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PubMed1993

Synaptic organization of dopaminergic amacrine cells in the larval tiger salamander retina.

The ultrastructural features and synaptic interactions of tyrosine hydroxylase-like-immuno-reactive amacrine cells in the larval tiger salamander retina were examined using routine immunoelectron microscopy. The somas of tyrosine hydroxylase-like-immunoreactive amacrine cells were immunostained evenly throughout their cytoplasm. Their nuclei were generally unstained and possessed indented nuclear membranes. The processes of tyrosine hydroxylase-like-immunoreactive amacrine cells were homogeneously stained with the exception of their mitochondria, whose morphology was often disrupted by the staining procedure. Tyrosine hydroxylase-like-immunoreactive amacrine cell processes were characterized by an occasional dense-cored vesicle(s), in addition to a generally homogeneous population of small, round, agranular synaptic vesicles. They formed conventional synaptic junctions that were characterized by symmetrical synaptic membrane densities. A total of 168 synapses were observed that involved tyrosine hydroxylase-like-immunoreactive amacrine cell processes. A large percentage (79.8%) of these synaptic arrangements were found in sublayer 1 of the inner plexiform layer, while substantially lower percentages were observed in sublayers 3 (9.5%) and 5 (10.7%). They served as pre- and postsynaptic elements 63.1 and 36.9% of the time, respectively. Tyrosine hydroxylase-like-immunoreactive amacrine cell processes were presynaptic to amacrine cell processes (36.9% of total synaptic involvement) and processes that lack synaptic vesicles and whose origin remains uncertain (26.2%). They received synaptic input primarily from amacrine cell processes (31.0%). Tyrosine hydroxylase-like-immunoreactive amacrine cell processes also received a few ribbon synapses from bipolar cells (5.9%). Each of these synaptic relationships were observed in each of sublayers 1, 3 and 5 of the inner plexiform layer, with the majority of each arrangement being found in sublayer 1.

Ambystoma↗
PubMed1995

Comparison of the actions of glycine and related amino acids on isolated third order neurons from the tiger salamander retina.

Whole cell voltage and current clamp recordings were obtained from third order neurons isolated from the salamander retina. Using cross desensitization, the structure-function relationship of short chain amino acids on the glycine receptor were examined. L-Serine, L-alanine, beta-alanine and taurine all cross desensitized with glycine, but did not show significant cross desensitization with GABA. This indicates that these amino acids act at the glycine receptor. The order of potency was glycine >> beta-alanine > taurine >> L-alanine > L-serine. TAG, a reputed selective taurine antagonist, was equally effective in blocking taurine and glycine currents. There is no evidence for distinct receptors for taurine. Amino acids with larger moieties at the alpha carbon, such as threonine and valine, produced inactive ligands. Placing a methyl group on the amine of glycine or esterification of the carboxyl group also greatly reduced activity. Based on these modifications of the glycine molecule, it appears that selectivity at the glycine receptor results in part from steric restrictions at all three sites in the glycine chain. The steric interference is most critical at the carboxyl and amino ends, and less limiting at the alpha carbon. Doses of L-serine that had only slight effects in voltage clamp experiments, nevertheless produced large effects in current clamp experiments. This indicates that several endogenous amino acids can have significant effects on membrane voltage, even when their shunting activity may be small. High concentrations of agonists produced desensitization in the voltage clamp records, but there was little evidence of desensitization in the current clamp experiments. These results indicate that several endogenous amino acids can activate the glycine receptor, but there is no evidence for a discrete receptor for taurine, beta-alanine, L-alanine or L-serine. Since all these endogenous amino acids have similar amino and acid terminals, reduction in potency results from steric interference around the alpha carbon. This graded potency may have functional significance in mediating inhibition.

Amino Acids↗
PubMed1990

Anterior decerebration blocks visual habituation in the larval salamander (Ambystoma punctatum).

