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PubMed1997

A cyclic nucleotide-dependent chloride conductance in olfactory receptor neurons.

Whole-cell membrane currents were recorded from olfactory receptor neurons from the neotenic salamander Necturus maculosus. Cyclic nucleotides, released intracellularly by flash photolysis of NPE-caged cAMP or NPE-caged cGMP, activated a transient chloride current. The chloride current could be elicited at constant voltage in the absence of extracellular Ca2+ as well as in the presence of 3 mM intracellular Ca2+, suggesting that the current did not require either voltage or Ca2+ transients for activation. The current could be elicited in the presence of the protein kinase inhibitors H-7 and H-89, and in the absence of intracellular ATP, indicating that activation was independent of protein kinase A activity. These results suggest that Necturus olfactory receptor neurons contain a novel chloride ion channel that may be directly gated by cyclic nucleotides.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗
PubMed2004

Impact of ammonium nitrate on growth and survival of six European amphibians.

We conducted static experiments to assess the effects of ammonium nitrate fertilizer on embryos and larvae of six European amphibians: sharp-ribbed salamander (Pleurodeles waltl), Iberian painted frog (Discoglossus galganoi), western spadefoot toad (Pelobates cultripes), common toad (Bufo bufo), natterjack toad (Bufo calamita), and common tree frog (Hyla arborea). Embryos were exposed to different and environmentally relevant concentrations of ammonium nitrate (0 to 200 mg NO(3)(-)/L) for 15 days. Hatching took place during the experiments. H. arborea was extremely sensitive and had high mortality after 8 days of exposure even at the lower fertilizer levels. D. galganoi and B. bufo were also very sensitive and had significant mortality after 15 days of exposure. The rest of the species did not suffer lethal effects but suffered abnormalities or decreased growth at the highest fertilizer concentrations. Chemical fertilizers or manure could have contributed to the observed decrease of B. bufo and D. galganoi in agricultural areas of the Iberian Peninsula during recent years. H. arborea was the most sensitive species studied. The results of our study showed that environmentally relevant levels of ammonium nitrate can induce mortality and might affect population dynamics of this species in agricultural environments.

Amphibians↗
PubMed2004

Effects of chlorinated solvents on four species of North American amphibians.

Tetrachloroethylene (PCE), a dry cleaning and degreasing solvent, can enter groundwater through accidental leaks or spills, and concentrations as high as 75 mg/L have been reported in Canadian aquifers. Amphibians in wetlands receiving contaminated groundwater may be exposed to PCE and its degradation products, but little information is available on the impacts of these compounds on indigenous amphibian species. Acute (96-h static renewal) exposures to PCE and its major degradation products, trichloroethylene (TCE) and cisand trans-dichloroethylene, were conducted on embryos of four North American amphibian species: wood frogs (Rana sylvatica), green frogs (R. clamitans), American toads (Bufo americanus), and spotted salamanders (Ambystoma maculatum). Subsequently, chronic exposures to PCE and TCE were conducted with the larvae of American toads. Both PCE and TCE were teratogenic to amphibian embryos; median effective concentrations (EC50s) for developmental deformities produced by PCE and TCE exposure for wood frogs and green frogs were 12 and 40 mg/L, respectively. Embryonic survivorship, however, was not compromised at these concentrations. American toads were less sensitive; the EC50 for developmental abnormalities was not attained at the highest test concentrations, 45 and 85 mg/L PCE and TCE, respectively. These results are pertinent in assessing the impact of groundwater pollution on an aquifer-fed wetland.

Amphibians↗
PubMed2002

Sequencing and expression of the CD3 gamma/delta mRNA in Pleurodeles waltl (urodele amphibian).

