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Genetic variation in an endemic salamander, Salamandra atra, using amplified fragment length polymorphism.

The pattern of genetic differentiation of the endemic alpine salamander, Salamandra atra, has been studied using amplified fragment length polymorphism (AFLP) from 11 populations throughout the range of the two currently recognized subspecies, atra and aurorae. Five different primer combinations produced 706 bands and were analyzed by constructing a phylogenetic tree using NJ and principal component analysis. Significant genetic variation was revealed by AFLP between and within populations but, our results show a lack of genetic structure. AFLP markers seems to be unsuitable to investigate complex and recent diversification.

Animals↗

Phylogeography of the salamander genus Pseudobranchus in the southeastern United States.

This study provides an extensive set of mitochondrial cytochrome b sequences for the salamander genus Pseudobranchus of the southeastern United States. These sequences were analysed by multiple phylogenetic methods that support a single set of major phylogeographic divisions for its two corroborated species (P. axanthus and P. striatus). These phylogeographic divisions overlap with the geographic breakpoints of other freshwater and terrestrial taxa in this region. Collectively, these overlapping patterns highlight the Central Highland and Tifton/Vidalia uplands as a significant barrier to Atlantic vs. Gulf coast groups, while reconfirming the phylogeographic significance of the Altamaha and Apalachicola river drainages. Despite their distinct phylogeographic split, P. striatus from west and east of these uplands are not currently recognizable as separate species according to the concordance principles for species definition.

Animals↗

The ontogeny of feeding kinematics in a giant salamander Cryptobranchus alleganiensis: Does current function or phylogenetic relatedness predict the scaling patterns of movement?

Studies of the scaling of feeding movements in vertebrates have included three species that display both near-geometric growth and isometry of kinematic variables. These scaling characteristics allow one to examine the "pure" relationship of growth and movement. Despite similar growth patterns, the feeding movements of toads (Bufo) slow down more with increasing body size than those of bass (Micropterus), and sharks (Ginglymostoma). This variation might be due to major differences in the mechanism of prey capture; the bass and sharks use suction to capture prey in water, while the toad uses tongue prehension to capture prey on land. To investigate whether or not these different scaling patterns are correlated with differences in feeding mechanics, we examined the ontogenetic scaling of prey capture movements in the hellbender salamander (Cryptobranchus alleganiensis), which also has near-geometric growth. The hellbender suction feeds in the same general manner as the teleosts and shark, but is much more closely related to the toad. The feeding movements of the hellbender scale more similarly to the feeding movements of toads than to those of fishes or sharks, indicating that phylogenetic relatedness rather than biomechanical similarity predicts ontogenetic scaling patterns of movement.

Age Factors↗

Spectral tuning in salamander visual pigments studied with dihydroretinal chromophores.

In visual pigments, opsin proteins regulate the spectral absorption of a retinal chromophore by mechanisms that change the energy level of the excited electronic state relative to the ground state. We have studied these mechanisms by using photocurrent recording to measure the spectral sensitivities of individual red rods and red (long-wavelength-sensitive) and blue (short-wavelength-sensitive) cones of salamander before and after replacing the native 3-dehydro 11-cis retinal chromophore with retinal analogs: 11-cis retinal, 3-dehydro 9-cis retinal, 9-cis retinal, and 5,6-dihydro 9-cis retinal. The protonated Schiff's bases of analogs with unsaturated bonds in the ring had broader spectra than the same chromophores bound to opsins. Saturation of the bonds in the ring reduced the spectral bandwidths of the protonated Schiff's bases and the opsin-bound chromophores and made them similar to each other. This indicates that torsion of the ring produces spectral broadening and that torsion is limited by opsin. Saturating the 5,6 double bond in retinal reduced the perturbation of the chromophore by opsin in red and in blue cones but not in red rods. Thus an interaction between opsin and the chromophoric ring shifts the spectral maxima of the red and blue cone pigments, but not that of the red rod pigment.