Amputation of the rostral half of the cerebrum induces a compulsion-like reaction in larval Ambystoma punctatum towards Enchytraeus protected within glass vials. Normal and craniotomized larvae are visually attracted to worm-containing vials, as revealed by time-lapse video taping but, after several unsuccessful attempts to get the prey, habituate and depart. The video tapes revealed that anteriorly decerebrated animals spent as much as 100 of 120 minutes at the worm-containing vial, repeatedly but futilely attacking the glass. The data indicate that the telencephalon plays an active negative role in the salamander larva's visually guided behavior.

Ambystoma↗
PubMed1986

Cloning and sequencing of mRNAs coding for two adult alpha globin chains of the salamander Pleurodeles waltlii.

A cDNA library of erythrocyte mRNAs was established from immature red blood cells of the adult amphibian, Pleurodeles waltlii (urodele; salamander). The cDNA clones corresponding to the four adult globin chains were first identified and characterized by positive selection and the cDNAs derived from the two (major and minor) alpha-globin chains sequenced. The sequences presented contain both the complete 3'-noncoding region and the coding region of both chains, with the exception of the first nine codons of the minor alpha-chain, and a portion of the 5'-noncoding region of the major chain. The amino acid sequences of the encoded alpha-globin polypeptides have been deduced and compared with those of Xenopus laevis and of man. These comparative studies suggest that the alpha-globins of Pleurodeles waltlii and Xenopus laevis may have diverged from a common ancestral gene at the time when mammalian and amphibian lines diverged, and that they then evolved separately. Duplication of the alpha-gene, which is responsible for the polypeptide heterogeneity, appears to have occurred earlier in Pleurodeles waltlii than in Xenopus laevis.

Amino Acid Sequence↗
PubMed1988

Growth and synapse formation among major classes of adult salamander retinal neurons in vitro.

Adult neurons, isolated from the salamander retina, were maintained in low-density cell culture and examined for synapse formation by electrophysiological and electron microscopic techniques. Morphologically identifiable rod, cone, horizontal, bipolar, and amacrine/ganglion cells survived for many months, grew processes, and formed numerous cell contacts. Intracellular recordings showed the presence of a variety of voltage- and time-dependent conductances and both electrical and chemical transmission among these cells. At the ultrastructural level, gap junctions, monad ribbon synapses, and conventional synapses, like those present in the intact retina, were observed in sibling cultures. Thus, all major classes of adult retinal neurons, in addition to ganglion cells, are able to regenerate processes and reform synapses. The regenerated synaptic contacts are functional and structurally diverse.

Animals↗
PubMed2005

Notes on cranial ontogeny and delayed metamorphosis in the hynobiid salamander Ranodon sibiricus Kessler, 1866 (Urodela).

The skull of larvae, juveniles and adults of the rare and primitive hynobiid salamander Ranodon sibiricus was re-examined using transparencies and illustrated by new graphics. The earliest larva available for investigations already had the dominant bones. The maxillary, however, was still lacking. Previous descriptions regarding the appearance and growth of bones could be largely confirmed. The vomer, first seen as a relatively small obliquely arranged dentate bar in the 3.8 cm long larva, became larger during ontogeny, but did not change its position remarkably. The vomerine pars dentalis with only a single tooth line was straight in larvae and juveniles, but was slightly curved in adults allowing for distinction of an outer and inner portion. This feature is typical and more pronounced in most other hynobiids. The significance of the vomer and vomerine dentition for systematic and phylogenetic purposes and its changes during metamorphosis are briefly discussed. Two of the specimens examined showed delayed metamorphosis very likely caused by low temperatures. Here the temporal course of transformation was "stretched" and therefore some alterations, e.g. regression of the palatinal portion of the palatopterygoid, were shown more clearly. Continuous growth of some skull elements in these individuals suggested a relative independence from metamorphosis perhaps due to variable thyroid activity and/or independent changes in individual tissue sensitivities. It is suggested that remodelling of the mouth roof could be used for staging urodele ontogeny.

