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The thermal dependence of locomotor performance and muscle contractile function in the salamander Ambystoma tigrinum nebulosum.

The thermal dependence of locomotor performance and in vitro muscle mechanical properties were studied after acclimation at 10 degrees and 20 degrees C in the salamander Ambystoma tigrinum nebulosum Hallowell. Performance measurements included burst and endurance capacities on land and in water. No significant differences in locomotor performance or muscle contractile properties were found between acclimation groups. Locomotor performance had only a slight thermal dependence, with Q10 values of 0.99-1.36 for running and swimming burst capacities (i.e. maximum speed and leg/tail cycling frequency). Running and swimming endurance capacities had thermal ratios of 1.58-1.66. Thermal dependence of muscle contractile rates was higher than that of locomotor performance: rates of force development for both isometric twitch and tetanus and maximal shortening velocity had Q10 values of 1.89-2.01. Maximal power output was also thermally dependent (Q10 = 2.33) and occurred at 38% of maximal (tetanic) force output. Force-generating capacities in isometric twitch and tetanus were relatively temperature-independent.

Acclimatization↗

Blood flow patterns in the salamander, Ambystoma tigrinum, before, during and after metamorphosis.

The patterns of blood flow through the complex circulation of the tiger salamander, Ambystoma tigrinum, were investigated during aquatic normoxia and hypoxia by application of the microsphere technique. The distribution of differently labelled microspheres injected into the bloodstream towards the left and right atria, respectively, was used to evaluate the role of the ductus arteriosus in lung perfusion before, during and after metamorphosis, as well as the general contribution of right and left atrial outputs to the blood flow in gills and lungs in neotenic and postmetamorphic animals. The distribution patterns of radioactive microspheres among pulmonary, branchial and systemic tissues indicated that the ductus arteriosus is the major pulmonary perfusion pathway in neotenic and metamorphosing animals, whereas after metamorphosis the main perfusion pathway is down the entire length of the pulmonary artery. In neotenes, the ductus arteriosus becomes even more important during aquatic hypoxia. The anterior branchial arches receive blood richer in pulmonary venous blood than the posterior arches. Approximately 26% of left atrial output and 36% of right atrial output perfuses the brachial respiratory lamellae during normoxia in neotenes. Severe aquatic hypoxia appears to increase the fraction of cardiac output flowing to the lung and decrease the fraction flowing into the first branchial arch in neotenes. This decrease into the first arch may facilitate lung perfusion and also reduce branchial O2 loss. In postmetamorphic animals, approximately 55% of right atrial output and 32% of left atrial output is directed to the lungs. The flow patterns in postmetamorphic animals remain unaffected by aquatic hypoxia.

Ambystoma↗

Anguilliform locomotion in an elongate salamander (Siren intermedia): effects of speed on axial undulatory movements

Many workers interested in the mechanics and kinematics of undulatory aquatic locomotion have examined swimming in fishes that use a carangiform or subcarangiform mode. Few empirical data exist describing and quantifying the movements of elongate animals using an anguilliform mode of swimming. Using high-speed video, I examine the axial undulatory kinematics of an elongate salamander, Siren intermedia, in order to provide data on how patterns of movement during swimming vary with body position and swimming speed. In addition, swimming kinematics are compared with those of other elongate vertebrates to assess the similarity of undulatory movements within the anguilliform locomotor mode. In Siren, most kinematic patterns vary with longitudinal position. Tailbeat period and frequency, stride length, Froude efficiency and the lateral velocity and angle of attack of tail segments all vary significantly with swimming speed. Although swimming speed does not show a statistically significant effect on kinematic variables such as maximum undulatory amplitude (which increases non-linearly along the body), intervertebral flexion and path angle, examination of the data suggests that speed probably has subtle and site-specific effects on these variables which are not detected here owing to the small sample size. Maximum lateral displacement and flexion do not coincide in time within a given tailbeat cycle. Furthermore, the maximum orientation (angle with respect to the animal's direction of forward movement) and lateral velocity of tail segments also do not coincide in time. Comparison of undulatory movements among diverse anguilliform swimmers suggests substantial variation across taxa in parameters such as tailbeat amplitude and in the relationship between tailbeat frequency and swimming speed. This variation is probably due, in part, to external morphological differences in the shape of the trunk and tail among these taxa.

Journal Article↗

Behavioural, physiological and metabolic responses to long-term starvation and refeeding in a blind cave-dwelling (Proteus anguinus) and a surface-dwelling (Euproctus asper) salamander.