The CD3 complex is an essential component of the T-cell receptor (TCR) implicated in T-cell maturation and activation. This TCR has been identified in both cartilaginous and bony vertebrates. In different studies where the CD3 chains were cloned and sequenced, it appeared that the CD3 complex is composed of several chains, all susceptible to phosphorylation and able to transduce signals. Here, by an approach combining degenerative oligonucleotide primers and RACE-PCR, we report the cloning and sequencing of a CD3 cDNA from the salamander Pleurodeles waltl, highly homologous to the Xenopus and chicken CD3 gamma/delta cDNAs. Using semi-quantitative PCR and Northern blot analysis, we found the highest CD3 gamma/delta mRNA expression in the thymus; weaker expression was observed in the spleen and blood, followed by the intestine, therefore confirming the tissue and lymphoid specificities of this mRNA. The signals in the spleen, blood and intestine represented 55%, 33% and 16%, respectively, of the signal detected in the thymus. During the embryonic and larval stages of Pleurodeles waltl development, CD3 gamma/delta mRNA expression begins early at the neurula stage (stage 15, 69 h after laying), increases up to stage 33 (9 days after laying) and afterwards remains stable, at least until the larval stage 42 (28 days after laying). As the thymus primordium appears much later, the question of the formation and maturation of the first T-cell precursors outside this organ is posed.

Amino Acid Sequence↗
PubMed2001

The recombination-activating gene 1 of Pleurodeles waltl (urodele amphibian) is transcribed in lymphoid tissues and in the central nervous system.

The recombination-activating gene 1 (RAG1) product is required for the somatic rearrangement of immunoglobulin and T-cell receptor genes. We cloned and sequenced the large continuous open reading frame coding for the salamander Pleurodeles waltl RAG1 protein. Semi-quantitative RT-PCR experiments were performed to quantify the expression of RAG1 in different tissues. The strongest signal was observed in the thymus of juvenile animals, confirming the primary lymphoid nature of that organ. Weaker expression was observed in the spleen, brain, and eyes of adults. Signals in these tissues represented 5.5%, 4.6%, and 2.0%, respectively, of the signal detected in the thymus. Expression in brain was confirmed by in situ hybridization. Similarly, low amounts of RAG1 transcripts were previously detected in the mouse brain. Moreover, the transcription of RAG1 begins as early as the neurula stages of development. These data suggest that the RAG1 protein could play a role in the central nervous system of vertebrates.

Amino Acid Sequence↗
PubMed1998

Wide tissue distribution of axolotl class II molecules occurs independently of thyroxin.

Unlike most salamanders, the Mexican axolotl (Ambystoma mexicanum) fails to produce enough thyroxin to undergo anatomical metamorphosis, although a "cryptic metamorphosis" involving a change from fetal to adult hemoglobins has been described. To understand to what extent the development of the axolotl hemopoietic system is linked to anatomical metamorphosis, we examined the appearance and thyroxin dependence of class II molecules on thymus, blood, and spleen cells, using both flow cytometry and biosynthetic labeling followed by immunoprecipitation. Class II molecules are present on B cells as early as 7 weeks after hatching, the first time analyzed. At this time, most thymocytes, all T cells, and all erythrocytes lack class II molecules, but first thymocytes at 17 weeks, then T cells at 22 weeks, and finally erythrocytes at 26-27 weeks virtually all bear class II molecules. Class II molecules and adult hemoglobin appear at roughly the same time in erythrocytes. These data are most easily explained by populations of class II-negative cells being replaced by populations of class II-positive cells, and they show that the hemopoietic system matures at a variety of times unrelated to the increase of thyroxin that drives anatomical metamorphosis. We found that administration of thyroxin during axolotl ontogeny does not accelerate or otherwise affect the acquisition of class II molecules, nor does administration of drugs that inhibit thyroxin (sodium perchlorate, thiourea, methimazole, and 1-methyl imidazole) retard or abolish this acquisition, suggesting that the programs for anatomical metamorphosis and some aspects of hemopoietic development are entirely separate.