Ambystoma↗

Differential modulation of light-evoked on- and off-EPSCs by paired-pulse stimulation in salamander retinal ganglion cells.

Short-term plasticity of On- and Off-EPSPs, and its potential role in regulation of signal processing was studied in salamander retinal On-Off ganglion cells by whole-cell recording. Paired-pulse light stimulation resulted in a depression of On-, and an enhancement of Off-EPSCs. Recovery from depression and enhancement was exponential and complete by 20 s. Paired-pulse enhancement, but not depression, was abolished with increasing stimulus duration. Blockade of On-EPSC by L-2-amino-4-phosphonobutyrate (AP-4), an agonist at group III mGluRs, significantly increased Off-EPSCs evoked by short (<2 s) duration conditioning light stimuli, resulting in a reversal of the paired-pulse enhancement to depression. The acetylcholinesterase inhibitor eserine reduced Off-EPSC1 and increased the ratio of enhancement. An opposite effect was observed in the presence of the nACh receptor antagonist d-tubocurarine. AP-7, an antagonist of NMDA receptors attenuated the enhancement of Off-EPSCs. In current clamp mode paired-pulse stimulation resulted in a modulation of light evoked, as well as the depolarization-induced spike firing pattern of ganglion cells. The present study suggests that paired light stimulation differently modulates On and Off EPSPs, and the light-evoked spike firing pattern of On-Off ganglion cells.

Action Potentials↗

Characterization of glutamate transporter function in the tiger salamander retina.

Glutamate transporters in the tiger salamander retina were studied by autoradiographic and intracellular recording techniques. When the retina was incubated with 15 microM L-[3H]glutamate, photoreceptors and Muller cells were labeled, indicating that these cells had high-affinity glutamate uptake transporters. A much higher dose of glutamate than kainate was required in the bath to produce the same membrane depolarization in horizontal cells (HCs), and the time course of glutamate-induced depolarization was much slower than that of the kainate-induced depolarization. Since glutamate is a substrate of glutamate transporters whereas kainate is not, we attribute these differences to the buffering of extracellular glutamate by glutamate transporters in the retina. D-aspartate (D-asp) increased the efficacy of bath-applied glutamate. Dihydrokainate (DHKA) exerted little effect on glutamate efficacy when applied alone, but it increased glutamate efficacy in the presence of D-asp. These results are consistent with the notion that glutamate transporters in Muller cells are D-asp sensitive and those in photoreceptors are DHKA and D-asp sensitive. Application of DHKA (1-2 mM) did not affect the dark membrane potential or the light responses in rods and cones, but it depolarized the HC dark membrane potential and reduced the HC peak and tail light responses. Our results suggest that DHKA-sensitive glutamate transporters in photoreceptors regulate glutamate levels in rod and cone synaptic clefts. They modulate dark membrane potential and the relative rod cone inputs in retinal HCs.

ATP-Binding Cassette Transporters↗

Organochlorine pesticides and polychlorinated biphenyls (PCBs) in eggs of red-legged frogs (Rana aurora) and northwestern salamanders (Ambystoma gracile) in an agricultural landscape.

Organochlorine pesticides were widely used in the past in Sumas Prairie, British Columbia, Canada. In the 1990s, the hatching success of amphibians from agricultural sites was significantly depressed in the Sumas Prairie compared to reference sites. Therefore, in this study, organochlorine pesticides and polychlorinated biphenyls (PCBs) were measured in eggs of red-legged frogs (Rana aurora) and northwestern salamanders (Ambystoma gracile) from the Sumas Prairie. Egg masses were sampled from three agricultural sites that were exposed to agricultural runoff, and three reference sites that had lower agricultural runoff exposures. Not only was there little difference in contamination between agricultural and reference sites, but the levels of pesticides and PCBs found were lower than the concentrations reported to cause developmental or behavioural problems in amphibians.

Agriculture↗

The detection of certain predators via olfaction by small-mouthed salamander larvae (Ambystoma texanum).