Aging↗
PubMed2004

Signal transmission from cones to amacrine cells in dark- and light-adapted tiger salamander retina.

Amacrine cells (ACs) are third-order interneurons in the retina that mediate antagonistic surround inputs to retinal ganglion cells and motion-related signals in the inner retina. Previous studies have revealed that rod-to-AC signals in dark-adapted retina are mediated by a nonlinear high-gain synaptic pathway. In this study, we investigated how cone signals are transmitted to ACs under dark- and light-adapted conditions. By using the spectral subtraction method, we found that the voltage gain of the cone-AC synaptic pathway in dark-adapted salamander retina (GD) is between 28 and 72, which is about one order of magnitude lower than the voltage gain of the rod-AC pathway. This suggests that, in darkness, rod signals are more efficiently transmitted to the ACs than cone signals. The voltage gain of the cone-AC synaptic pathway in the presence of 500 nm/-2.4 background light, GL, ranges between 28 and 56. Linear regression analysis indicates that GD and GL are strongly, positively, and linearly correlated. The average GL/GD ratio is 0.73, suggesting that, on average, GL in any given AC is about 73% of GD. This adaptation-induced change in cone-AC voltage gain exemplifies use-dependent modulations of synaptic transmission in the retina, and possible mechanisms underlying light-mediated alterations of retinal synaptic function are discussed.

Adaptation, Ocular↗
PubMed2006

Structure and evolution of the mitochondrial control region and flanking sequences in the European cave salamander Proteus anguinus.

The European cave salamander Proteus anguinus Laurenti 1768 is one of the best-known subterranean animals, yet its evolutionary history and systematic relationships remain enigmatic. This is the first comprehensive study on molecular evolution within the taxon, using an mtDNA segment containing the control region (CR) and adjacent sequences. Two to seven tandem repeats of 24-32 bp were found in the intergenic spacer region (VNTR1), and three, four or six repeats, 59-77 bp each, in the 3' end of the CR (VNTR2). Different molecular mechanisms account for VNTR2 formation in different lineages of Proteus. The overall CR variation was lower than that of the spacer region, the 3' end of the cytb gene, or the tRNA genes. Individual genes and the concatenated non-repetitive sequences produced similar, well resolved maximum likelihood, Bayesian inference and parsimony trees. The numbers of repeat elements as well as the genealogy of the VNTR2 repeat units were mostly inconsistent with the groupings of the non-repetitive sequences. Different degrees of repeat array homogenization were detected in all major groups. Orthology was established for the first and the second VNTR2 elements of some populations. These two copies may therefore be used for analyses at the population level. The pattern of CR sequence variation points to strong genetic isolation of hydrographically separated populations. Genetic separation of the major groups of populations is incongruent with the current division into subspecies.

Animals↗
PubMed2003

Kinetics of synaptic transfer from rods and cones to horizontal cells in the salamander retina.

We examined synaptic transmission between rods or cones and horizontal cells, using perforated patch recording techniques in salamander retinal slices. Experimental conditions were established under which horizontal cells received nearly pure rod or pure cone input. The response-intensity relation for both photoreceptors and horizontal cells was described by a Michaelis-Menten function with an exponent close to 1. A dynamic model was developed for the transduction from photoreceptor voltage to postsynaptic current. The basic model assumes that: (i) photoreceptor light-evoked voltage controls Ca2+ entry according to a Boltzmann relation; (ii) the rate of glutamate release depends linearly on the voltage-gated Ca2+ current (ICa) in the synaptic terminal; (iii) glutamate concentration in the synaptic cleft reflects the balance of release and reuptake in which reuptake obeys first order kinetics; (iv) the binding of glutamate to its receptor and channel gating are fast compared with glutamate kinetics in the synaptic cleft. The good fit to the model confirms that these are the key features of synaptic transmission from rods and cones. The model accommodated changes in kinetics induced by the glutamate uptake blocker, dihydrokainate. The match between model and response was not improved by including an estimate of alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) receptor desensitization or by making glutamate uptake voltage dependent.

