Chromosome 1 in crested and marbled newts (Triturus)

The heteromorphic chromosomes 1 of Triturus cristatus carnifex and T. marmoratus were studied in mitotic metaphase after staining with the Giemsa C-banding technique and with the fluorochromes, DAPI (AT-specific) and mithramycin (GC-specific). They were also examined in the lampbrush form under phase-contrast before fixation and after fixation and staining with Giemsa. Chromosomes 1 of T.c. carnifex are asynaptic and achiasmatic throughout most of their long arms. They are also heteromorphic in most of their long arms for the patterns of Giemsa and fluorochrome staining and the distribution of distinctive lampbrush loops. The heteromorphic regions correspond to the regions that are asynaptic and achiasmatic. They stain more strongly with mithramycin and more weakly with DAPI than the remainder of the chromosomes, signifying that their DNA is relatively rich in GC. The patterns of staining with Giemsa and fluorochromes and the distributions of distinctive lateral loops vary from one animal to another in the same species and even in the same population. The asynaptic and achiasmatic regions of chromosomes 1 in T. marmoratus extend throughout the whole of the long arms and well beyond the heterochromatic region. Chiasmata form only in the short arm and occasionally in the short euchromatic segment at the tip of the long arms. The staining patterns of chromosomes 1 in T. marmoratus differ from those in T.c. carnifex although, like carnifex, their DNA is relatively GC-rich. The chromosomes 1 of T. marmoratus are more submetacentric than those of T.c. carnifex. In T. marmoratus chromosome 1B is about 12% shorter than 1A. There is a short paracentric inversion heterozygosity in the long arm of chromosome 1B in T. marmoratus which probably accounts for the lack of chiasmata in the euchromatin that separates the centromere from the start of the heterochromatin. In both carnifex and marmoratus, embryos that are homomorphic for chromosome 1 arrest and die at the late tailbud stage of development. The same applies to F₁ hybrid embryos T.c. carnifex x T. marmoratus, and this has permitted identification of chromosomes 1A and 1B in both species. There is no correspondence between patterns of Giemsa or fluorochrome staining of the heteromorphic regions of chromosome 1 and any feature of the lampbrush chromosomes. However, the short euchromatic ends of the long arms of chromosomes 1 in both species are distinguished in the lampbrush form by a series of uniformly small loops of fine texture associated with very small chromomeres. The Giemsa C-staining patterns of both chromosomes 1A and 1B are different in each of the four subspecies of T. cristatus. T.c. karelinii stands out by having unusually large masses of Giemsa C-staining centromeric heterochromatin on all but 1 of its 12 chromosomes. A scheme is proposed for the evolution of chromosome 1 in T. cristatus and T. marmoratus, based on all available cytological and molecular data.     

Microvascular anatomy of testes in the adult pipid frog,Xenopus laevis Daudin: A scanning electron microscopic study of vascular corrosion casts

The three-dimensional microvascular anatomy of the testes of adult Xenopus laevis was studied by scanning electron microscopy of vascular corrosion casts. Results showed that testicular arteries branch off from urogenital arteries and approach the testes via the mesorchium. At the dorsal surface of testes, arteries course towards the medial and lateral surfaces of the ovoid testes, pierce the tunica albuginea and run in the subalbugineal space and then penetrate the testicular parenchyma. In the parenchyma, they branch in all directions. At sites where larger branches arise, sphincters are present. Intimal cushions are found at the origin of smaller branches. Terminal arterioles capillarize and form a wide-meshed capillary network around convoluted seminiferous tubules. Inter- and peritubular capillaries cannot be differentiated reliably. Arterio-venous transition distances are short. Arterio-arterial anastomoses and veno-venous anastomoses are present. Testicular venules empty into a venous subalbugineal plexus which shows veno-venous anastomoses of different lengths and calibres. Signs of the ongoing non-sprouting angiogenesis, particularly intussusceptive branch remodelling, reflect the dynamics of the testicular parenchyma. The present study is the first that demonstrates the testicular microvascular anatomy in an anuran species and shows that X. laevis and men share the intricate convoluted seminiferous tubules with associated vascular patterns as a feature in common.     