Ambystoma↗
PubMed2002

A GIS-based approach for assessing the regional conservation status of genetic diversity: an example from the southern Appalachians.

Conserving genetic diversity requires an assessment of the distribution of genetic variants in relation to patterns of land use and environmental variation at a regional scale. This assessment requires a novel approach to integrating and analyzing the genetic and environmental data across spatial scales. To explore the integration of genetic data with other geospatial data sets, we developed a GIS-based approach for examining patterns of genetic diversity for several species of salamanders in southern Appalachians. The genetic data, from allozyme surveys in the genetics literature, were integrated into a GIS database along with related attributes including population identifications and spatial locations. Using existing geospatial data, we classified sample locations as being either protected from anthropogenic disturbance (e.g., National Parks, Wilderness Areas) or as unprotected (e.g., private lands, multiple-use lands in National Forests). We used multidimensional scaling of allelic frequencies and contributions of populations to interpopulation differences in allelic richness to determine which populations had genetic characteristics most different from other populations in the sample. Measures of genetic differentiation were integrated into the GIS database to facilitate spatial analysis and visualization of the indices in relation to land use. This approach was useful for both identification of populations with components of genetic variation that were not well represented at protected sites and for identifying areas of species distributions where more genetic sampling would be necessary to make informed management decisions. Our approach could be readily adapted for use by managers and geneticists working with other species and types of genetic markers.

Animals↗
PubMed2001

Object segmentation model: analytical results and biological implications.

A simple, biologically motivated neural network for segmentation of a moving object from a visual scene is presented. The model consists of two parts: an object selection model which employs a scaling approach for receptive field sizes, and a subsequent network implementing a spotlight by means of multiplicative synapses. The network selects one object out of several, segments the rough contour of the object, and encodes the winner object's position with high accuracy. Analytical equations for the performance level of the network, e.g., for the critical distance of two objects above which they are perceived as separate, are derived. The network preferentially chooses the object with the largest angular velocity and the largest angular width. An equation for the velocity and width preferences is presented. Additionally it is shown that for certain neurons of the model, flat receptive fields are more favourable than Gaussian ones. The network exhibits performances similar to those known from amphibians. Various electrophysiological and behavioral results--e.g., the distribution of the diameters of the receptive fields of tectal neurons, of the tongue-projecting salamander Hydromantes italicus and the range of optimal prey velocities for prey catching--can be understood on the basis of the model.

Animals↗
PubMed2002

Cell cycle reentry of ventricular and atrial cardiomyocytes and cells within the epicardium following amputation of the ventricular apex in the axolotl, Amblystoma mexicanum: confocal microscopic immunofluorescent image analysis of bromodeoxyuridine-labeled nuclei.

To understand how to reinitiate cell division in adult human myocardium, a heart regeneration model was examined in the amphibian axolotl salamander, Amblystoma mexicanum. The ventricular apex was surgically amputated and resected for 3 weeks. At 14 days of recovery, the thymidine analog 5-bromo-2'-deoxyuridine (BrdU) was injected intraperitoneally to identify cell types undergoing S-phase by indirect immunofluorescence using primary anti-BrdU antibodies. This is the first report showing a concentrated area of cell division in the ventricle and cells throughout the atrial epicardium by confocal microscopic image analysis in response to wounding of the ventricle. Tissues coimmunostained with anti-BrdU and sarcomeric myosin-specific MF20 antibodies showed 12.8 +/- 4.10% of myocytes contained BrdU(+) nuclei in a 75 microm to 750 microm zone in the ventricular myocardium subjacent to the amputation plane. BrdU(+) nuclei also were present in newly formed ventricular epicardium that surround dividing myocytes, and in epicardial mesothelium (74.3 +/- 6.36 %) and connective tissue (44.9 +/- 13.31%) cells distal to the wound. Unexpectedly, immunofluorescent BrdU(+) nuclei were observed in isolated atrial myocytes (13.9 +/- 1.45%) and in uninjured atrial epicardial mesothelium (64.3 +/- 1.55%) and connective tissue (29.4 +/- 5.50%). No BrdU(+) nuclei were present in cardiomyocytes or epicardium from sham-operated and BrdU-treated controls. These results suggest that renewed cell division is a specific response to wounding of the ventricle. Additionally, release of a growth factor may be responsible for the specific localized mitotic ventricular cardiomyocyte response adjacent to the wound, and the more generalized atrial proliferative response distal to the amputation.

