Scanning and transmission electron microscopic study of the lung of the newt,Triturus alpestris Laur.

Summary The lungs of Triturus alpestris Laur. were investigated with the scanning and transmission electron microscopes. Dimensions of the cell bodies of pneumocytes and ciliated cells, as well as the thickness of the air-blood barrier, were determined. The lungs of the newt form two simple sacs without septa. A ciliated epithelium containing goblet cells lines the pulmonary vein and partially the pulmonary artery. The remainder of the lung surface is covered internally by respiratory epithelium consisting of one type of cell and only occasionally showing the presence of single ciliated cells. All cells, ciliated, goblet and pneumocytes, contain in their cytoplasm lamellar bodies. Multivesicular bodies and numerous vesicles of variable electron density also occur in the cytoplasm of pneumocytes. Atypical mitochondria can be found in all cell types of the lung. Fixation with addition of tannic acid reveals the surface lining film. Tubular myelin figures were not observed.     

Distribution patterns of the paraneuronal endocrine cells in the skin, gills and the airways of fishes as determined by immunohistochemical and histological methods

Summary The neuro-endocrine cells of fish skin and respiratory surfaces, and their bioactive secretion as far as is known, are reviewed, and compared with similar elements in tetrapods, particularly amphibians. In the skin of teleost fish, immunohistochemistry has shown that Merkel cells react for serotonin, neuron-specific enolase and enkephalins. The pharmacology is not established in dipnoans or lampreys. In some teleosts, neuromasts react for substance P and leu-enkephalins; substance P is also reported from some ampullary organs (electroreceptors). Taste buds of teleosts may react for enkephalin and substance P. Basal cells of taste buds react for serotonin and neuron-specific enolase. Some unicellular skin glands of teleosts express bioactive compounds, including serotonin and some peptides; this ectopic expression is paralleled in amphibian skin glands. The dipnoan Protopterus has innervated pulmonary neuro-endocrine cells in the pneumatic duct region with dense-cored vesicles. In Polypterus and Amia the lungs have serotonin-positive neuro-endocrine cells that are apparently not innervated. In fish gills, a closed type of neuro-endocrine cell reacts for serotonin, an open type for enkephalins and some calcium-binding proteins (calbindin, calmodulin and S-100 protein). The functions of neuro-endocrine cells in fishes await investigation, but it is assumed they are regulatory.     

Endocrine-like cells in the lungs of the newt,Triturus alpestris Laur.

Summary Endocrine-like cells (ELC) scattered in the basal part of the ciliated epithelium were identified in the lungs of the newt, Triturus alpestris. These ELC have a clear cytoplasm containing large amounts of secretory vesicles (69–180 nm in diameter), especially in their basal parts, but do not display formaldehyde-induced fluorescence. The ELC may be associated with nerve fibres.     

Chromosome banding in amphibia

The distribution of constitutive heterochromatin on the chromosomes of Triturus a. alpestris, T. v. vulgaris and T. h. helveticus (Amphibia, Urodela) was investigated. Sex-specific chromosomes were determined in the karyotypes of T. a. alpestris (chromosomes 4) and T. v. vulgaris (chromosomes 5). The male animals have one heteromorphic chromosome pair, of which only one homologue displays heterochromatic telomeres in the long arms; the telomeres of the other homologue are euchromatic. This chromosome pair is always homomorphic and without telomeric heterochromatin in the female animals. There is a highly reduced crossing-over frequency between the heteromorphic chromosome arms in the male meiosis of T. a. alpestris; in T. v. vulgaris no crossing-over at all occurs between the heteromorphic chromosome arms. No heteromorphisms between the homologues exist on the corresponding lampbrush chromosomes of the female meiosis. In T. h. helveticus no sex-specific heteromorphism of the constitutive heterochromatin could be determined. The male animals of this species, however, already possess a chromosome pair with a greatly reduced frequency of chiasma-formation in the long arms. The C-band patterns and the pairing configurations of the sex-specific chromosomes in the male meiosis indicate an XX/XY-type of sex-determination for the three species. A revision of the literature about experimental interspecies hybridizations, gonadic structure of haploid and polyploid animals, and sex-linked genes yielded further evidence in favor of male heterogamety. The results moreover suggest that the heterochromatinization of the Y-chromosome was the primary step in the evolution of the sex chromosomes.     

