Showing papers on "Toad published in 1995"

Pre‐MPF is absent in immature oocytes of fishes and amphibians except Xenopus

Abstract: Maturation-promoting factor (MPF), a final trigger for initiating oocyte maturation, is activated in the oocyte cytoplasm, in response to maturation-inducing hormone (MIH) secreted from follicle cells surrounding the oocyte. MPF consists of cdc2 and cyclin B. We investigated the state of cdc2 and cyclin B in immature and mature oocytes of fishes (carp, catfish and lamprey) and amphibians (Xenopus, frog [Rana] and toad [Bufo]) using monoclonal antibodies raised against mouse cdc2, which also recognize fish and amphibian cdc2, and monoclonal antibodies against goldfish cyclin B1 and polyclonal antibodies against Xenopus cyclins B1 and B2. Anti-cdc2 and anti-cyclin B immunoblotting of oocyte extracts fractionated by gel filtration chromatography showed that immature oocytes from all of these species with the exception of Xenopus contained only monomeric cdc2. Cyclin B-bound inactive cdc2 (pre-MPF) was present only in immature Xenopus oocytes. Cdc2-cyclin B complex was, however, found in mature oocytes from all the species examined. After the oocyte is induced to mature by MIH, cdc2 should therefore bind to cyclin B in all of these species, except Xenopus. These results suggest that the complex formation of cdc2 and cyclin B in response to MIH stimulation at the oocyte surface is a critical step for initiating oocyte maturation in fishes and amphibians, with the exception of Xenopus, in which pre-MPF already exists in immature oocytes and only its chemical modification is required for MPF activation.

Distribution of mRNA encoding the FA-CHIP water channel in amphibian tissues: effects of salt adaptation.

Abstract: A water channel, the frog aquaporin-CHIP (FA-CHIP) was recently cloned from Rana esculenta urinary bladder. The 28.9 kDa encoded protein shows 78.8%, 77.4%, 42.4% and 35.6% identity with rat CHIP28, human CHIP28, rat WCH-CD and γ-TIP, other members of the new transmembrane water channel family (Aquaporin-CHIP). We have now studied membranes from different frog (R. esculenta) organs employing semiquantitative PCR using FA-CHIP specific primers and an internal standard to quantify the PCR products. The FA-CHIP mRNA was abundantly expressed in the frog urinary bladder, skin, lung and gall bladder, while a lower expression was detected in the colon, liver and oviduct. FA-CHIP mRNA was not detected in the frog kidney, erythrocytes and brain but its expression was observed in the toad (Bufo arenarum) urinary bladder and skin, showing that FA-CHIP is probably a general amphibian water channel. Salt acclimation is known to increase the water permeability of frog and toad epithelia. We have now observed that salt acclimation for 1, 3, 4 or 5 days markedly increased skin and urinary bladder FA-CHIP mRNA expression. It is generally accepted that water permeability is controlled in these tissues by the rate of water channel transfer from subapical vesicles (aggrephores) to the apical membrane. Our results indicate that water permeability is also regulated at the level of the FA-CHIP transcription.

Galanin-like immunoreactive expression in the central nervous system of the toad.

Abstract: The distribution of galanin (GAL)-like immunoreactivity (-LI) was studied in the CNS of the toad (Bufo arenarum Hensel). Tissue sections were incubated with antibodies directed toward rat or porcine GAL and processed either for the avidin-biotin complex, or for the indirect immunofluorescence techniques. In the telencephalon GAL-immunoreactive (-IR) perikarya were observed in the ventral part of the striatum and in the septal accumbens nuclei. Immunopositive neurons were also observed in the medial amigdala with some intermingled cells between the fibers of the anterior commissure. Numerous GAL-IR perikarya were present along the rostrocaudal medial preoptic nucleus. Occasionally lightly immunoreactive cells were detected in the magnocellular region. The most numerous accumulation of GAL-IR cells was present in the ventral hypothalamus around the infundibular region, in the posterior tubercle and in the nucleus of the paraventricular organ. Immunostained cells were also present in the pretectal gray, solitary nucleus, gracil nucleus and in the spinal cord in the intermediate gray and in large motoneurons of the ventral horn. The widespread distribution found of GAL-LI suggests that GAL in the toad, as well as in mammalian species, may serve a variety of functions with a preponderant role in neuroendocrine processes. A role for GAL as a trophic factor in the brain of the toad is also suggested.

Drugs activating G proteins disturb cycling of ADH-dependent water channels in toad urinary bladder.

Abstract: In the toad urinary bladder, antidiuretic hormone (ADH)-mediated changes in water permeability depend on exocytic insertion and endocytic retrieval of water channels into and from the apical membrane, respectively. Because GTP-binding proteins (G proteins) are well-recognized regulators of vesicular trafficking throughout the cell, we tested the hypothesis that drugs interfering with G protein would modify the hydrosmotic response to ADH and the ADH-regulated formation of endosomes, as assessed by luminal incorporation of a fluid-phase marker [fluorescein isothiocyanate (FITC)-dextran, 70 kDa]. Mastoparan (4 microM) and compound 48/80 (poly-p-methoxyphenylethylmethylamine; 50 micrograms/ml), added to the luminal side of the toad urinary bladder, as well as AlF3 added to the serosal side (400 microM), inhibited ADH- and 8-bromoadenosine 3',5'-cyclic monophosphate-induced transepithelial water flow by > 50% and simultaneously enhanced cellular incorporation of FITC-dextran by > 200%. The pattern of FITC-dextran uptake observed using fluorescence microscopy both in scraped cells and in the intact bladder was granular, suggesting fluid-phase endocytosis. Mastoparan and AlF3, which are both probes of G proteins, increased FITC-dextran uptake only in the presence of ADH and a transepithelial osmotic gradient, i.e., under conditions where water channel-carrying endosomes presumably cycle. Therefore, we suggest that the ADH-dependent cycling of water channels could be controlled by one or more G proteins associated with the apical membrane and/or the water channel-carrying vesicles.

Application of impedance spectra to the permeability study of toad bladder

Abstract: The electrochemical impedance spectra for toad urinary bladder were abtained and we show a new easy way to seperate the apical membrane conductance from that of basolateral membrane without using microelectrode. The mucosal addition of nystatin to toad bladder presented a graeatly declined main semicircle which decreased with time. These indicated that ion transport through the apical membrane affected by nystain may include two processes in series, one of which perhaps resulted from the incorporation of nystatin into the apical membrane.