Ambystoma mexicanum↗
PubMed2006

Enameloid/enamel transition through successive tooth replacements in Pleurodeles waltl (Lissamphibia, Caudata).

Study of the evolutionary enameloid/enamel transition suffers from discontinuous data in the fossil record, although a developmental enameloid/enamel transition exists in living caudates, salamanders and newts. The timing and manner in which the enameloid/enamel transition is achieved during caudate ontogeny is of great interest, because the caudate situation could reflect events that have occurred during evolution. Using light and transmission electron microscopy, we have monitored the formation of the upper tooth region in six successive teeth of a tooth family (position I) in Pleurodeles waltl from late embryos to young adult. Enameloid has only been identified in embryonic tooth I(1) and in larval teeth I(2) and I(3). A thin layer of enamel is deposited later by ameloblasts on the enameloid surface of these teeth. From post-metamorphic juvenile onwards, teeth are covered with enamel only. The collagen-rich enameloid matrix is deposited by odontoblasts, which subsequently form dentin. Enameloid, like enamel, mineralizes and then matures but ameloblast participation in enameloid matrix deposition has not been established. From tooth I(1) to tooth I(3), the enameloid matrix becomes ever more dense and increasingly comes to resemble the dentin matrix, although it is still subjected to maturation. Our data suggest the absence of an enameloid/enamel transition and, instead, the occurrence of an enameloid/dentin transition, which seems to result from a progressive slowing down of odontoblast activity. As a consequence, the ameloblasts in post-metamorphic teeth appear to synthesize the enamel matrix earlier than in larval teeth.

Animals↗
PubMed2005

Dose and host characteristics influence virulence of ranavirus infections.

Parasites play a prominent role in the ecology, evolution, and more recently, conservation of many organisms. For example, emerging infectious diseases, including a group of lethal ranaviruses, are associated with the declines and extinctions of amphibians around the world. An increasingly important basic and applied question is: what controls parasite virulence? We used a dose-response experiment with three laboratory-bred clutches of tiger salamander larvae (Ambystoma tigrinum) to test how the size of inoculum and host genetic factors influence the dynamics and outcome of ranavirus infections. We found that infection rates increased with dose and were strongly affected by clutch identity and host life history stage. Case mortality increased with dose of inoculum, but was unaffected by host characteristics. Average survival time decreased with dose and differed among clutches, but this was largely due to differences in the time to onset of symptoms. Overall, our results suggest that dose of inoculum and host characteristics (life history stage and genetic background) influence the establishment and early virus replication, and therefore the virulence of ranavirus infections.

Animals↗
PubMed2006

Density of an intraguild predator mediates feeding group size, intraguild egg predation, and intra- and interspecific competition.