The effects of long-term starvation and subsequent refeeding on haematological variables, behaviour, rates of oxygen consumption and intermediary and energy metabolism were studied in morphologically similar surface- and cave-dwelling salamanders. To provide a hypothetical general model representing the responses of amphibians to food stress, a sequential energy strategy has been proposed, suggesting that four successive phases (termed stress, transition, adaptation and recovery) can be distinguished. The metabolic response to prolonged food deprivation was monophasic in the epigean Euproctus asper (Salamandridae), showing an immediate, linear and large decrease in all the energy reserves. In contrast, the hypogean Proteus anguinus (Proteidae) displayed successive periods of glucidic, lipidic and finally lipido-proteic-dominant catabolism during the course of food deprivation. The remarkable resistance to long-term fasting and the very quick recovery from nutritional stress of this cave organism may be explained partly by its ability to remain in an extremely prolonged state of protein sparing and temporary torpor. Proteus anguinus had reduced metabolic and activity rates (considerably lower than those of most surface-dwelling amphibians). These results are interpreted as adaptations to a subterranean existence in which poor and discontinuous food supplies and/or intermittent hypoxia may occur for long periods. Therefore, P. anguinus appears to be a good example of a low-energy-system vertebrate.

Adaptation, Physiological↗

The effect of stimulus features on the visual orienting behaviour of the salamander Plethodon jordani.

The effects of the visual features of prey-like objects on the orienting behaviour of the salamander Plethodon jordani were studied. Two stimuli (cricket dummies, rectangles), moving in opposite directions, were presented simultaneously on a computer screen. They differed in size, contrast, velocity and movement pattern of the entire body or the body appendages. Size and velocity appeared to be the dominant features; shape was of lesser importance. Contrast and movement pattern were of intermediate importance and local motion of little importance. This rank order was the same when the probability of a response to the different stimuli was estimated by means of the maximum-likelihood method. Cluster analysis revealed that in all animals stimuli could be grouped into five clusters. Among individuals, the rank order of stimuli was similar for high- and low-ranking stimuli and varied for those of intermediate rank; stimuli could be grouped into 3-5 clusters. Our findings favour the view that, in amphibians, prey recognition is guided by a number of visual features acting either alone or in combination and depending on internal motivational or attentional states and individual experience.

Animals↗

P450 aromatase expression in the temperature-sensitive sexual differentiation of salamander (Hynobius retardatus) gonads.

Sex differentiation of gonads in amphibians is believed to be controlled genetically, but altered epigenetically or environmentally. When larvae of the salamander Hynobius retardatus were reared at defined temperatures from hatching to metamorphic stages, a high temperature (28 degrees C) induced exclusively female gonads (ovaries), whereas intermediate (20 and 23 degrees C) or lower (16 degrees C) temperatures produced a 1:1 sex ratio of the morphological gonads. The thermosensitive period was determined to be restricted from 15 to 30 days after hatching, just before or when sexual differentiation occurred. Hynobius P450 aromatase (P450arom) cDNA was isolated from adult gonads and the partial nucleotide or deduced amino acid sequences were determined, showing a high level of identity with various vertebrate species. The P450arom gene was expressed predominantly in the adult ovary and brain, weakly in testis, but not in other somatic organs. A typical sexual dimorphism in P450arom expression was detected in normally developing larvae by a quantitative competitive RT-PCR; strong expression in the female gonads but very weak in male gonads. The dimorphism was detected much earlier than the morphological sexual differentiation of the gonads. When larvae were reared at the female-producing temperature (28 degrees C), strong expression was detected in all the temperature-treated larvae, suggesting that P450arom was up-regulated, even in genetic males. Our results confirm the importance of the P450arom regulation in the sexual differentiation of gonads and demonstrate that an up-regulation of P450arom is involved in the process of temperature-sensitive sex reversal in this species.

Amino Acid Sequence↗

Color vision in salamander larvae.

In behavioral experiments using monochromatic prey-patterns, larval salamanders (Salamandra salamandra) are able to discriminate colored from white light. In the retina only blue-yellow opponent-color ganglion cells were recorded. Thus the color vision in these animals is dichromatic.

Animals↗

Large-conductance calcium-activated potassium channels facilitate transmitter release in salamander rod synapse.

