolymphatic sac of the tree frog and its crystals were observed by light- and electron microscopy. Scanning electron microscopy revealed that the crystals have a faceted body and two pointed ends. Light- and transmission electron microscopy revealed that the endolymphatic sac is composed of many small chambers. In their lumina, numerous "ghosts" of crystals that resulted from decalcifications were observed. The ghosts were demarcated by a linear dense material or embedded in a flocculent substance. The epithelium of the endolymphatic sac is simple squamous or cuboidal and peculiar cytoplasmic granules are found in most cells. The granules are surrounded by a limiting membrane and have varying electron density. Some granules contain a core and/or tubular structures. Vacuoles containing large ghosts are also found in the epithelial cells. These ghosts were quite similar to those in the lumen and sometimes coexist with cell debris. The fine structure of the endolymphatic sac and its crystals is discussed.

Comparative studies on the electric nature of amphibian gonadotropin

The isoelectric focusing (IEF) profiles of pituitary gonadotropic (GTH) activity in six species of adult male amphibians were examined in addition to the newt and bullfrog. The GTH activity was measured by two radioreceptor assay (RRA) systems: one with Xenopus laevis testicular homogenate as the receptor preparation, and the other with Anolis carolinensis testicular homogenate, both employing 125I-labeled rat FSH as the radioligand, or by a radioimmunoassay (RIA) system for bullfrog basic GTH (LH). The IEF profiles were classified into four groups: (1) Cynops pyrrhogaster pyrrhogaster, Onychodactylus japonicus, (2) X. laevis, (3) Rana japonica, Bufo japonicus formosus, (4) Rana nigromaculata, Hyla arborea japonica, Rana catesbeiana. In Onychodactylus japonicus (Group 1), Anolis RRA-positive peaks and Xenopus RRA-positive peaks were found in almost the same fractions. They were in the alkaline to neutral pH region. This IEF profile was very similar to that of newt pituitary GTH in July when spermatogenesis was active. In X. laevis (Group 2), the IEF profile was similar to that of urodeles. However, the most alkaline peak was found at pH 9.02, which was lower than those of urodeles. These peaks showed some differences in the reactivity to the RRA systems. In R. japonica (Group 3), the major peak obtained by Xenopus RRA was found at pH 10.27, which corresponded to the only positive peak obtained by the RIA system for bullfrog basic GTH. Furthermore, this peak was also recognized by Anolis RRA as one of the eight peaks, which appeared in the alkaline to acidic region. In B. japonicus formosus (Group 3), similar results were observed. In H. arborea japonica, R. nigromaculata and R. catesbeiana (Group 4), Anolis RRA-positive and Xenopus RRA-positive peaks were quite independent, indicating distinct separation of LH and FSH, though p Is of FSH-like GTH in the frogs were not so acidic as those of mammalian and avian FSH.

Metallothionein present or induced in the three species of frogs Bombina Orientalis, Bufo Bufo Japonicus and Hyla arborea Japonica

1. A low molecular weight copper (Cu)- and/or zinc (Zn)-binding protein was present in the livers (and kidneys) of the three species of frogs Bombina orientalis, Bufo bufo japonicus and Hyla arborea japonica. 2. Cadmium (Cd) accumulated in the livers (and kidneys) after repeated i.m. injections of Cd 2+ was mostly bound to the same metal-binding protein as Cd, Zn- or Cd, Zn, Cu-thionein. 3. Concentrations of Cd and several essential elements in the livers and kidneys of the three species of frogs were determined by inductively coupled plasma-atomic emission spectrophotometry. 4. Metallothionein in the three species of frogs consists of a single isoform both in control and Cd-injected animals.

Limb regenerative capacity of four species of Japanese frogs of the families Hylidae and Ranidae.

The regenerative capacity of limbs was investigated by amputation of limbs at the zeugopodium in postmetamorphic froglets and adults of various sizes in four species of Japanese frogs, all of which showed some regeneration at these ages. In Hyla arborea japonica and Rana brevipoda porosa most young froglets regenerated their limbs well; however, the rate of regeneration decreased with the age of amputation, and the limb became nonregenerative in adults. Limbs of adults in Rana rugosa and R. japonica, on the other hand, exhibited good regeneration. All of the regenerates in the four species were heteromorphic, consisting histologically of well-developed cartilaginous rods surrounded by connective tissue and skin. Limited development of muscle was apparent in regenerates of the three ranid species. The relations between body size, innervation of limbs, and regenerative capacity are discussed.