Ontogeny of skull size and shape changes within a framework of biphasic lifestyle: a case study in six <Emphasis Type="Italic">Triturus</Emphasis> species (Amphibia,Salamandridae)

As with many other amphibians, Triturus species are characterized by a biphasic life cycle with abrupt changes in the cranial skeleton during metamorphosis. The post-metamorphic shape changes of the cranial skeleton were investigated using geometric morphometric techniques in six species: Triturus alpestris, T. vulgaris, T. dobrogicus, T. cristatus, T. carnifex, and T. karelinii. The comparative analysis of ontogenetic trajectories revealed that these species have a conserved developmental rate with divergent ontogenetic trajectories of the ventral skull shape that mainly reflect phylogenetic relatedness. A striking exception in the ontogenetic pattern was possibly found in T. dobrogicus, characterized by a marked increase in the developmental rate compared to the other newt species. The size-related shape changes explained a large proportion of shape change during post-metamorphic growth within each species, with marked positive allometric growth of skull elements related to foraging.     

The Different Vascular Patterns of Slime Glands in the Hagfishes,Myxine glutinosa Linnaeus and Eptatretus stouti Lockington A Scanning Electron Microscope Study of Vascular Corrosion Casts*

The vascular patterns of the epidermally derived slime glands of the Atlantic hagfish, Myxine glutinosa, and of the Pacific hagfish, Eptatretus stouti, have been studied by scanning electron microscopy of vascular corrosion casts. In Myxine a simple two-dimensional vascular network sheaths the slime glands, while in Eptatretus there is also a great number of capillary loops of different lengths arising from the sheathing network and extending into the interior of the glands. These basic differences in slime glands vascular patterns are thought to reflect substantially different physiological behaviour of the slime glands in Myxine and Eptatretus.     

Silver staining of the lampbrush chromosomes ofTriturus cristatus carnifex

Lampbrush chromosomes fromTriturus cristatus carnifex were stained using the ammoniacal silver staining (AgAS) technique. Many of the recognized marker structures proved to be silver positive, plus between six and fifteen lateral loop pairs. None of the stained loop pairs corresponded to known sites of the nucleolus organizers, although the extrachromosomal nucleoli were silver positive. The ammoniacal silver staining technique does not demonstrate the specificity for active ribosomal cistrons in lampbrush chromosomes that it does in a wide variety of mammalian mitotic chromosomes.     

Lampbrush chromosomes and chiasmata of sex-reversed crested newts

Triturus cristatus carnifex provides a particularly clear example of sexual dimorphism for chiasma frequency and localisation. Oocytes from normal XX females routinely carry one proximal chiasma on each arm of their lampbrush bivalents. Spermatocytes from normal XY males have more numerous and relatively distal chiasmata. Lampbrush chromosomes from the oocytes of sex-reversed XY neofemales are found to resemble those from normal oocytes in having one proximal chiasma on each bivalent arm. A comparison of particular markers on the heteromorphic long arm of chromosome 1 provides evidence to equate the lampbrush 1A to somatic 1A, and confirms previous reports that lampbrush chromosome 1A is slightly longer than 1B. The XY sex bivalent of neofemales does not show any obvious heteromorphy of recognised marker loops. Received: 9 September 1997 / Accepted: 16 October 1997     

Microvascularization and histomorphology of lateral line organs in adult Xenopus laevis

The microvasculariaztion of the lateral line organs (LLOs) of the adult pipid frog, Xenopus laevis was studied by scanning electron microscopy of vascular corrosion casts (VCCs) and correlative light microscopy of paraplast embedded tissues sections. Scanning electron micrographs of VCCs revealed that each neuromast within the LLO rests on a distinct bowl‐like capillary network (vascular bowl). One to three vascular bowls were supplied by an ascending arteriole and drained by a descending venule towards the skin deep dermal vascular network. Blood flow regulation mechanisms in form of intimal cushions were present at the origin of ascending arterioles supplying LLOs, microvenous valves were present at the confluence of deep dermal venules and veins. This together with sprouting and nonsprouting angiogenesis (intussusceptive microvascular growth) found in vascular bowls demonstrate that in adult Xenopus the capillary bed of LLO's still can be adjusted to changing energetic needs. J. Morphol. 275:497–503, 2014. © 2013 Wiley Periodicals, Inc.     