Large-conductance calcium-activated potassium (BK) channels are colocalized with calcium channels at sites of exocytosis at the presynaptic terminals throughout the nervous system. It is expected that their activation would provide negative feedback to transmitter release, but the opposite is sometimes observed. Attempts to resolve this apparent paradox based on alterations in action potential waveform have been ambiguous. In an alternative approach, we investigated the influence of this channel on neurotransmitter release in a nonspiking neuron, the salamander rod photoreceptors. Surprisingly, the BK channel facilitates calcium-mediated transmitter release from rods. The two presynaptic channels form a positive coupled loop. Calcium influx activates the BK channel current, leading to potassium efflux that increases the calcium current. The normal physiological voltage range of the rod is well matched to the dynamics of this positive loop. When the rod is further depolarized, then the hyperpolarizing BK channel current exceeds its facilitatory effect, causing truncation of transmitter release. Thus, the calcium channel-BK channel linkage performs two functions at the synapse: nonlinear potentiator and safety brake.

Action Potentials↗

Detection sensitivity and temporal resolution of visual signals near absolute threshold in the salamander retina.

Several studies have suggested that the visual system can detect dim lights with a fidelity limited only by Poisson fluctuations in photon absorption and spontaneous activation of rhodopsin. If correct, this implies that neural processing of responses produced by rod photoreceptors is efficient and effectively noiseless. However, experimental uncertainty makes this conclusion tenuous. Furthermore, previous work provided no information about how accurately stimulus timing is represented. Here, the detection sensitivity and temporal resolution of salamander rods and retinal ganglion cells (RGCs) are compared in nearly matched experimental conditions by using recorded responses to identify the time of a flash. At detection threshold, RGCs could reliably signal the absorption of 20-50 photons, but the rods within the RGC receptive field could signal stimuli 3-10 times weaker. For flash strengths 10 times higher than detection threshold, some RGCs could distinguish stimulus timing with a resolution finer than 100 msec, within a factor of 2 of the rod limit. The relationship between RGC and rod sensitivity could not be explained by added noise in the retinal circuitry but could be explained by a threshold acting after pooling of rod signals. Simulations of rod signals indicated that continuous noise, rather than spontaneous activation of rhodopsin or fluctuations in the single-photon response, limited temporal resolution. Thus, detection of dim lights was limited by retinal processing, but, at higher light levels, synaptic transmission, cellular integration of synaptic inputs, and spike generation in RGCs faithfully conveyed information about the time of photon absorption.

Ambystoma↗

Evolution of salamander life cycles: a major-effect quantitative trait locus contributes to discrete and continuous variation for metamorphic timing.

The evolution of alternate modes of development may occur through genetic changes in metamorphic timing. This hypothesis was examined by crossing salamanders that express alternate developmental modes: metamorphosis vs. paedomorphosis. Three strains were used in the crossing design: Ambystoma tigrinum tigrinum (Att; metamorph), wild-caught A. mexicanum (Am; paedomorph), and laboratory Am (paedomorph). Att/Am hybrids were created for each Am strain and then backcrossed to their respective Am line. Previous studies have shown that a dominant allele from Att (met(Att)) and a recessive allele from lab Am (met(lab)) results in metamorphosis in Att/Am hybrids, and met(Att)/met(lab) and met(lab)/met(lab) backcross genotypes are strongly associated with metamorphosis and paedomorphosis, respectively. We typed a molecular marker (contig325) linked to met and found that met(Att)/met(lab) and met(Att)/met(wild) were associated with metamorphosis in 99% of the cases examined. However, the frequency of paedomorphosis was 4.5 times higher for met(lab)/met(lab) than for met(wild)/met(wild). We also found that met(Att)/met(wild) and met(wild)/met(wild) genotypes discriminated distributions of early and late metamorphosing individuals. Two forms of phenotypic variation are contributed by met: continuous variation of metamorphic age and expression of discrete, alternate morphs. We suggest that the evolution of paedomorphosis is associated with genetic changes that delay metamorphic timing in biphasic life cycles.

Age Factors↗

A comprehensive expressed sequence tag linkage map for tiger salamander and Mexican axolotl: enabling gene mapping and comparative genomics in Ambystoma.

Expressed sequence tag (EST) markers were developed for Ambystoma tigrinum tigrinum (Eastern tiger salamander) and for A. mexicanum (Mexican axolotl) to generate the first comprehensive linkage map for these model amphibians. We identified 14 large linkage groups (125.5-836.7 cM) that presumably correspond to the 14 haploid chromosomes in the Ambystoma genome. The extent of genome coverage for these linkage groups is apparently high because the total map size (5251 cM) falls within the range of theoretical estimates and is consistent with independent empirical estimates. Unlike most vertebrate species, linkage map size in Ambystoma is not strongly correlated with chromosome arm number. Presumably, the large physical genome size ( approximately 30 Gbp) is a major determinant of map size in Ambystoma. To demonstrate the utility of this resource, we mapped the position of two historically significant A. mexicanum mutants, white and melanoid, and also met, a quantitative trait locus (QTL) that contributes to variation in metamorphic timing. This new collection of EST-based PCR markers will better enable the Ambystoma system by facilitating development of new molecular probes, and the linkage map will allow comparative studies of this important vertebrate group.