The assignment of a Thinopyrum distichum (Thunb.) Löve-derived translocation to the long arm of wheat chromosome 7D using endopeptidase polymorphisms

Summary Endopeptidase zymograms of the translocation line Indis revealed the presence of several major and minor bands that had differential expression in coleoptile and seed tissues. While Indis lacks Ep-D1a, which is present in the parental cultivar Inia 66, it also may not express any of the Th. distichum bands. The Indis zymogram was found to be identical to that of an isogenic line of Inia 66 possessing Lr19. Since the absence of an Ep-D1a product appears to be linked to the 7DL translocation, it is possible to use the null condition as a marker for both the Lr19 or Indis translocations. The Indis translocation also did not show recombination with the cn-D1 chlorophyl mutant on 7DL, confirming that a part of 7D was involved. The results of a telocentric mapping experiment involving the 7D telosomes indicated that in Indis a chromosome segment from Th. distichum replaced a large section of 7DL of Inia 66.     

Surpellic films, lung surfactant, and their cellular origin in newt, caecilian, and frog

The lung lining and its surfactant have been studied in one species from each of the amphibian sub-classes Anura, Urodela, and Apoda. The surfactant was studied by observation of bubbles derived from the lungs, the morphology by electron microscopy. Other details of the apodan are also given.
The surfactant layers of Rana temporaria and Ichthyophis (species unknown) resemble one another, being somewhat less stable than that of mammals. That of Triturus vulgaris is even less stable, and shows qualitative differences from the other two. Nevertheless, it can reduce the surface tension to 2 mN/m.
The epithelial cells of amphibian lungs cannot be divided into two clearly defined types, as can those of mammals. Microvilli are present on the alveolar surface of all these cells. In the lungs of Rana and Ichthyophis the lining cells contain numerous lamellated osmiophilic bodies (which are intracellular depots of surfactant); in the former their appearance is mainly cross-barred, in the latter mainly concentric. Similar bodies exist in the lung epithelial cells of Triturus but are relatively rare.     

Die Ausbildung der Asymmetrie der Nuclei habenulae des Zwischenhirns bei Amphibien in Unabhängigkeit vom Blutkreislauf

Zusammenfassung Zur Prüfung der Fräge, ob der Blutkreislauf bei der Bestimmung der Asymmetrie der Nuclei habenulae des Zwischenhirns eine Rolle spielt, wurden Larven vonTriturus alpestris ohne Blutkreislauf durch Entfernung der Herzanlage hergestellt. Die Nuclei habenulae entwickelten auch ohne Kreislauf ihre normale Asymmetrie.Isolate aus dem Kopfbezirk der Neurula vonTriturus alpestris, die aus cephalem Ekto und Mesoderm bestanden und manchmal auch vorderes Entoderm enthielten, entwickelten in günstigen Fallen an ihrem Gehirn die Nuclei habenulae. Diese besaßen trotz Abwesenheit von Blutkreislauf und der anderen asymmetrischen Organe die normale Asymmetrie.

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The development of asymmetrical nuclei habenulae in the diencephalon of amphibians, independent of blood circulation

Summary The problem whether the blood circulation influences or determines the asymmetry of Nuclei habenulae was tested by removing the heart-anlage of the embryo ofTriturus alpestris. The resulting larvae without any blood circulation developed their normal asymmetry of habenulae.Isolates from the head region of neurulae ofTriturus alpestris, consisting of cephalic ecto and mesoderm, sometimes also of entoderm, under favorable conditions could develop brains with Nuclei habenulae. These were normal, asymmetric in spite of missing the blood circulation and other asymmetrical organs.

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Ausgefürhrt mit Unterstützung der Deutschen Forschugagemeinschaft.     

Serotonin-like immunoreactive cells in the pulmonary epithelium of ancient fish species

The pulmonary mucosa of three species of ancient fish was studied immunohistochemically to show the distribution of serotonin, regarded as the main monoamine of mammalian bronchopulmonary paraneurons. Serotonin-like immunoreactive cells, dispersed through the airway epithelium as single cells, were found in all the fish species studied. They are presumably equivalent to the neuroendocrine cells reported in the lungs of mammalian and submammalian vertebrates. However, the precise role and the function of these cells remain unknown. Since the species studied belong to the most primitive extant groups of ancient fish, the present investigation suggests that serotonin is widely distributed in the lungs of the vertebrates. Several peptides, known to be specific cytochemical markers for the identification of the pulmonary neuroendocrine cells of mammals, are being investigated in the lungs of the fish species studied. They may help to trace the phylogeny of the pulmonary neuroendocrine cell system and to elucidate its function in lower vertebrates.     

