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PubMed1993

Glutamate uptake from the synaptic cleft does not shape the decay of the non-NMDA component of the synaptic current.

To study the role of glutamate uptake at central glutamatergic synapses, we used the uptake blocker L-transpyrrolidine-2,4-dicarboxylate (PDC). The effects of PDC on the glutamate uptake current in salamander retinal glia indicated that PDC competes with glutamate for transport on the uptake carrier and that 300 microM PDC should significantly reduce the uptake of glutamate during the synaptic current. In isolated rat hippocampal neurons, 300 microM PDC did not affect non-N-methyl-D-aspartate (NMDA) receptor currents, but reduced NMDA receptor currents by 30%. In hippocampal and cerebellar slices, whereas 300 microM PDC reduced the NMDA component of excitatory synaptic currents by 50%, it reduced the non-NMDA component only slightly with no change in its decay time constant. Thus, the decay rate of the non-NMDA component is not set by the rate of glutamate uptake from the synaptic cleft into the presynaptic terminal.

Animals↗
PubMed1994

A cGMP-gated current can control exocytosis at cone synapses.

The voltage-gated Ca2+ current in cone photoreceptors operates over only a small part of the physiological voltage range produced by light and, consequently, appears insufficient for controlling transmitter release. We have used a whole-cell voltage clamp to measure membrane current and the capacitance change produced by exocytosis in solitary cone and rod photoreceptors isolated from the salamander retina. In both types of photoreceptor, Ca2+ influx through voltage-gated Ca2+ channels initiated exocytosis. In addition, Ca2+ influx through a cGMP-gated channel in the inner segment and synaptic processes of cones also initiated exocytosis. The cGMP-gated current sustained exocytosis over the entire physiological voltage range.

Ambystoma↗
PubMed1994

Modulation of ion channels in rod photoreceptors by nitric oxide.

Subcellular compartments in the outer retina of the larval tiger salamander were identified as likely sites of production of nitric oxide (NO), a recently recognized intercellular messenger. NADPH diaphorase histochemistry and NO synthase immunocytochemistry labeled photoreceptor ellipsoids and the distal regions of bipolar and glial cells apposing photoreceptor inner segments, suggesting a role for NO in visual processing in the outer retina. We investigated the actions of NO on several rod photoreceptor ion channels. Application of the NO-generating compound S-nitrosocysteine increased Ca2+ channel current and a voltage-independent conductance, but had no affect on voltage-gated K+ or nonspecific cation currents. Given the steep relation between voltage-dependent Ca2+ influx and photoreceptor synaptic output, these results indicate that NO could modulate transmission of the photoresponse to second order cells.

Amino Acid Oxidoreductases↗
PubMed1991

Toward a pharmacology of odor receptors and the processing of odor images.

Odor molecules may be considered as molecular ligands which bind to receptors in the olfactory sensory neurons to give rise to the sensory response. Binding studies in whole sensory epithelia suggest that the receptors also bind muscarinic cholinergic antagonists. Preliminary electrophysiological evidence indicates that muscarinic and beta adrenergic antagonists block odor-elicited membrane currents in single isolated salamander sensory neurons. These results support the idea that models developed for analyzing ligand binding by members of the 7 transmembrane domain family of membrane receptors may apply rather closely to olfactory transduction. We suggest that sensory neurons express single receptor types with differing degrees of affinity for different ligands. We further suggest that glomeruli in the olfactory bulb function as labeled lines for particular sets of odor ligand determinants, and that interglomerular circuits bind together similar glomeruli and enhance contrast between dissimilar glomeruli. The odor image laid down in the sensory neuron population is thus subjected to abstracting and enhancement at the glomerular stage, prior to being transmitted for further processing in the deeper layers of the olfactory bulb and in the olfactory cortex.

Animals↗
PubMed2005

Genetic basis of phenotypic plasticity for predator-induced morphological defenses in anuran tadpole, Rana pirica, using cDNA subtraction and microarray analysis.