Animals↗
PubMed2004

Actin filaments regulate voltage-gated ion channels in salamander retinal ganglion cells.

The regulation of voltage-activated K(+), and Ca(2+) currents by actin filaments was studied in salamander retinal ganglion cells, using the whole-cell patch clamp technique and Ca(2+) imaging. Disruption of F-actin by cytochalasin B or latrunculin B resulted in a reduction of L-type Ca(2+) current by 55+/-4%, and a sustained outward K(+) current (I(k)) by 41+/-3%. The effect was diminished when the F-actin stabilizing agent phalloidin was present in the patch pipette. In a group of cells where I(K) exhibited a small degree of inactivation, the effect of F-actin disruption on current was dual; it increased it by 89+/-16%, at -10 mV, and reduced it by 37+/-5% at +50 mV voltage step from the same holding potential of -70 mV. This was accompanied by a shift in a voltage of half-maximal activation toward negative potentials by approximately 20 mV. In Ca(2+) imaging experiments, 30 min incubation of isolated neurons with latrunculin A reduced a depolarization-induced Ca(2+) accumulation by 45+/-5%. These results suggest a role for the actin cytoskeleton in regulating voltage-gated ion channels in retinal ganglion cells.

Actins↗
PubMed2005

Dynamic response properties of visual neurons and context-dependent surround effects on receptive fields in the tectum of the salamander Plethodon shermani.

Neuronal responses to complex prey-like stimuli and rectangles were investigated in the tectum of the salamander Plethodon shermani using extracellular single-cell recording. Cricket dummies differing in size, contrast or movement pattern or a rectangle were moved singly through the excitatory receptive field of a neuron. Paired presentations were performed, in which a reference stimulus was moved inside and the different cricket dummies or the rectangle outside the excitatory receptive field. Visual object recognition involves much more complex spatial and temporal processing than previously assumed in amphibians. This concerns significant changes in absolute number of spikes, temporal discharge pattern, and receptive field size. At single presentation of stimuli, the number of discharges was significantly changed compared with the reference stimulus, and in the majority of neurons the temporal pattern of discharges was changed in addition. At paired presentation of stimuli, neurons mainly revealed a significant decrease in average spike number and a reduction of excitatory receptive field size to presentation of the reference stimulus inside the excitatory receptive field, when a large-sized cricket stimulus or the rectangle was located outside the excitatory receptive field. This inhibition was significantly greater for the large-sized cricket stimulus than for the rectangle, and indicates the biological relevance of the prey-like stimulus in object selection. The response properties of tectal neurons at single or paired presentation of stimuli indicate that tectal neurons integrate information across a much larger part of visual space than covered by the excitatory receptive field. The spike number of a tectal neuron and the spatio-temporal extent of its excitatory receptive field are not fixed but depend on the context, i.e. the stimulus type and combination. This dynamic processing corresponds with the selection of the stimuli in the visual orienting behavior of Plethodon investigated in a previous study, and we assume that tectal processing is modulated by top down processes as well as feedback circuitries.

Animals↗
PubMed2005

Complex secretory products in the oviduct of the plethodontid salamander Bolitoglossa dofleini (Amphibia, Urodela).