Serotonin-like immunoreactive cells in the pulmonary epithelium of ancient fish species

Summary The pulmonary mucosa of three species of ancient fish was studied immunohistochemically to show the distribution of serotonin, regarded as the main monoamine of mammalian bronchopulmonary paraneurons. Serotonin-like immunoreactive cells, dispersed through the airway epithelium as single cells, were found in all the fish species studied. They are presumably equivalent to the neuroendocrine cells reported in the lungs of mammalian and submammalian vertebrates. However, the precise role and the function of these cells remain unknown. Since the species studied belong to the most primitive extant groups of ancient fish, the present investigation suggests that serotonin is widely distributed in the lungs of the vertebrates. Several peptides, known to be specific cytochemical markers for the identification of the pulmonary neuroendocrine cells of mammals, are being investigated in the lungs of the fish species studied. They may help to trace the phylogeny of the pulmonary neuroendocrine cell system and to elucidate its function in lower vertebrates.     

Di‐ and tetranucleotide microsatellite markers for the Alpine newt (Triturus alpestris): characterization and cross‐priming in five congeners

We developed a set of di‐ and tetranucleotide microsatellite markers for the Alpine newt, Triturus alpestris. Polymorphism as detected in 39 individuals ranged from 3 to 32 alleles at a locus. Cross‐priming with samples of five other Triturus species showed extremely poor levels of cross‐species utility. Still, these markers are suitable for studies of inter‐ and intrapopulation genetic diversity in the focal species.     

Gesamtbestand an organischen Substanzen der Spinne Tegenaria atrica im Vergleich zu Protophormia terrae novae (Diptera) und Orconectes limosus (Crustacea,Decapoda)

Zusammenfassung Die Pulsationen der Lymphherzen von Normaltieren von Triturus vulgaris L. und T. alpestris Laur. sind unregelmäßig und verändern sich bei Erregung der Tiere. Die Lymphherzfrequenzen sind temperaturabhängig und folgen der R-G-T-Regel. Der Q ₁₀ beträgt bei T. vulgaris 2,5, bei T. alpestris 2,1. In Narkose oder bei Spinaltieren sinkt die Lymphherzfrequenz gegenüber der von Normaltieren ab, der Rhythmus wird regelmäßig. Bei Spinaltieren tritt homolaterale Synchronie auf. Jedes Lymphherz hat ein eigenes motorisches Zentrum im Rückenmark; die Impulse werden über Spinalnerven geleitet. Die Ergebnisse werden mit den von Anuren bekannten verglichen.

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Physiological investigations on the lymph hearts of urodelans

Summary The heart beats of normal individuals of Triturus vulgaris L. and T. alpestris Laur. are irregular and can be altered by excitation of the animal. The frequencies vary with alterations in temperature according to the rule of van t'Hoff. The Q ₁₀ was measured as 2.5 for T. vulgaris and 2.1 for T. alpestris. In anesthetized and in spinal animals the frequencies are strongly reduced and the pulsations become regular. Spinal animals show homolateral synchrony of the lymph heart beats. The motor centres, separated for each lymph heart, are situated in the spinal cord; the impulses are mediated by spinal nerves. The results are compared with those of the Anurans.

     

The distribution of GABA-like-immunoreactive neurons in the brain of the newt,Triturus cristatus carnifex,and the green frog,Rana esculenta

Summary The distribution of -aminobutyric acid (GABA) immunoreactivity was studied in the brain of two amphibian species (Triturus cristatus carnifex, Urodela; Rana esculenta, Anura) by employing a specific GABA antiserum. A noteworthy immunoreactive neuronal system was found in the telencephalic dorsal and medial pallium (primordium pallii dorsalis and primordium hippocampi) and in the olfactory bulbs. In the diencephalic habenular nuclei there was a rich GABAergic innervation, and immunoreactive neurons were observed in the dorsal thalamus. In the hypothalamus the GABA immunoreactivity was found in the preoptic area, the paraventricular organ and in the hypothalamo-hypophysial complex. In the preoptic area of the frog some GABA-immunoreactive CSF-contacting cells were shown. In the optic tectum immunolabeled neurons were present in all the cellular layers. A rich GABAergic innervation characterized both the fibrous layers of the tectum and the neuropil of the tegmentum and interpeduncular nucleus. In the cerebellum, in addition to the Purkinje cells showing a variable immunopositivity, some immunoreactive cell bodies appeared in the central grey. Abundant immunolabeled nerve fibers in the acoustico-lateral area and some immunopositive neurons in the region of the raphe nucleus were observed. In conclusion, the GABAergic central systems, well-developed in the amphibian species studied, were generally characterized by close similarities to the pattern described in mammals.Dedicated to Professor Valdo Mazzi (Dipartimento di Biologia Animale, Università di Torino), in honor of his 70th birthday     