Anuran tadpoles (Rana pirica) are induced to develop a higher tail and a bulgy body as predator-specific morphological responses when they are exposed to predatory larval salamanders. Subtractive hybridization was performed using induced tadpole body skin and normal tadpoles' body skin. A total of 196 clones showed higher expression, and 104 clones showed lower expression, when they formed bulgy bodies. In the subtraction, carboxypeptidase B, trypsinogen, elastase I, fibrinogen, elastase II, triacyl-glycerol lipase, and alpha1-antitrypsin genes showed lower expression. In contrast, RT-like protein, bullous pemphigoid antigen, phosphoserine aminotransferase, uromodulin, tetranectin, chaperonin-like protein, zinc finger protein, osteonectin, aldehyde dehydrogenase, Sec 23A protein, and ribosomal protein showed higher gene expression. Microarray analysis was also performed using this subtracted cDNA (nine replicates). Results of the microarray data essentially corresponded with those of the subtraction data, and the degree of the suppressed genes was much stronger than that of the expressed genes. Carboxypeptidase B showed the strongest suppression, and its inhibition range was from 1/100 to 3/100 compared with that of control body skin. Strong suppression was also observed with trypsinogen, elastase I, fibrinogen, and elastase II as well. These results can be interpreted as increases of fibrinolysis by strong depression of both carboxypeptidase B and other genes simultaneously, resulting in the retention of blood vessels and facilitating the circulation of blood. Expression was observed in phosphoserine aminotransferase, aldehyde dehydrogenase, RT-related protein, chaperonin-like protein, tetranectin, bullous pemphigoid antigen, uromodulin, and Sec 23A protein. They were significantly (p<0.05) increased and were at least 1.5 times greater compared with the control. From the appearance, it seems that the bulgy shaped body is highly connecting to the bullous pemphigoid (BP) antigen that causes the skin blistering disorder, and tetranectin and uromodulin may be related to the extracell matrix through myogenesis, protein secretion, and ion transport, respectively. Since the RT-related protein gene derived from retrotransposon (L1) is known to disrupt mammalian transcriptomes, retrotransposon may be involved with phenotypic plasticity for morphological defense by Rana prica against predator threat.

Animals↗
PubMed2006

Vasopressin-induced translocation and proteolysis of protein kinase Calpha in an amphibian brain: modulation by corticosterone.

In urodele amphibians, the hypothalamic neuropeptide arginine vasotocin and the adrenal steroid corticosterone interact to regulate reproductive behavior by actions in the brain. The present study investigated signal transduction pathways underlying acute effects of vasotocin and corticosterone, presumably mediated via "non-genomic" steroid action, in an amphibian brain. We used Western blot to examine the effects of corticosterone and the vasotocin receptor agonist arginine vasopressin, alone and in combination, on the subcellular localization and proteolytic processing of protein kinase C-alpha (PKCalpha) in tiger salamander brain tissue. Treatment of whole brain minces with vasopressin or vasotocin led to increases in PKCalpha in membrane fractions and concurrent decreases in PKCalpha in cytosolic fractions. Vasopressin or vasotocin treatment also induced the appearance in membrane and cytosolic fractions of a PKCalpha-immunoreactive band that corresponds to PKMalpha, the proteolytically generated, free catalytic subunit of PKCalpha. Treatment with corticosterone alone had no consistent effect on either PKCalpha or PKMalpha in either fraction. However, pretreatment with corticosterone reliably blocked vasopressin-induced increases in cytosolic PKMalpha. These data provide new information about the cellular mechanisms of action of vasopressin and corticosterone in the vertebrate brain and suggest a cellular mechanism by which the two hormones interact to regulate neuronal physiology and behavior.

Ambystoma↗
PubMed2005

Chloride concentration affects Kv channel voltage-gating kinetics: Importance of experimental anion concentrations.