Scanning electron microscopic study of the capillary loops in the dermal papillae. Skin of the hand of the Japanese monkey (Macaca fuscata)

The microvasculature of the skin of the hand of Japanese monkeys was examined by means of scanning electron microscopy of corrosion casts. The vasculature of all areas of the skin of the hand was examined and divided into three structures excluding the nail bed: (1) In the ball of the finger, the typical structure of the capillary loops was studied. Capillary loops were formed out of not just one capillary vessel, but two or three vessels. Each capillary vessel arose and divided into several branches at the papilla, and they became descending limbs. After the loop passed a hairpin turn, the descending limbs were 1.5 times larger than the ascending limbs in the intrapapillary portion, and they became extrapapillary venules. The descending limbs connected with the postcapillary venules in the postpapillary portion and with the horizontal network. The postcapillary venules fused with each other (1-5 loops) to form the primary and secondary venous arcades. (2) In the thenar eminence, the capillary loops were a little lower, and their grooves were wider than in the ball of the finger. The characteristic structure in this area was the interpapillar capillary network. (3) In the lateral side of the finger, the number of capillary loops formed by the arterial capillary network of the subepidermal layer was smaller. The capillary loops here had the lowest height and a simple structure.     

Microvascular anatomy of the brain of the adult pipid frog,Xenopus laevis (Daudin): A scanning electron microscopic study of vascular corrosion casts

To demonstrate the 3D microvascular anatomy of the brain of the model organism Xenopus laevis Daudin scanning electron microscopy of vascular corrosion casts was correlated with light microscopy of stained 7 µm thick serial tissues sections. Results showed that supplying arteries descended from the leptomeningeal surface without remarkable branchings straight to the subventricular zone where they branched and capillarized. Capillaries showed few H‐ and/or Y‐shaped anastomoses during their centrifugal course toward the leptomeningeal surface where they drained into cerebral venules and veins. Apart from the accessory olfactory bulb and the vestibule‐cochlear nucleus where capillaries were densely packed, capillaries formed a wide‐meshed 3D network throughout the brain parenchyma and thus contrasted to urodelian brains where hairpin‐shaped capillaries descend from the leptomeningeal vessels into varying depths of the brain parenchyma. In about two‐third of specimens, a closed arterial circle of Willis was found at the base of the brain. If this circle in Xenopus might serve the same two functions as in men is briefly discussed. Choroid plexuses of third and fourth ventricle were found to have a high venous, but a low arterial inflow via one small choroidal artery only. Findings are compared with previous studies on the vascularization of the anuran brain and discrepancies in respect to presence or absence of particular arteries and/or veins in Ranids, Bufonids, and Pipids studied so far are discussed with particular emphasis on the techniques used in the various studies published so far.     

Microvascular anatomy of the esophagus in larval and adult Xenopus laevis Daudin: a scanning electron microscope study of microvascular corrosion casts and correlative light microscopy

Larval to adult microvascular anatomy of the esophagus was studied in the South African Clawed Toad, Xenopus laevis (Daudin) by scanning electron microscopy of vascular corrosion casts and correlative light microscopy of paraplast embedded stained tissue sections. Analyses of vascular corrosion casts of tadpole esophaguses at premetamorphosis revealed a wide‐meshed, but mature looking capillary bed which during following prometamorphosis increased in density and gained the adult‐like pattern during late metamorphic climax by sprouting and nonsprouting angiogenesis. In adult Xenopus, the esophageal mucosa possessed a dense subepithelial capillary bed fed by one or two esophageal arteries that originated from right and/or left thoracic aorta just distal to the origin of the subclavian arteries. In the adult undistended esophagus, esophageal arteries revealed an undulating course, a pattern that guarantees a continuous blood supply when the esophagus is extremely wide expanded as it is the case when adult Xenopus swallows large prey.     

Peripheral vascularization of the dermal laminae of the equine hoof

The vascular architecture of the dermal laminae was studied by scanning electron microscopy of vascular corrosion casts. Ultrastructurally, the laminar vasculature consisted of arterioles, capillaries, venules and veins, arranged in a sheet-like network. Through the laminae, arterioles ran parallel to the solar surface and branched at two levels to form a continuous arteriolar arcade, parallel to the hoof wall. Capillaries originating from these arcades formed hairpin loops joining the marginal vein prior to forming an axially situated venous network. Additional capillaries were also given off by the arterioles, forming an abaxially arranged capillary plexus.     