Asymmetric imine-ene reactions: Diastereofacial selective reactions with chiral glyoxylate-derivedα-imino esters and asymmetric catalysis of enantiofacial selective reactions with prochiralα-imino esters

Summary The diastereofacial selective imine-ene reactions with-imino esters, prepared from (–)-8-phenylmenthyl glyoxylate, are shown to provide an efficient entry to the asymmetric synthesis of-amino acids. The feasibility study of the asymmetric catalysis is also reported on the enantiofacial selective ene reactions with prochiral-imino esters.     

L'action du dioxyde de sélénium sur des œufs d'amphibiens d'espèces résistantes

The eggs of some Amphibian species (Triturus helveticus Raz., Triturus alpestris Laur., Xenopus laevis Daud.) are more resistant to selenium dioxyde M/100 than eggs of Pleurodeles waltlii Michah. Minimal mitotic abnormalities are observed: 1. “empty” poles, 2. cupular poles, 3. escape of centrosphere, which separates from the spindle, and sometimes torsion of polar regions of the achromatic figure. Cytasters and newly formed spindles are also observed in the diastematic clear zone, which is dissociated and inefficient. These abnormalities are believed to be consequences of the progressive diminution of a dominant factor associated with the centriole. When dominance is weakened, secondary poles appear with or without relation to the spindle. The balance between fibrillogenesis and reproduction of centrioles is newly demonstrated. These mechanisms are compared with phenomena occuring during fertilization and parthenogenesis     

Untersuchungen an Oozytenkernen von Amphibien und Teleosteern mit Pseudoisozyanin

Zusammenfassung Das fluoreszenzmikroskopische Nachweisverfahren für DNS-Nukleoproteide mit N,N-Diäthylpseudoisozyaninchlorid (PIC-Reaktion nach Sterba, 1963) wird auf seine Spezifität für DNS und auf den zugrundeliegenden Mechanismus untersucht.Es wird der Nachweis erbracht, daß nur solche Strukturen dargestellt sind, deren DNS-Gehalt durch Vergleichsfärbungen oder durch spezifische Extraktionen gesichert werden kann. Als wesentlich für das Zustandekommen der Reaktion werden die Phosphatgruppen der DNS erkannt. Eine mögliche Mitbeteiligung von SH-Gruppen und Disulfidbrücken am Zustandekommen der Reaktion wird diskutiert. Es wird der Nachweis erbracht, daß RNS bei der PIC-Reaktion aus den Schnitten extrahiert wird.Vergleiche mit anderen Nukleinsäurenachweisen zeigen, daß die PIC-Reaktion diesen überlegen ist.Im Oozytenkern von Triturus alpestris konnten extrachromosomale DNS enthaltende Strukturen (Mikronukleolen) nachgewiesen werden.Morphologie und Histochemie der Oozytenkerne einiger Amphibien und Fische werden vergleichend untersucht.

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Studies on oocyte nuclei of amphibians and teleosts by means of pseudoisocyanine

Summary The fluorescence-microscopical method for demonstration of DNA-proteids with N,N-diethyl-pseudoisocyanine chloride is investigated for specificity for DNA and for the chemism of it. It is stated that only such structures are demonstrated which contain DNA.The phosphoric acid groups in the DNA are responsible for the positive results with the pseudoisocyanine-reaction. The possibility is discussed that SH and SS-groups of the residual protein may be responsible for the positive reaction, too.It is suggested, that RNA is extracted by the manipulations during the pseudoisocyanine-method.In oocyte nuclei of Triturus alpestris we were able to demonstrate extrachromosomal DNA-containing structures (micronucleoli). Morphology and histochemistry of the oocyte nuclei of some Amphibiens and fishes are investigated comparatively.