Chloride concentration has been shown to have a dramatic impact on protein folding and subsequent tertiary conformation [K.D. Collins, Ions from the Hofmeister series and osmolytes: effects on proteins in solution and in the crystallization process, Methods 34 (2004) 300-311; I. Jelesarov, E. Durr, R.M. Thomas, H.R. Bosshard, Salt effects on hydrophobic interaction and charge screening in the folding of a negatively charged peptide to a coiled coil (leucine zipper), Biochemistry 37 (1998) 7539-7550]. As it is known that Kv channel gating is linked to the stability of the cytoplasmic T1 multimerization domain conformation [D.L. Minor, Y.F. Lin, B.C. Mobley, A. Avelar, Y.N. Jan, L.Y. Jan, J.M. Berger, The polar T1 interface is linked to conformational changes that open the voltage-gated potassium channel, Cell 102 (2000) 657-670; B.A. Yi, D.L. Minor Jr., Y.F. Lin, Y.N. Jan, L.Y. Jan, Controlling potassium channel activities: interplay between the membrane and intracellular factors, Proc. Natl. Acad. Sci. U.S.A. 98 (2001) 11016-11023] and that intracellular chloride concentration has been linked to Kv channel kinetics [L.K. Bekar, W. Walz, Intracellular chloride modulates A-type potassium currents in astrocytes, Glia 39 (2002) 207-216; W.B. Thoreson, S.L. Stella, Anion modulation of calcium current voltage dependence and amplitude in salamander rods, Biochim. Biophys. Acta 1464 (2000) 142-150], the objective of the present study was to address how chloride concentration changes affect Kv channel kinetics more closely in an isolated expression system. Initially, no significant chloride concentration-dependent effects on channel steady-state gating kinetics were observed. Only after disruption of the cytoskeleton with cytochalasin-D did we see significant chloride concentration-dependent shifts in gating kinetics. This suggests that the shift in gating kinetics is mediated through effects of intracellular chloride concentration on cytoplasmic domain tertiary conformation as cytoskeletal interaction appears to mask the effect. Furthermore, as cytoskeletal disruption only impacts channel gating kinetics at low physiological intracellular chloride concentrations, these studies highlight the importance of paying close attention to anion concentrations used under experimental conditions.

Cell Line↗
PubMed2005

The complete mitochondrial genome of a tree frog, Polypedates megacephalus (Amphibia: Anura: Rhacophoridae), and a novel gene organization in living amphibians.

In this study, we have determined the complete sequence of the mitochondrial genome of an Old World tree frog Polypedates megacephalus (Anura: Rhacophoridae) by using a long polymerase chain reaction (PCR) technique and shotgun strategy of sequencing. The entire mtDNA sequence is 16,473 nt long with a novel mitogenomic gene organization in amphibians. Unlike other neobatrachian frogs, the transfer ribonucleic acid (tRNA)-Leu(CUN) and tRNA-Thr genes exchange their positions in P. megacephalus and form a Thr-Leu(CUN)-Pro-Phe tRNA gene tetrad. Moreover, we found that the ATP8 gene was replaced by a noncoding sequence of 853 nt long and that the ND5 gene was absent in the new mitogenome. These peculiar features of P. megacephalus mtDNA were further studied among related anuran species by PCR amplification. The new sequence data was used to assess the phylogenetic relationships of the three living amphibian orders using neighbor-joining, maximum likelihood, and Bayesian methods. In agreement with most morphological studies, phylogenetic analyses of a whole mitochondrial genome data set suggest a close relationship between salamanders and frogs. Moreover, using a molecular clock-independent Bayesian approach for inferring dating information from molecular phylogenies, we have provided a rough timescale for living amphibian evolution. This timescale provides a working framework for future paleontological researches on amphibian evolution and improves our understanding of the evolutionary history of modern amphibians.

Animals↗
PubMed2005

A computational model of the ribbon synapse.