Larvae of the small-mouthed salamander (Ambystoma texanum) showed an increase in refuge use when exposed to chemical cues from a predaceous fish. The percentage of time spent outside of refuge was significantly reduced relative to controls when larvae were exposed to chemical cues from fish, but was not different from the controls when larvae were exposed to chemical cues from three other potential predators. A. texanum larvae responded to these chemical cues via olfaction. Larvae that had their external nares plugged with a gelatinous paste did not respond to fish cues, while sham-treated larvae showed an avoidance of fish cues.

Animal Communication↗

Pseudo-immunolabelling with the avidin-biotin-peroxidase complex (ABC) due to the presence of endogenous biotin in retinal Müller cells of goldfish and salamander.

Immunodetection techniques are dependent on enzyme-protein conjugates for the visualisation of antigen-antibody complexes. One of the most widely used is the avidin-biotin-peroxidase complex (ABC) method. The present study demonstrates that direct treatment of goldfish and salamander retinal sections with ABC, followed by an incubation with the chromogenic substrate 3,3-diaminobenzidine tetrahydrochloride (DAB) and H2O2, manifested a punctate staining pattern across the neural retinae, presumably through binding of avidin to endogenous biotin. Incubation with a primary antiserum against biotin followed by immunoprocessing with the peroxidase--anti-peroxidase (PAP) method showed a pattern similar to the punctuate framework as detected with solo ABC-treated sections. Moreover, the ABC-DAB/H2O2 mediated pattern corresponded to the spatial orientation of Müller cells as identified by GFAP immunostaining. These findings indicate the presence of endogenous biotin in Müller cells and calls for caution in the application of the ABC method in immunotechniques in retinal research.

Animals↗

Dihydropyridine-sensitive calcium currents in bipolar cells of salamander retina are inhibited by reductions in extracellular chloride.

Dihydropyridine-sensitive calcium currents (I(Ca)) in photoreceptors are unusual in that they can be inhibited by reductions in extracellular chloride. The present study examined whether I(Ca) in retinal bipolar cells, which as in photoreceptors mediates sustained neurotransmission, is also inhibited by reductions in chloride. Nystatin-perforated patch, whole cell recordings were obtained from bipolar cells in a retinal slice preparation of larval tiger salamander. In the presence of Ba(2+), voltage steps above -40 mV evoked sustained inward currents, which were enhanced by the dihydropyridine, (-)BayK8644, and inhibited by nisoldipine. Similar to photoreceptors, replacing Cl(-) with gluconate or CH(3)SO(4) inhibited bipolar cell I(Ca) and produced a negative shift in the current/voltage relationship. Thus, sensitivity to Cl(-) may be a more general property of L-type Ca(2+) channel subtypes that mediate sustained neurotransmission.

3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethy↗

The complete mitochondrial genome of the Chinese giant salamander, Andrias davidianus (Amphibia: Caudata).

The mitochondrial genome of the Chinese giant salamander Andrias davidianus was isolated using the long-and-accurate polymerase chain reaction (LA PCR) method. The sequencing work adopted the shotgun strategy accompanying with seven internal primers to cover the gaps where overlapping clones were not available. The entire mtDNA sequence is 16,503 bp long, with a gene content of 13 protein-coding, two ribosomal RNA and 22 transfer RNA genes, and order identical to that observed in most other vertebrates except for an additional 318 bp non-coding sequence between tRNA-Thr and tRNA-Pro genes. In order to carry out molecular phylogenetic analyses, all 13 protein sequences deduced from whole mitochondrial genomes for eight vertebrate species (six amphibians, two lobe-finned fishes) were combined to a single data set. This data set was refined by the program Gblocks using a stringent parameter setting and then subjected to MP, ML and NJ analyses. Thus phylogenetic relationships among living amphibians were discussed.

Animals↗

Population-specific behavioral electrosensitivity of the European blind cave salamander, Proteus anguinus.