愛知県における壷形吸虫メタセルカリアの分布と人工感染猫における病理組織学的所見

Five species of amphibians, Rana nigromaculata, R.catesbeiana, R.rugosa, Hyla arborea japonica, and Triturus pyrrhogaster pyrrhogaster, and reptilian species, Rhabdophis tigrinus tigrinus, were collected from Toyohashi, Aichi Prefecture, from June to September, 1979, and examined for parasites.Metacercariae were found in the mesentery and intramuscular connective tissue, and identified as Pharyngostomum cordatum by morphological examination of adult flukes obtained from cats experimentally infected with those metacercariae.In these cats, the epithelium was lost from the sites of the intestine harboring the flukes.

Behavior of foreign nuclei transplanted into a Rana nigromaculata egg and DNA synthesis by them.

Several early-gastrula nuclei from Rana nigromaculata, R. brevipoda, R. japonica, R. rugosa, Hyla arborea japonica, Bombina orientalis, Xenopus laevis or Cynops pyrrhogaster were transplanted into an unfertilized or enucleated R. nigromaculata egg. Immediately after the nuclear transplantation, 3H-thymidine was injected into these eggs. The behavior of the transplanted nuclei and the incorporation of 3H-thymidine by them were examined.All the transplanted nuclei swelled remarkably in a great majority of the eggs transplanted with nuclei from each of the four Rana species or Hyla as well as in a small number of eggs transplanted with Bombina or Xenopus nuclei. There was a mixture of swollen and degenerative nuclei in a small number of eggs transplanted with Rana or Hyla nuclei as well as in many eggs transplanted with Bombina or Xenopus nuclei. In a small number of eggs transplanted with Bombina or Xenopus nuclei as well as in all the eggs transplanted with Cynops nuclei, the transplanted nuclei were all degenerative.No remarkable difference was found in the amount of 3H-thymidine incorporation by each nucleus between swollen nuclei from different species.

Studies On The Parasites Of Korean Amphibia: 1. Trematodes Of Amphibians

An investigation on the trematodes of Korean amphibians was carried out during the period from April to September of 1976. A total of 883 amphibians of 6 species were examined. These frogs, toads and salamanders were collected in the areas of the Han river side. And also these amphibians comprised 775 Rana nigromaculata, 53 Rana rugosa, 21 Hyla arborea japonica, 2 Bufo bufo gargarizans, 2 Gallula tornieri and 30 Hynobius leechi. Out of the results of this work, 7 species of trematodes belonging to 2 orders were found as follows: 1. Monogenea: Diplorchis ranae Ozaki, 1931. 2. Digenea: Diplodiscus japonicus (Yamaguti, 1936), Gorgodera japonica Yamaguti, 1936, Haematoloechus sibiricus japonicus(Yamaguti, 1936), Mesocoelium brevicaecum Ochi, 1929, Loxogenes liberum Seno,1980, Pleurogenoides japonicus (Yamaguti, 1936).

Bile salts of anura.

Bile salts of six different species of the Anura were investigated by means of thin-layer chromatography, gas-liquid chromatography, and mass spectrometry. Bile of Xenopus laevis contained taurine-conjugated and free forms of 3α, 7α, 12α-trihydroxy-5β-cholestanoic acid and 5β-cyprinol sulfate2) as its chief constituents. Smaller amounts of cholic acid and chenodeoxycholic acid were also detected in the hydrolyzed bile. Neither conjugated bile acids nor bile alcohol sulfates were found in the bile of Bombina orientalis, the chief constituents of which were free 3α, 7α, 12α-trihyboxy- and 3α, 7α, 12α, 24-tetrahydroxy-5β-cholestanoic acids. Hyla arborea japonica contained 5β-ranol sulfate as almost the only bile salt. Rana brevipoda contained taurine-conjugated and free forms of cholic acid and 3α, 7α, 12α-trihydroxy-5β-cholestanoic acid, and 5β-cyprinol sulfate. Rana rugosa had mainly 5α-cyprinol sulfate in its bile. There was also a minor amount of tauro-3α, 7α, 12α-trihydroxy-5α-cholestanoate. Rana limnocharis limnocharis contained taurine-conjugated and free forms of cholic acid and 3α, 7α, 12α-trihydroxy-5β-cholestanoic acid, and 5β-bufol sulfate.