A model of the ribbon synapse was developed to replicate both pre- and postsynaptic functions of this glutamatergic juncture. The presynaptic portion of the model is rich in anatomical and physiological detail and includes multiple release sites for each ribbon based on anatomical studies of presynaptic terminals, presynaptic voltage at the terminal, the activation of voltage-gated calcium channels and a calcium-dependent release mechanism whose rate varies as a function of the calcium concentration that is monitored at two different sites which control both an ultrafast, docked pool of vesicles and a release ready pool of tethered vesicles. The postsynaptic portion of the program models diffusion of glutamate and the physiological properties of glutamatergic neurotransmission in target cells. We demonstrate the behavior of the model using the retinal bipolar cell to ganglion cell ribbon synapse. The model was constrained by the anatomy of salamander bipolar terminals based on the ultrastructure of these synapses and presynaptic contacts were placed onto realistic ganglion cell morphology activated by a range of ribbon synapses (46-138). These inputs could excite the cell in a manner consistent with physiological observations. This model is a comprehensive, first-generation attempt to assemble our present understanding of the ribbon synapse into a domain that permits testing our understanding of this important structure. We believe that with minor modifications of this model, it can be fine tuned for other ribbon synapses.

Animals↗
PubMed2005

Maximum longevities of chemically protected and non-protected fishes, reptiles, and amphibians support evolutionary hypotheses of aging.

Evolutionary hypotheses of aging predict that species with low rates of mortality from extrinsic sources, such as predation, should senesce more slowly and have longer maximum life spans than related species with higher rates of extrinsic mortality. We tested this prediction by synthesizing information on maximum body lengths and life spans in captivity of 1193 species of chemically protected (venomous or poisonous) and non-chemically protected fishes, snakes, caudatans (salamanders and newts), and anurans (frogs and toads). In every phylogenetic group maximum longevity was positively correlated with body size and, when size was controlled for statistically, chemically protected species and genera usually had longer maximum life spans than non-protected species. These results reemphasize the importance of life history traits, particularly protection from predation, in the evolution of senescence.

Animals↗
PubMed2006

The rise of the boy-genius: psychological neoteny, science and modern life.

The mid-20th century saw the rise of the boy-genius, probably because a personality type characterized by prolonged youthfulness is advantageous both in science and modern life generally. This is the evolution of 'psychological neoteny', in which ever-more people retain for ever-longer the characteristic behaviours and attitudes of earlier developmental stages. Whereas traditional societies are characterized by initiation ceremonies marking the advent of adulthood, these have now dwindled and disappeared. In a psychological sense, some contemporary individuals never actually become adults. A child-like flexibility of attitudes, behaviours and knowledge is probably adaptive in modern society because people need repeatedly to change jobs, learn new skills, move to new places and make new friends. It seems that this adaptation is achieved by the expedient of postponing cognitive maturation - a process that could be termed psychological neoteny. ('Neoteny' refers to the biological phenomenon whereby development is delayed such that juvenile characteristics are retained into maturity.) Psychological neoteny is probably caused by the prolonged average duration of formal education, since students' minds are in a significant sense 'unfinished'. Since modern cultures favour cognitive flexibility, 'immature' people tend to thrive and succeed, and have set the tone of contemporary life: the greatest praise of an elderly person is to state that they retain the characteristics of youth. But the faults of youth are retained with well as its virtues: short attention span, sensation- and novelty-seeking, short cycles of arbitrary fashion and a sense of cultural shallowness. Nonetheless, as health gets better and cosmetic technologies improve, future humans may become somewhat like an axolotl - the cave-dwelling salamander which retains its larval form until death.

Beginning of Human Life↗
PubMed2005

D-Serine uptake by isolated retinas is consistent with ASCT-mediated transport.

Uptake of the neuromodulator D-serine by isolated larval tiger salamander (Ambystoma tigrinum) retinas was measured using capillary electrophoresis (CE). Excised retinas were incubated in Ringer's solution in the presence of 5 microM D-serine. The supernatant was removed after 30 min, mixed with 4-fluoro-7-nitrobenz-2-oxa-1,3-diazole (NBD-F) to fluorescently label amines and analyzed using CE. Significant D-serine uptake was observed over a period of 1.5h. This is the first observation of D-serine uptake by an intact retinal tissue. D-Serine uptake in the retina was Na(+)-dependent and blocked by l-alanine, l-threonine, and l-cysteine. This pharmacology is consistent with the sodium dependent heteroexchange expected of system ASC-type transporters.