Microvascularization of the spleen in larval and adult Xenopus laevis: Histomorphology and scanning electron microscopy of vascular corrosion casts

Microvascular anatomy and histomorphology of larval and adult spleens of the Clawed Toad, Xenopus laevis were studied by light microscopy of paraplast embedded serial tissue sections and scanning electron microscopy (SEM) of vascular corrosion casts (VCCs). Histology showed i) that white and red pulp are present at the onset of metamorphic climax (stage 57) and ii) that splenic vessels penetrated deeply into the splenic parenchyma at the height of metamorphic climax (stage 64). Scanning electron microscopy of VCCs demonstrated gross arterial supply and venous drainage, splenic microvascular patterns as well as the structure of the interstitial (extravasal) spaces representing the “open circulation routes.” These spaces identified themselves as interconnected resin masses of two distinct forms, namely “broccoli‐shaped” forms and highly interconnected small resin structures. Arterial and venous trees were clearly identified, as were transitions from capillaries to interstitial spaces and from interstitial spaces to pulp venules. Venous sinuses were not diagnosed (nonsinusal spleen). The splenic circulation in Xenopus laevis is “open.” It is hypothesized that red blood cells circulate via splenic artery, central arteries, penicillar arteries, and red pulp capillaries primarily via “broccoli‐shaped” interstitial spaces, pulp venules and veins into subcapsular veins to splenic veins while lymphocytes circulate also via the interstitial spaces represented by the highly interconnected small resin structures in vascular corrosion casts. In physiological terms, the former most likely represent the fast route for blood circulation, while the latter represent the slow route. J. Morphol. 277:1559–1569, 2016. © 2016 Wiley Periodicals, Inc.     

Hoof-Like Unguals,Skin, and Foot Movements Deduced from Deltapodus Casts of the Galve Basin (Upper Jurassic-Lower Cretaceous,Teruel, Spain)

Galve (Teruel, Spain) is a town in the interior of a synclinal fold with Upper Jurassic marine limestones along its flanks, and, in its core, Upper Jurassic–Lower Cretaceous continental and shoreline sediments crop out. The core sediments cover an area about 8 km², and contain a concentration of sites with footprints, bones, and eggshells of dinosaurs. The footprints are both shafts and natural casts. Some casts are attributed to stegosaurs (Deltapodus). The Deltapodus casts are characterized by features that allow us to make direct observations on the skin formed by polygonal scales, and ellipsoidal “hooves,” as well as deductions on the movement of the limbs during walking. According to the opinion of some authors, dinosaur footprints are indicators of the motion of their limbs and sometimes of the whole body. So far, results have been deduced from theropod, ornithopod, and sauropod footprints. This article shows the results obtained from analis of the aforementioned Deltapodus casts, i.e., forelimb movement similar to that of the forelimbs of sauropods, and the rigid structure of the autopodial part of the hind limb.     

Microvascularization on collared peccary placenta: a microvascular cast study [corrected] in late pregnancy

The microvascularization of the collared peccary (Tayassu tajacu) placenta was studied by vascular casts and immunolocalization of α-smooth muscle actin and vimentin, to identify the three dimensional organization and vascular flow interrelation in the microvasculature between the maternal and fetal compartments of the placentae. The immunolocalization of vimentin in the vascular endothelium and in the smooth muscle cells of blood vessels showed indented capillaries along the uterine epithelium and the trophoblast at the sides of complementary maternal and fetal microfolds, or rugae. This confers the three-dimensional structure observed in vascular casts. On the maternal side, casts demonstrated uterine folds coated by with primary and secondary ridges, and by areolae dispersed between these ridges. The arteriole runs through the center/middle of ridges, branching at the top into a microvascular network wall in a basket-like fashion. At the base of these baskets venules were formed. On the fetal side, arterioles branched centrally in the fetal rugae into a capillary network in a bulbous form, complementary to the opposite maternal depressions forming the baskets. At the base of the bulbous protrusions, the fetal venules arise. The blood vessel orientation in the materno-fetal interface of the placentae of collared peccaries suggests a blood flow pattern of the type countercurrent to cross current. The same pattern has been reported in domestic swine demonstrating that, even after 38 million years, the Tayassuidae and Suidae families exhibit similar placental morphology, which is here characterized at the microvascular level.     