Ambystoma mexicanum↗

Biogeography of the southeastern United States: a comparison of salamander phylogeographic studies.

Most phylogeographic studies of species from the southeastern United States have shown a simple east-west division of mtDNA variation. However, a study of the salamander Ambystoma maculatum resulted in a more complex pattern that includes a close affinity between populations from the Central Highlands of Missouri and Arkansas and the Coastal Plain separated by a genetically distinct central group of populations. We test the generality of this observation by surveying mitochondrial DNA (mtDNA) variation in the closely related species A. talpoideum. An Ambystoma-specific intergenic spacer was amplified and sequenced. The 26 resulting haplotypes varied from 380 to 800 base pairs, and alignments, including the outgroup, required 101 insertions/deletions. Sequence divergence among haplotypes ranged from 0.001 to 0.758. Population subdivision was extensive (theta = 0.64). Phylogenetic analysis of A. talpoideum mtDNA sequence reveals a close relationship between the populations from the Central Highlands and the Coastal Plain. This result is similar to that obtained for A. maculatum, although the A. talpoideum clade is not as well differentiated from its sister clades. We discuss the differences and similarities between the two Ambystoma species and previous studies and call for increased focus on multiple species with similar ecologies as a way to detect subtle biogeographic events.

Ambystoma↗

Observations on the migration and differentiation of neural crest cells in somite extirpated salamander larvae.

In order to research the influences of the somites or their derivatives on the migration and differentiation of neural crest cells, unilateral extirpation of somites was performed in the larvae of the salamander (Hynobius lichenatus Boulenger). Two types of neural crest cells appeared in the somite-free environment. They could already be distinguished from each other shortly after beginning of migration. One type of the cells was distributed in the expanded space between the epidermis and the neural tube, with melanocytes differentiated from them. The other type of the cells migrated closely to the lateral wall of the neural tube. They aggregated as a cord on the ventrolateral side of the neural tube and gave rise either to neurons or to satellite cells of spinal ganglia. The sequence of cytological events along the development of the cells and the time required for each event were essentially identical with those of the melanocytes and spinal ganglion cells developing on the control side. The present results suggest not only that migration and differentiation of neural crest cells are independent of the possible regulation of the somites or their derivatives but also that the fate of neural crest cells is determined before the onset of migration.

Animals↗

Correlated histochemical and electron microscopic studies of the esophageal epithelium in the salamander, Hynobius nebulosus.

The esophageal epithelium of the adult salamander, Hynobius nebulosus, was studied by histochemistry, including periodic acid-Shiff (PAS), alcian blue both at pH 2.5 and pH 1.0, and alcian blue (pH 2.5)-PAS with or without neuraminidase-digestion, and scanning and transmission electron microscopy. The epithelium was columnar in type, comprising partly pseudostratified and partly two-layered architecture; it consisted mainly of ciliated cells, goblet cells, and basal cells. The ciliated cells consisted of two types, light and dark: both types, especially the latter, frequently contained numerous small mucous granules in their apical portion. Ciliated-mucous cells were also occasionally present. Immature ciliated cells were frequently found. Goblet cells were divided into two types: Type I cells possessed electron lucent mucous granules, which frequently contained dense specific inclusions, and frequently bulged into the lumen; Type II cells had moderately electron-dense mucous granules with no inclusions and a conical apex which did not exceed the level of the lumen. The Type I cells were closely distributed throughout the esophagus, while the Type II were mainly dispersed in the cranial portion, remarkably decreasing in the caudal portion. Correlated histochemical and electron microscopic observations suggested that, in the Type I cell, mucous granules contain acid mucosubstances, while in the Type II, they possess neutral mucosubstances, and that in the Type I cell mucous granules consist of sialic acid-containing glycoproteins and their swollen portions are more highly sulfated than the non-swollen ones.

Animals↗

Linking direct and indirect data on dispersal: isolation by slope in a headwater stream salamander.