Intraguild predation (IGP) is common in most communities, but many aspects of density-dependent interactions of IG predators with IG prey are poorly resolved. Here, we examine how the density of an IG predator can affect feeding group size, IG egg predation, and the growth responses of IG prey. We used laboratory feeding trials and outdoor mesocosm experiments to study interactions between a social intraguild predator (larvae of the wood frog; Rana sylvatica) and its prey (spotted salamander; Ambystoma maculatum). Larvae of R. sylvatica could potentially affect A. maculatum by consuming shared larval food resources or by consuming eggs and hatchlings. However, successful egg predation requires group feeding by schooling tadpoles. We established from five to 1,190 hatchlings of R. sylvatica in mesocosms, then added either 20 A. maculatum hatchlings to study interspecific competition, or a single egg mass to examine IGP. Crowding strongly suppressed the growth of R. sylvatica, and IGP was restricted to the egg stage. In the larval competition experiment, growth of A. maculatum was inversely proportional to R. sylvatica density. In the predation experiment, embryonic mortality of A. maculatum was directly proportional to the initial density of R. sylvatica and the mean number of tadpoles foraging on egg masses. IGP on eggs reduced A. maculatum hatchling density, which accelerated larval growth. Surprisingly, the density of R. sylvatica had no overall effect on A. maculatum growth because release from intraspecific competition via egg predation was balanced by increased interspecific competition. Our results demonstrate that the density of a social IG predator can strongly influence the nature and intensity of interactions with a second guild member by simultaneously altering the intensity of IGP and intra- and interspecific competition.

Ambystoma↗
PubMed2006

Isotopic evidence for methane-based chemosynthesis in the Upper Floridan aquifer food web.

Anecdotal observations of the Dougherty plain cave crayfish (Cambarus cryptodytes), the Georgia blind cave salamander (Haideotriton wallacei), and albinistic isopods (Caecidotea sp.) at great depths below the land surface and distant from river corridors suggest that obligate aquifer-dwelling (troglobitic) organisms are widely distributed throughout the limestone Upper Floridan aquifer (UFA). One mechanism by which subterranean life can proliferate in an environment void of plant productivity is through a microbial food web that includes chemosynthesis. We examined this possibility in the UFA by measuring the isotopic composition ((13)C, (14)C, and (15)N) of tissues from troglobitic macrofauna. Organisms that were captured by cave divers entering into spring conduits had delta(13)C values that suggested plant matter as a primary food resource (cave crayfish, -24.6 +/- 2.7 per thousand, n = 9). In contrast, delta(13)C values were significantly depleted in organisms retrieved from wells drilled into areas of the UFA remote from spring and sinkhole conduits (cave crayfish -34.7 +/- 9.8 per thousand, n = 10). Depleted (13)C values in crayfish were correlated with radiocarbon (Delta(14)C) depletion relative to modern values. The results suggest that methane-based microbial chemosynthetic pathways support organisms living in the remote interior of the aquifer, at least in part.

Alabama↗
PubMed2002

Ranaviruses (family Iridoviridae): emerging cold-blooded killers.

Although possessing novel replicative and structural features, the family Iridoviridae has not been as extensively studied as other families of large, DNA-containing viruses (e.g., poxviridae and herpesviridae). This oversight most likely reflects the inability of iridoviruses to infect mammals and birds, and their heretofore low pathogenicity among cold-blooded animals and invertebrates. In fact, the original frog virus isolates (e.g., frog viruses 1-3) would likely have been considered orphan viruses since they were isolated from apparently healthy frogs. However, recent disease outbreaks among commercially and recreationally important fish, cultured and wild frogs, and endangered salamanders has challenged this benign view and have implicated several members of the genus Ranavirus as pathogens. This review explores three facets of ranavirus biology. In the first the salient features of ranavirus replication are summarized using frog virus 3 as a model. Secondly, criteria for characterizing new ranavirus isolates, based on biochemical (viral protein profiles, DNA restriction fragment length polymorphisms, and nucleotide sequence analysis), ecological (host range, tissue tropism), and clinical considerations, are detailed. Lastly, the principal agents of ranavirus-mediated disease and immune responses to these viruses are discussed. In light of the above, it is clear that ranaviruses are no longer orphan viruses, and that they have a significant impact on diverse populations of ectothermic animals.

Amphibians↗
PubMed2005

Testing population genetic structure using parametric bootstrapping and MIGRATE-N.