Dynamics of the active site loops in catalyzing aminoacylation reaction in seryl and histidyl tRNA synthetases

Aminoacylation reaction is the first step of protein biosynthesis. The catalytic reorganization at the active site of aminoacyl tRNA synthetases (aaRSs) is driven by the loop motions. There remain lacunae of understanding concerning the catalytic loop dynamics in aaRSs. We analyzed the functional loop dynamics in seryl tRNA synthetase from Methanopyrus kandleri (^mkSerRS) and histidyl tRNA synthetases from Thermus thermophilus (^ttHisRS), respectively, using molecular dynamics. Results confirm that the motif 2 loop and other active site loops are flexible spots within the catalytic domain. Catalytic residues of the loops form a network of interaction with the substrates to form a reactive state. The loops undergo transitions between closed state and open state and the relaxation of the constituent residues occurs in femtosecond to nanosecond time scale. Order parameters are higher for constituent catalytic residues which form a specific network of interaction with the substrates to form a reactive state compared to the Gly residues within the loop. The development of interaction is supported from mutation studies where the catalytic domain with mutated loop exhibits unfavorable binding energy with the substrates. During the open-close motion of the loops, the catalytic residues make relaxation by ultrafast librational motion as well as fast diffusive motion and subsequently relax rather slowly via slower diffusive motion. The Gly residues act as a hinge to facilitate the loop closing and opening by their faster relaxation behavior. The role of bound water is analyzed by comparing implicit solvent-based and explicit solvent-based simulations. Loops fail to form catalytically competent geometry in absence of water. The present result, for the first time reveals the nature of the active site loop dynamics in aaRS and their influence on catalysis.     

Genetic variation in the Italian crested newt, Triturus carnifex, and the origin of a non-native population north of the Alps

Genetic variation over 40 protein loci and 46 populations representing three taxa of crested newts revealed moderate genetic distances between Triturus carnifex carnifex, T. c. macedonicus and T. cristatus. Two populations from the Geneva Basin (presumed to be introduced) were genetically similar to T. c. carnifex and dissimilar to T. c. macedonicus and T. cristatus, showing that they belong to T. c. carnifex and not to native T. cristatus. A significant pattern of spatial genetic variation was found within T. c. carnifex along a north to south axis, from Croatia to Calabria. The Genevan populations showed highest genetic similarity with T. carnifex from Tuscany, suggesting that the propagule originated from that area. Effects of a population genetic bottleneck associated with the introduction could not be documented. The observed high allelic variation in Genevan T. c. carnifex could not be directly explained by introgression from T. cristatus. Comparisons across the range, including zones of hybridization within the T. cristatus superspecies, indicated that some alleles typical for the Genevan population may represent the so-called `hybrizymes'.     

3-Dimensional Resin Casting and Imaging of Mouse Portal Vein or Intrahepatic Bile Duct System

In organs, the correct architecture of vascular and ductal structures is indispensable for proper physiological function, and the formation and maintenance of these structures is a highly regulated process. The analysis of these complex, 3-dimensional structures has greatly depended on either 2-dimensional examination in section or on dye injection studies. These techniques, however, are not able to provide a complete and quantifiable representation of the ductal or vascular structures they are intended to elucidate. Alternatively, the nature of 3-dimensional plastic resin casts generates a permanent snapshot of the system and is a novel and widely useful technique for visualizing and quantifying 3-dimensional structures and networks.A crucial advantage of the resin casting system is the ability to determine the intact and connected, or communicating, structure of a blood vessel or duct. The structure of vascular and ductal networks are crucial for organ function, and this technique has the potential to aid study of vascular and ductal networks in several ways. Resin casting may be used to analyze normal morphology and functional architecture of a luminal structure, identify developmental morphogenetic changes, and uncover morphological differences in tissue architecture between normal and disease states. Previous work has utilized resin casting to study, for example, architectural and functional defects within the mouse intrahepatic bile duct system that were not reflected in 2-dimensional analysis of the structure^1,2, alterations in brain vasculature of a Alzheimer''s disease mouse model³, portal vein abnormalities in portal hypertensive and cirrhotic mice⁴, developmental steps in rat lymphatic maturation between immature and adult lungs⁵, immediate microvascular changes in the rat liver, pancreas, and kidney in response in to chemical injury⁶.Here we present a method of generating a 3-dimensional resin cast of a mouse vascular or ductal network, focusing specifically on the portal vein and intrahepatic bile duct. These casts can be visualized by clearing or macerating the tissue and can then be analyzed. This technique can be applied to virtually any vascular or ductal system and would be directly applicable to any study inquiring into the development, function, maintenance, or injury of a 3-dimensional ductal or vascular structure.