In nine salamanders from different Slovenian populations of the urodele Proteus anguinus, including three specimens of its 'black' variety, P anguinus parkelj, thresholds of an overt avoidance response to electrical field stimuli were estimated as a function of frequency (continuous sine-waves in water). Thresholds down to 0.3V/cm (ca 100 nA/cm2) and up to 2 mV/cm (670 nA/cm2), at 'best frequencies' of around 30 Hz were found. Sensitivity covered a total frequency range of below 1 Hz, excluding DC, up to 1-2 kHz with up to 40 dB higher thresholds. Thresholds and tuning curves are compared with those of a Proteus population raised in captivity for more than 35 years. The biological significance and the apparently still ongoing evolution of the electrical sense in urodeles, ie in the genus Proteus, are interpreted in terms of comparative sensory physiology and ethological ecology as a result of more recent evolutionary diversification during and since glaciation in the Pleistocene.

Animals↗

Teeth and their sex-dependent dimorphic shape in three species of Costa Rican plethodontid salamanders (Amphibia: Urodela).

The shape of the teeth and their sex-dependent dimorphic expression in three species of Costa Rican plethodontids (Bolitoglossa subpalmata, Oedipina uniformis and Nototriton abscondens) were studied using light microscopy and scanning electron microscopy. The teeth of the vomerine tooth patches are about one third larger than the teeth of the jaws in B. subpalmata and O. uniformis, whereas all teeth of N. abscondens are of about uniform size. The occurrence of bicuspid tooth germs in the fetus proves that primary teeth are bicuspid in these directly developing plethodontids. Females possess only bicuspid teeth consisting of a pedicel and a crown, as is considered characteristic for urodeles after metamorphosis. Adult males possess conical monocuspid teeth on the premaxillary. These teeth--which are similar to the typical late larval tooth of salamanders presenting a larval stage--are about twice as big as the neighbouring bicuspid maxillary teeth. N. abscondens males possess some monocuspid teeth and teeth of aberrant shapes on the premaxillary and the maxillaries. A tendency to build more monocuspid teeth in the premaxillary region than in the maxillary region can be observed in this species. We suppose that different degrees of sensitivity to androgens in each section of the dental lamina of the upper jaw cause the secondary occurrence of conical monocuspid teeth predominantly on the premaxillary section.

Animals↗

The structure and development of the skull of Costa Rican plethodontid salamanders (Amphibia: Urodela).

We describe the structure of the skulls of the Costa Rican plethodontid salamanders Bolitoglossa subpalmata, Oedipina uniformis and Nototriton abscondens, and the characteristic sequences of development and ossification of the bony elements resulting from direct development using mainly cleared and stained specimens. Significant differences between the species studied are observed. N. abscondens possesses the broadest premaxillary pars dentalis and O. uniformis the narrowest one. The premaxillary dorsal processes are fused over their rostral third only in B. subpalmata; over half their extention in N. abscondens and almost completely in O. uniformis. A prefrontal is always present in N. abscondens; it is hidden underneath the nasal or missing in B. subpalmata, and it is always absent in O. uniformis. The skull bones, with the exception of the orbitosphenoid, develop and ossify sequentially from caudal to rostral in these directly developing species. A more massive pars dentalis of a generally narrower premaxillary are found as typical characters in males.

Aging↗

The skull structure of six species of Mesoamerican plethodontid salamanders (Amphibia, Urodela).

The skulls of six species of plethodontid salamanders of the genus Bolitoglossa from Costa Rica and Panama, Bolitoglossa colonnea, B. dofleini, B. lignicolor, B. marmorea, B. schizodactyla and B. striatula are comparatively analysed. All species are terrestrial or slightly arboreal and show no life-mode-dependent skull characteristics. Heads cleared by transparent preparation and stained in toto were used for examination of the skull structure and paraffin sections of the gonads were prepared to confirm sexual maturity of each individual. Focussing on the size of the premaxillary pars dentalis, the degree of fusion of the processus dorsales praemaxillares and the presence or absence of the paired prefrontals we state that B. dofleini and B. striatula possess more ancestral characteristics, B. marmorea is situated in an intermediate position and B. colonnea, B. lignicolor and B. schizodactyla show more advanced characteristics.