We present a method for investigating genetic population structure using sequence data. Our hypothesis states that the parameters most responsible for the formation of genetic structure among different populations are the relative rates of mutation (micro) and migration (M). The evolution of genetic structure among different populations requires rates of M << p because this allows population-specific mutation to accumulate. Rates of micro << M will result in populations that are effectively panmictic because genetic differentiation will not develop among demes. Our test is implemented by using a parametric bootstrap to create the null distribution of the likelihood of the data having been produced under an appropriate model of sequence evolution and a migration rate sufficient to approximate panmixia. We describe this test, then apply it to mtDNA data from 243 plethodontid salamanders. We are able to reject the null hypothesis of no population structure on all but smallest geographic scales, a result consistent with the apparent lack of migration in Plethodon idahoensis. This approach represents a new method of investigating population structure with haploid DNA, and as such may be particularly useful for preliminary investigation of non-model organisms in which multi-locus nuclear data are not available.

Animals↗
PubMed1975

Sialic acid, electrophoretic mobility and transmembrane potentials of the Amphiuma red cell.

Red cells from the giant salamander Amphiuma means are shown to contain sialic acid. The amount removed by the action of neuraminidase is equal to that released by acid hydrolysis, indicating that all of the sialic acid is present on the outer surface of the plasma membrane. These cells have a negative electrophoretic mobility and 100% enzymatic removal of sialic acid results in a 40% reduction in the mobility, suggesting that either a fraction of the sialic acid carboxyl groups are unavailable to the action of external electric fields, or other negatively charged groups contribute to the surface charge. A further reduction in mobility of normal and sialic acid-free cells is caused by an increased extracellular calcium concentration. The negative groups affected by calcium are most likely to be phosphate groups, since the isoelectric point of the cells is found to lie between the pK values for H2PO-4 groups and the carboxyl groups of sialic acid. Membrane potentials of single cells, from which 80% or more of the total sialic acid had been removed, were identical to those measured in normal cells, confirming that sialic acid plays little, if any, direct role in the maintenance of membrane potentials and ionic permeabilities.

Animals↗
PubMed1976

Effect of antihistamines and chlorpromazine on the calcium-induced hyperpolarization of the Amphiuma red cell membrane.

1. It has previously been demonstrated that an increase in extracellular Ca2+ conce-tratio- induces a trandient increase in K+ permeability and associated hyperpolarization of the red cell membrane of the giant salamander, Amphiuma meand. This phenomenon is analogous to the Ca2+-induced KCl loss observed in ATP-depleted human red cells and red cell ghosts. 2. Histamine, which enhances the Ca2+-induced K+ loss from depleted human red cells, is without effect on this Ca2+-induced hyperpolarization of Amphiuma red cells. 3. Promethazine (10 muM) and mepyramine (1 mM), which inhibit the Ca2+-induced K+ loss in depleted human red cells, also block the Ca2+-related hyperpolarization of Amphiuma erythrocytes. 4. Chlorpromazine (25 muM), despite being a weak antihistamine, is equally effective in blocking the Ca2+-induced hyperpolarization of Amphiuma red cells. 5. Ionophore A23187 causes a large and sustained Ca2+/K+-dependent hyperpolarization even in the presence of normal (1.8 mM) concentrations of Ca2+. This hyperpolarization is relatively insensitive to chlorpromazine and promethazine. 6. The inhibition of the Ca2+-induced hyperpolarization of the Amphiuma red cell membrane by chlorpromazine and promethazine may berelated to their properties as local anaesthetics.

Animals↗
PubMed1981

Evolution of inner-ear auditory apparatus in the frog.

The morphology of typical anuran amphibian papillae is thoroughly distinct from that of urodeles. However, the morphological discontinuity lies not between the frogs and the salamanders, but between the most primitive living frog, Ascaphus truei, and the more derived anurans. Three features distinguishing the papillae of more derived anurans from that of Ascaphus apparently provide peripheral tonotopy in the former. The adaptive significance of a fourth feature, kinociliary bulbs, is not clear.

Animals↗