There is growing recognition of the need to incorporate information on movement behavior in landscape-scale studies of dispersal. One way to do this is by using indirect indices of dispersal (e.g., genetic differentiation) to test predictions derived from direct data on movement behavior. Mark-recapture studies documented upstream-biased movement in the salamander Gyrinophilus porphyriticus (Plethodontidae). Based on this information, we hypothesized that gene flow in G. porphyriticus is affected by the slope of the stream. Specifically, because the energy required for upstream dispersal is positively related to slope, we predicted gene flow to be negatively related to change in elevation between sampling sites. Using amplified DNA fragment length polymorphisms among tissue samples from paired sites in nine streams in the Hubbard Brook Watershed, New Hampshire, USA, we found that genetic distances between downstream and upstream sites were positively related to change in elevation over standardized 1-km distances. This pattern of isolation by slope elucidates controls on population connectivity in stream networks and underscores the potential for specific behaviors to affect the genetic structure of species at the landscape scale. More broadly, our results show the value of combining direct data on movement behavior and indirect indices to assess patterns and consequences of dispersal in spatially complex ecosystems.

Animals↗

Lethal and sublethal effects of atrazine, carbaryl, endosulfan, and octylphenol on the streamside salamander (Ambystoma barbouri).

Agricultural contaminants may be contributing to worldwide amphibian declines, but little is known about which agrichemicals pose the greatest threat to particular species. One reason for this is that tests of multiple contaminants under ecologically relevant conditions are rarely conducted concurrently. In this study, we examined the effects of 37-d exposure to the agrichemicals atrazine (4, 40, and 400 micrograms/L), carbaryl (0.5, 5, and 50 micrograms/L), endosulfan (0.1, 1, and 10 micrograms/L for 31 d and 0.1, 10, and 100 micrograms/L for the last 6 d), and octylphenol (5, 50, and 500 micrograms/L) and to a solvent control on streamside salamanders (Ambystoma barbouri) in the presence and absence of food. We found that none of the agrichemicals significantly affected embryo survival, but that hatching was delayed by the highest concentration of octylphenol. In contrast to embryos, larval survival was reduced by the highest concentrations of carbaryl, endosulfan, and octylphenol. Growth rates were lower in the highest concentrations of endosulfan and octylphenol than in all other treatments, and the highest concentration of endosulfan caused respiratory distress. Significantly more carbaryl, endosulfan, and octylphenol tanks had larvae with limb deformities than did control tanks. Refuge use was independent of chemical exposure, but 10 micrograms/L of endosulfan and 500 micrograms/L of octylphenol decreased larval activity. Systematically tapping tanks caused a greater activity increase in larvae exposed to 400 micrograms/L of atrazine and 10 micrograms/L of endosulfan relative to solvent controls, suggesting underlying nervous system malfunction. Hunger stimulated a decrease in refuge use and an increase in activity, but this response was least pronounced in larvae exposed to the highest concentration of any of the four agrichemicals, possibly because these larvae were the most lethargic. More studies are needed that concurrently examine the effect of multiple contaminants on amphibians so we can better identify effective mitigating measures.

Ambystoma↗

Variation in cannibalistic polyphenism between populations in the salamander Hynobius retardatus.

Organisms sometimes change their phenotype to maximize fitness according to local environments. If the frequency of the broad-headed "cannibal" morph in the larvae of the salamander Hynobius retardatus has been evolutionarily maintained at a certain level within a population as a result of local adaptation, variations in its frequency should be found among different populations with environmental variation. We investigated whether variations in the frequency of the broad-headed morph were present in 2 different populations, Nopporo (a low-density population) and Erimo (a high-density population), by raising larvae from the respective populations under the same experimental conditions. The occurrence rate of the broad-headed "cannibal" morph was significantly different between the 2 populations when examined with different experimental larval densities. These results suggest that the reaction norm with respect to the frequency of the broad-headed morph is different between the Nopporo and Erimo populations. Because the local populations are assumed to be selected for under different environments, the different reaction norm might have evolved in response to different selection pressures.

Animals↗

A new species of lentic breeding Korean salamander of the genus Hynobius (Amphibia, Urodela).

A new species of salamander, Hynobius yangi, is described from Kori (=Hyoam-ri), Busan-shi, southeastern Korea. It is a lentic breeder belonging to the H. nebulosus species group, and has long been confused with another Korean species H. leechii. The new species, however, is genetically substantially differentiated from Korean and Japanese relatives. Hynobius yangi is morphologically very similar to H. leechii, but could be differentiated from it by the tail shape, degree of limb separation, shape of vomerine teeth series, shape of egg sac, and dorsal coloration.

Animals↗