The ultrastructure of the secretory granules in the cells of the subdivisions of the oviduct in the neotropical plethodontid salamander Bolitoglossa dofleini was studied by transmission electron microscopy. In addition, we applied the cationic dye Cuprolinic Blue (CB) at different electrolyte concentrations to demonstrate proteoglycans, and the pyrogallol red-copper (PR-C) method to stain proteins at the ultrastructural level. The entire oviduct is lined by a simple epithelium that contains ciliated and microvillous cells in the first subdivision, the aglandular pars recta; microvillous cells show a moderate secretory activity. The following pars convoluta is differentiated into five glandular subdivisions and the aglandular "uterine portion". Especially in the glandular parts, the epithelium is arranged in longitudinal folds. At their crests ciliated and microvillous cells similar to those in the pars recta occur. Gland cells are crowded with secretory granules that differ in their structural complexity (with and without electron-dense spheres or masses; elaborated, homogeneous or granular matrix; spherical; distorted) along the various subdivisions. Further, as suggested by the CB-technique, the cranial subdivisions contain large amounts of sulphated proteoglycans that decrease in the caudal direction. Carboxylated proteglycans appear to be present in all subdivisions examined. Electron-dense spheres of secretory granules are largely free of CB-precipitates, but stain more or less intensely with PR-C. The ultrastructure of the pars recta, and especially the "uterine portion" indicates transporting capability. The epithelial cells of the "uterus" have coated pits and a considerable amount of lysosome-like bodies.

Animals↗
PubMed2003

Genomic sequence of a ranavirus (family Iridoviridae) associated with salamander mortalities in North America.

Disease is among the suspected causes of amphibian population declines, and an iridovirus and a chytrid fungus are the primary pathogens associated with amphibian mortalities. Ambystoma tigrinum virus (ATV) and a closely related strain, Regina ranavirus (RRV), are implicated in salamander die-offs in Arizona and Canada, respectively. We report the complete sequence of the ATV genome and partial sequence of the RRV genome. Sequence analysis of the ATV/RRV genomes showed marked similarity to other ranaviruses, including tiger frog virus (TFV) and frog virus 3 (FV3), the type virus of the genus Ranavirus (family Iridoviridae), as well as more distant relationships to lymphocystis disease virus, Chilo iridescent virus, and infectious spleen and kidney necrosis virus. Putative open reading frames (ORFs) in the ATV sequence identified 24 genes that appear to control virus replication and block antiviral responses. In addition, >50 other putative genes, homologous to ORFs in other iridoviral genomes but of unknown function, were also identified. Sequence comparison performed by dot plot analysis between ATV and itself revealed a conserved 14-bp palindromic repeat within most intragenic regions. Dot plot analysis of ATV vs RRV sequences identified several polymorphisms between the two isolates. Finally, a comparison of ATV and TFV genomic sequences identified genomic rearrangements consistent with the high recombination frequency of iridoviruses. Given the adverse effects that ranavirus infections have on amphibian and fish populations, ATV/RRV sequence information will allow the design of better diagnostic probes for identifying ranavirus infections and extend our understanding of molecular events in ranavirus-infected cells.

Animals↗
PubMed2006

Cross-talk between ON and OFF channels in the salamander retina: indirect bipolar cell inputs to ON-OFF ganglion cells.

It has been widely accepted that ON and OFF channels in the visual system are segregated with little cross-communication, except for the mammalian rod bipolar cell-AII amacrine cell-ganglion cell pathway. Here, we show that in the tiger salamander retina the light responses of a subpopulation of ON-OFF ganglion cells are mediated by crossing the ON and OFF bipolar cell pathways. Although the majority of ON-OFF ganglion cells (type I cells) receive direct excitatory inputs from depolarizing and hyperpolarizing bipolar cells (DBCs and HBCs), about 5% (type II cells) receive indirect excitatory inputs from DBCs and 20% (type III cells) receive indirect excitatory inputs from HBCs. These indirect bipolar cell inputs are likely to be mediated by a subpopulation of amacrine cells that exhibit transient hyperpolarizing light responses (AC(H)s) and make GABAergic/glycinergic synapses on DBC or HBC axon terminals. GABA and glycine receptor antagonists enhanced the ON and OFF excitatory cation current (DeltaI(C)) in type I ganglion cells, but completely suppressed the ON DeltaI(C) mediated by DBCs in type II cells and the OFF DeltaI(C) mediated by HBCs in types III cells. Dendrites of type I cells ramify in both sublamina A and B, type II cells exclusively in sublamina A, and type III cells exclusively in sublamina B of the inner plexiform layer. These results demonstrate that indirect, amacrine cell-mediated bipolar cell-ganglion cell synaptic pathways exist in a non-mammalian retina, and that bidirectional cross-talk between ON and OFF channels is present in the vertebrate retina.