     

Antibodies to neurofilament protein and other brain proteins reveal the innervation of peripheral organs

Summary Monoclonal and polyclonal antibodies to neurofilament proteins, neuron-specific enolase, glial fibrillary acidic protein and S-100 have been used to demonstrate nerves, ganglion cells and the supportive glial system of the innervation of various organs. The female genitalia, the urinary tract, the respiratory system, the pancreas, the heart and the skin of several mammalian species, including rat, mouse, guinea pig, cat, pig, monkey and man were fixed in parabenzoquinone and portions of each organ were snap frozen. Serial or free-floating thick cryostat sections were stained using indirect immunofluorescence and peroxidase anti-peroxidase immunocytochemistry. In addition, the newly described and highly sensitive immunogold-silver staining technique was used on Bouin's-fixed and wax-embedded tissues.Antibodies to neurofilament proteins seemed to react with neuronal structures in all the species studied. Alternately stained serial sections showed a similar distribution of neurofilament proteins and neuron-specific enolase-containing nerves. Neuron-specific enolase staining had a diffuse appearance and was found to be highly variable, indicating that the neuron-specific enolase content might be related to the physiological state of the nerves and ganglion cells, whereas antibodies to neurofilament protein gave a consistently intense and very clear picture of the ganglion cells and nerve fibres. Antibodies to S-100 stained supportive elements of the peripheral nervous system in all tissues examined, whereas antibodies to glial fibrillary acidic protein were more selective.Abbreviations GFAP glial fibrillary acidic protein - NSE neuron-specific enolase - PBS phosphate-buffered saline - PAP peroxidase anti-peroxidase - FITC fluorescein-isothiocyanate     

Micro-habitat use within a guild of newt larvae (Trituridae) in an Alpine lake

Population and ecological parameters such as numbers of larvae, microhabitat use, niche breadth and niche overlap of three species of syntopic larval newts (Alpine newt Triturus alpestris, Italian crested newt T. carnifex, and common newt T. vulgaris) were studied for two years in a small pond at 1160 m a.s.l. in NE Slovenia. Differences in microhabitat partitioning among larval newts were small. The largest niche breadth was estimated for larval T. alpestris, and the narrowest estimate was for larval T. carnifex in both years. Ecological differences seem to be very small and quite variable among sites and years. It appears that the developmental stage and size of newt larvae are more important in explaining resource partitioning than the characteristics of each species. Because of the absence of potential invertebrate predators and adult newts in the second half of the breeding season, the injuries could only be caused by intra-and interspecific predation attempts.     

Changes of the cell surface charge of amphibian ectoderm after induction

Summary Isolated ectoderm of early gastrula stages ofTriturus alpestris was treated with vegetalizing factor for 24 h employing the sandwich method (induced ectoderm). Controls were incubated for the same period with -globulin which has no inducing activity. Explants of both series were labelled with cationized ferritin, which binds to negatively charged groups at physiological pH. In non-induced ectoderm, ferritin particles can be found as a thin layer all over the plasma membranes. In induced ectoderm the total amount of ferritin bound to the plasma membrane is much lower than in non-induced ectoderm. Ferritin is located in restricted areas only. In contrast to the controls, other membrane areas are free of ferritin particles. The correlation between these results and the change of cell affinity after induction with vegetalizing factor is discussed.     

Skull of the neotenic salamandrid amphibian Triturus alpestris and abbreviated development in the Tertiary Salamandridae

The skull of neotenic individuals of the Alpine Newt Triturus alpestris from the locality Drakolimni (Greece) is described on the basis of models made from magnified serial frontal sections. In order to recognize features associated with neoteny and paedomorphosis, the results were compared with normal development of contemporary Triturus and of other Caudata. The neotenic larvae from Greece correspond to advanced stages of metamorphosis in normal development of Triturus alpestris. Comparison with salamandrids from the Tertiary of Europe in which the hyobranchial skeleton was preserved, namely Brachycormus noachicus, Chelotriton paradoxus, and Palaeopleurodeles hauffi, revealed that both latter taxa were completely metamorphosed adults anatomically similar to their contemporary relatives Tylototriton, Echinotriton, and Pleurodeles, whereas Brachycormus, though apparently related to Chelotriton, was a neotenic amphibian. This is suggested by its incompletely metamorphosed but ossified hyobranchial skeleton. © 1996 Wiley-Liss, Inc.