4-Chloro-7-nitrobenzofurazan↗
PubMed2005

Redundancy in the population code of the retina.

We have explored the manner in which the population of retinal ganglion cells collectively represent the visual world. Ganglion cells in the salamander were recorded simultaneously with a multielectrode array during stimulation with both artificial and natural visual stimuli, and the mutual information that single cells and pairs of cells conveyed about the stimulus was estimated. We found significant redundancy between cells spaced as far as 500 mum apart. When we used standard methods for defining functional types, only ON-type and OFF-type cells emerged as truly independent information channels. Although the average redundancy between nearby cell pairs was moderate, each ganglion cell shared information with many neighbors, so that visual information was represented approximately 10-fold within the ganglion cell population. This high degree of retinal redundancy suggests that design principles beyond coding efficiency may be important at the population level.

Animals↗
PubMed2005

Release of L-aspartate by reversal of glutamate transporters.

Aspartate is released in the brain during metabolic inhibition and can activate NMDA receptors. We compared the characteristics of aspartate and glutamate release mediated by reversed operation of GLAST glutamate transporters in salamander retinal glial cells, when high [K(+)](o) solution was applied to mimic the ionic conditions of stroke or glaucoma. In the absence of Cl(-), to isolate the transport-associated current of the transporters, reversed uptake of aspartate and glutamate had similar characteristics. Both were increased strongly by depolarisation, inhibited by the transport inhibitor TBOA (DL-threo-beta-benzyloxyaspartate), and activated in a first order manner by intracellular amino acid (in the presence of 20mM [Na(+)](i)) with an EC(50) of 0.8mM for aspartate and 2.3mM for glutamate. In stroke the extracellular pH shifts acid by around a pH unit: this reduced the release of aspartate and glutamate by reversed uptake by a factor of 8-20. The external Cl(-) concentration had only a small effect on the current associated with reversed uptake of aspartate and glutamate. Tamoxifen, which reduces amino acid release through swelling-activated anion channels in glial cells, was found to inhibit reversed uptake with an IC(50) which was >100 microM. Part of the activation of NMDA receptors which occurs in ischaemia is likely to reflect the release of aspartate by reversed uptake.

Amino Acid Transport System X-AG↗
PubMed2006

The dynamic range and domain-specific signals of intracellular calcium in photoreceptors.

Vertebrate photoreceptors consist of strictly delimited subcellular domains: the outer segment, ellipsoid, cell body and synaptic terminal, each hosting crucial cellular functions, including phototransduction, oxidative metabolism, gene expression and transmitter release. We used optical imaging to explore the spatiotemporal dynamics of Ca(2+) signaling in non-outer segment regions of rods and cones. Sustained depolarization, designed to emulate photoreceptor activation in the darkness, evoked a standing Ca(2+) gradient in tiger salamander photoreceptors with spatially-averaged intracellular Ca(2+) concentration within synaptic terminals of approximately 2 microM and lower (approximately 750 nM) intracellular calcium concentration in the ellipsoid. Measurements from axotomized cell bodies and isolated ellipsoids showed that Ca(2+) enters the two compartments via both local L-type Ca(2+) channels and diffusion. The results from optical imaging studies were supported by immunostaining analysis. L-type voltage-operated Ca(2+) channels and plasma membrane Ca(2+) ATPases were highly expressed in synaptic terminals with progressively lower expression levels in the cell body and ellipsoid. These results show photoreceptor Ca(2+) homeostasis is controlled in a region-specific manner by direct Ca(2+) entry and diffusion as well as Ca(2+) extrusion. Moreover, quantitative measurement of intracellular calcium concentration levels in different photoreceptor compartments indicates that the dynamic range of Ca(2+) signaling in photoreceptors is approximately 40-fold, from approximately 50 nM in the light to approximately 2 microM in darkness.

Animals↗
PubMed2005

Identification of cDNA coding for a homologue to mammalian leptin from pufferfish, Takifugu rubripes.