Amacrine Cells↗
PubMed2006

Dopaminergic modulation and rod contribution in the generation of oscillatory potentials in the tiger salamander retina.

The roles of rod and cone input and of dopamine in the generation of oscillatory potentials were studied in tiger salamander retina. Under scotopic conditions, oscillations were elicited with a green, but not a red stimulus. With mesopic background illumination, both stimuli caused oscillations. Addition of quinpirole to a mesopic retina eliminated oscillations while SKF-38393 had no effect. Similarly, addition of sulpiride to a light-adapted retina elicited oscillatory activity, but SCH 22390 had no effect. These results suggest that oscillatory potentials are elicited through activation of the rod pathway and are modulated by dopamine through D2-receptors.

Adaptation, Physiological↗
PubMed2005

Effects of steroids and dioxin (2,3,7,8-TCDD) on the developing wolffian ducts of the tiger salamander (Ambystoma tigrinum).

This study was undertaken to investigate effects of the prototypical dioxin, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) on steroid-dependent development of the wolffian ducts of an amphibian, the tiger salamander (Ambystoma tigrinum). Larvae with immature gonads and undeveloped mullerian ducts were injected with the steroid hormones estradiol (E2), dihydrotestosterone (DHT), or vehicle alone. Additionally, steroid-treated and vehicle-control larvae were immersed in sub-lethal solutions of technical grade TCDD (0, 0.0003, 0.003, 0.03, 0.3, and 3.0 microg TCDD/L). Both steroid treatments stimulated hypertrophy of the wolffian duct epithelium and an increase in mean epithelial cell size. Only DHT treatment stimulated epithelial cell proliferation. TCDD stimulated wolffian duct hypertrophy through an increase in mean epithelial cell size. TCDD acted as an androgen agonist on wolffian duct epithelial area and epithelial cell size. TCDD had no effect on wolffian duct epithelium among E2-injected animals. Stimulatory effects on cell size were observed at 0.0003 microg/L TCDD in saline-injected animals and at 0.003 microg/L TCDD in DHT-injected animals. Both E2 and DHT stimulated growth of the wolffian ducts early in development. Technical grade TCDD alone mimics E2 and DHT action but exhibits an androgen-agonistic action in the presence of exogenously administered DHT. Implications of possible interactions between TCDD and xenosteroids are discussed.

Ambystoma↗
PubMed2006

Effects of androgens on behavioral and vomeronasal responses to chemosensory cues in male terrestrial salamanders (Plethodon shermani).

Chemosensory stimuli and sex steroid hormones are both required for the full expression of social behaviors in many species. The terrestrial salamander, Plethodon shermani, is an emerging nonmammalian system for investigating the nature and evolution of pheromonal communication, yet little is known regarding the role of sex steroid hormones. We hypothesized that increased circulating androgen levels in male P. shermani enhance chemoreception through morphological, behavioral, and physiological mechanisms. Experimental elevation of plasma androgens increased development of cirri, morphological structures thought to enhance the transfer of chemosensory cues from the substrate to the vomeronasal organ (VNO). Elevated plasma androgens also increased expression of a chemo-investigatory behavior (nose tapping) and increased preference for some female-derived chemosensory cues. Male-produced courtship pheromones activated a large number of cells in the VNO as measured by the method of agmatine uptake. However, androgen levels did not affect the total number of vomeronasal cells activated by male-produced courtship pheromones. Future studies will determine whether androgens potentially modulate responsiveness of the VNO to female-derived (as opposed to male-derived) chemosensory cues.

Analysis of Variance↗