Animals↗

An anatomical investigation of the muscles of the pelvic outlet in Japanese giant salamander (Cryptobranchidae Megalobatrachus japonicus) with special reference to their nerve supply.

Four pelvic halves from two Japanese giant salamanders (Cryptobranchidae Megalobatrachus japonicus, one male and one female) were dissected in order to obtain detailed morphological data on the lumbosacral plexus and the muscles of the pelvic outlet. According to the positions of the passage of the metazonal nerves, the nerves can be divided into three groups: 1) the nerves passing dorsal to the caudofemoralis muscle, 2) the nerves passing between the caudofemoralis and the caudoischiadicus, 3) the nerves passing ventral to the caudoischiadicus. In consideration of the sites of origin from the lumbosacral plexus, a three-way stratificational analysis can be made, group 1 arising craniodorsal to group 2, and group 3 caudoventral to group 2. It is suggested that the three pelvic outlet muscles, the caudofemoralis, caudocruralis, and caudoischiadicus, have derived from the ventral muscles of the posterior limb, and have extended ventrocaudalwards.

Animals↗

The phylogenetics of Desmognathine salamander populations across the southern Appalachians.

Salamanders in the genus Desmognathus (Caudata: Plethodontidae) are distributed along an aquatic to terrestrial habitat gradient in the southern Appalachian Mountains. The spatial distribution of species is believed to have formed as aquatic ancestors displaced lineages by competition and predatory interactions into less optimal terrestrial habitats. Aquatic and terrestrial species may also display different patterns of genetic diversity due to the differing likelihood of gene flow via aquatic corridors. To determine whether phylogenetic patterns were consistent with these hypotheses, we sequenced portions of the cytochrome oxidase I and 12S rRNA genes of the mitochondrial genome from 96 individuals belonging to 10 species in the genus Desmognathus. In addition, we combined our dataset with an earlier published dataset for the 12S rRNA genes. The order of species divergence is consistent with aquatic ancestors having displaced taxa into more terrestrial habitats, but the major lineages within the genus Desmognathus arose suddenly, and therefore, the specific sequence of events is not well resolved. The phylogenetic analyses among species suggest that direct-development and a terrestrial lifestyle are ancestral in the genus Desmognathus, but the degree of adult terrestriallity is labile, with some species having re-invaded terrestrial habitats. We present evidence of a clade of Desmognathus quadramaculatus from North Carolina that is distinct from the D. quadramaculatus/Desmognathus marmoratus clade. Within species, estimates of Tajima's D and Fu and Li's statistics suggest the species experienced population expansions at different times in the past. Current levels of sequence diversity in northern populations, therefore, reflect different arrival times, and hence, differences in the opportunity for among population divergence. The recent arrival of most species over large portions of their geographic ranges suggests that most extant communities have been assembled, a posteriori, by the recent assortment of species along the aquatic to terrestrial gradient according to their ecologies.

Animals↗

Hepatic metallothioneins in two neotenic salamanders, Proteus anguinus and Necturus maculosus (Amphibia, Caudata).

The presence of metallothionein (MT) and the subcellular distribution of copper, zinc and cadmium were investigated in livers of two neotenic salamanders, Proteus anguinus and Necturus maculosus. In P. anguinus, caught in the wild, hepatic MTs were present as a single isoform of (Zn, Cu, Cd)-thioneins, whose molecular weight was estimated to be approximately 12000 by size exclusion chromatography. The percentage of zinc and cadmium was higher in the cytosol and of copper in the pellet. Cytosolic cadmium was almost exclusively associated with MTs (80%), while zinc and copper were also present in the regions of higher-molecular weight proteins. In laboratory bred N. maculosus, MTs were isolated from the liver cytosol and extract of the pellet as (Cu, Zn)- and (Zn, Cu)-thioneins, respectively. According to the low amount of copper extracting from liver pellets of N. maculosus, the presence of water insoluble aggregated forms of Cu-thioneins should be checked in further investigations.

Animals↗