We identified cDNA coding for a homologue to mammalian leptin in puffer, Takifugu rubripes, using genomic synteny around the human leptin gene. In addition to significant sequence homologies, the puffer leptin (pLEP) displays characteristic structural features in common with mammalian leptin. The pLEP mRNA was expressed mostly in the liver that contained abundant lipids. In addition, homologues to pLEP were found in the databanks for three fish species (salmon, medaka, and Tetraodon) and two amphibians (salamander and Xenopus). The phylogenetic analysis shows rapid rates of molecular divergence among leptins from different vertebrate classes, but not between mammals and avians.

Amino Acid Sequence↗
PubMed2005

Developmental genetic basis for the evolution of pelvic fin loss in the pufferfish Takifugu rubripes.

Paired appendages were a key developmental innovation among vertebrates and they eventually evolved into limbs. Ancient developmental control systems for paired fins and limbs are broadly conserved among gnathostome vertebrates. Some lineages including whales, some salamanders, snakes, and many ray-fin fish, independently lost the pectoral, pelvic, or both appendages over evolutionary time. When different taxa independently evolve similar developmental morphologies, do they use the same molecular genetic mechanisms? To determine the developmental genetic basis for the evolution of pelvis loss in the pufferfish Takifugu rubripes (fugu), we isolated fugu orthologs of genes thought to be essential for limb development in tetrapods, including limb positioning (Hoxc6, Hoxd9), limb bud initiation (Pitx1, Tbx4, Tbx5), and limb bud outgrowth (Shh, Fgf10), and studied their expression patterns during fugu development. Results showed that bud outgrowth and initiation fail to occur in fugu, and that pelvis loss is associated with altered expression of Hoxd9a, which we show to be a marker for pelvic fin position in three-spine stickleback Gasterosteus aculeatus. These results rule out changes in appendage outgrowth and initiation genes as the earliest developmental defect in pufferfish pelvic fin loss and suggest that altered Hoxd9a expression in the lateral mesoderm may account for pelvis loss in fugu. This mechanism appears to be different from the mechanism for pelvic loss in stickleback, showing that different taxa can evolve similar phenotypes by different mechanisms.

Animals↗
PubMed2005

Historical perspective: Hormonal regulation of behaviors in amphibians.

This review focuses on research into the hormonal control of behaviors in amphibians that was conducted prior to the 21st century. Most advances in this field come from studies of a limited number of species and investigations into the hormonal mechanisms that regulate reproductive behaviors in male frogs and salamanders. From this earlier research, we highlight five main generalizations or conclusions. (1) Based on studies of vocalization behaviors in anurans, testicular androgens induce developmental changes in cartilage and muscles fibers in the larynx and thereby masculinize peripheral structures that influence the properties of advertisement calls by males. (2) Gonadal steroid hormones act to enhance reproductive behaviors in adult amphibians, but causal relationships are not as well established in amphibians as in birds and mammals. Research into the relationships between testicular androgens and male behaviors, mainly using castration/steroid treatment studies, generally supports the conclusion that androgens are necessary but not sufficient to enhance male behaviors. (3) Prolactin acts synergistically with androgens and induces reproductive development, sexual behaviors, and pheromone production. This interaction between prolactin and gonadal steroids helps to explain why androgens alone sometimes fail to stimulate amphibian behaviors. (4) Vasotocin also plays an important role and enhances specific types of behaviors in amphibians (frog calling, receptivity in female frogs, amplectic clasping in newts, and non-clasping courtship behaviors). Gonadal steroids typically act to maintain behavioral responses to vasotocin. Vasotocin modulates behavioral responses, at least in part, by acting within the brain on sensory pathways that detect sexual stimuli and on motor pathways that control behavioral responses. (5) Corticosterone acts as a potent and rapid suppressor of reproductive behaviors during periods of acute stress. These rapid stress-induced changes in behaviors use non-genomic mechanisms and membrane-associated corticosterone receptors.

Amphibians↗