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We established a normal embryonic development table for the Anji salamander Hynobius amjiensis, a critically endangered tailed amphibian of the family Hynobiidae with a very limited distribution in East China, following the standards set by the early developmental table of vertebrates. Put together 32 embryonic stages for the Anji salamander was defined. The total embryonic period from oviposition to hatching is approximately 30 days at 9 °C. Stages 1-16 represent early development from cleavage to neurulation. Stages 17-32 represent organogenesis documenting later developmental events such as tail, gill, and limb formation, and hatching (Stage 32). We provided a detailed description of the external morphology and color changes of the head, trunk, limbs, tail, external gills, and balancers at various stages from egg-laying to hatching. We also described several cases of abnormal embryonic development. The establishment of the embryonic development table in H. amjiensis contributes to better understanding of the ontogeny in tailed amphibians, distinguishing closely related species, and identifying abnormal embryonic amphibians., (Copyright © 2024 Elsevier Inc. All rights reserved.)

We present a complete staging table of normal development for the lungless salamander, Hemidactylium scutatum (Caudata: Plethodontidae). Terrestrial egg clutches from naturally ovipositing females were collected and maintained at 15 °C in the laboratory. Observations, photographs, and time-lapse movies of embryos were taken throughout the 45-day embryonic period. The complete normal table of development for H. scutatum is divided into 28 stages and extends previous analyses of H. scutatum embryonic development (Bishop, 1920; Humphrey, 1928). Early embryonic stage classifications through neurulation reflect criteria described for Xenopus laevis, Ambystoma maculatum and other salamanders. Later embryonic stage assignments are based on unique features of H. scutatum embryos. Additionally, we provide morphological analysis of gastrulation and neurulation, as well as details on external aspects of eye, gill, limb, pigmentation, and tail development to support future research related to phylogeny, comparative embryology, and molecular mechanisms of development., (Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.)

Studies of meso-endoderm and neural induction and subsequent body plan formation have been analyzed using mainly amphibians as the experimental model. Xenopus is currently the predominant model, because it best enables molecular analysis of these induction processes. However, much of the embryological information on these inductions (e.g., those of the Spemann-Mangold organizer), and on the morphogenetic movements of inductively interacting tissues, derives from research on non-model amphibians, especially urodeles. Although the final body pattern is strongly conserved in vertebrates, and although many of the same developmental genes are expressed, it has become evident that there are individually diverse modes of morphogenesis and timing of developmental events. Whether or not this diversity represents essential differences in the early induction processes remains unclear. The aim of this review is to compare the gastrulation process, induction processes, and gene expressions between a urodele, mainly Cynops pyrrhogaster, and an anura, Xenopus laevis, thereby to clarify conserved and diversified aspects. Cynops gastrulation differs significantly from that of Xenopus in that specification of the regions of the Xenopus dorsal marginal zone (DMZ) are specified before the onset of gastrulation, as marked by blastopore formation, whereas the equivalent state of specification does not occur in Cynops until the middle of gastrulation. Detailed comparison of the germ layer structure and morphogenetic movements during the pre-gastrula and gastrula stages shows that the entire gastrulation process should be divided into two phases of notochord induction and neural induction. Cynops undergoes these processes sequentially after the onset of gastrulation, whereas Xenopus undergoes notochord induction during a series of pre-gastrulation movements, and its traditionally defined period of gastrulation only includes the neural induction phase. Comparing the structure, fate, function and state of commitment of each domain of the DMZ of Xenopus and Cynops has revealed that the true form of the Spemann-Mangold organizer is suprablastoporal gsc-expressing endoderm that has notochord-inducing activity. Gsc-expressing deep endoderm and/or superficial endoderm in Xenopus is involved in inducing notochord during pre-gastrulation morphogenesis, rather than both gsc- and bra-expressing tissues being induced at the same time., (Copyright © 2012 Elsevier Inc. All rights reserved.)

Among the urodele amphibians, only Cynops pyrrhogaster and Pleurodeles waltl, two species of the Salamandridae family, were used in space experiments. The advantages for using urodeles reside (i) in reproduction: a few months after natural breeding, females can lay eggs in absence of males after a hormonal treatment, because spermatozoa were preserved in the cloacal pelvic glands of matted females, (ii) in the rate of development which is slower in Cynops and Pleurodeles than in the anuran Xenopus, (iii) in their physiological properties: they can live in a closed water container or in a moisturized environment, and they can fast during several days. Moreover, urodeles have an important phylogenetic interest. Many biological phenomena differ from those of anurans, such as fertilization events, the germ cell origin and the migration toward the differentiating gonads, and their regeneration capabilities. The main goals of the space experiments were to answer the following questions. On the one hand, does fertilization occur normally in microgravity? Is subsequent embryonic development normal in microgravity? Is further development and reproduction normal after return to Earth? On the other hand, does microgravity affect the organs in adult animals? Does microgravity affect the regeneration of organs? Fertilization in space is clearly demonstrated. However, subsequent embryonic development appears to be altered in microgravity. In Pleurodeles, abnormalities such as cortical cytoplasmic movements, decrease of cell adhesion, and loss of cells were observed. Although, early development was not strictly normal as a consequence of embryological regulation phenomena, young hatching larvae had normal morphological phenotypes and swimming behavior. After landing, no differences were observed between born-in-space animals to standard ones during the embryonic development to adulthood. The analyses of their offspring demonstrated that the percentages of fertilization and development were in accordance with the control animals. No genetic abnormalities were detected during the analysis of the offspring. The development of their progenies were also without characteristic differences compared to control Pleurodeles. Microgravity seems to have effects on the morphological and histological structures of organs of flight adults. However, as was the case in several experiments the number of analyzed adults was low, and it is too early to conclude on specific effects of microgravity. Moreover, in certain flights the temperature was not regulated, and an increase in temperature occurred. Conditions of these space flights had certainly influenced the samples, and consequently the interpretations of results. Space flights have clear effects on organs in regeneration. But more specifically, they have long term effects that last several weeks after the return of the animals to Earth. A similar result was also obtained for otoconia several months after landing. So far, however, no clear hypothesis could be proposed to interpret these observations.

Urodeles begin gastrulation with much of their presumptive mesoderm in the superficial cell layer, all of which must move into the deep layers during development. We studied the morphogenesis of superficial mesoderm in the urodeles Ambystoma maculatum, Ambystoma mexicanum, and Taricha granulosa. In all three species, somitic, lateral, and ventral mesoderm move into the deep layer during gastrulation, ingressing through a "bilateral primitive streak" just inside the blastopore. The mesodermal epithelium appears to slide under the endodermal epithelium by a mechanism we term "subduction." Subduction removes the large expanse of superficial presumptive somitic and lateral-ventral mesoderm that initially separates the sub-blastoporal endoderm from the notochord, leaving the endoderm bounding the still epithelial notochord along the gastrocoel roof. Subduction may be a common feature of urodele gastrulation, differing in this regard from anurans. Subducting cells constrict their apices and become bottle-shaped as they approach the junction of the mesodermal and endodermal epithelia. Subducting bottle cells endocytose apical membrane and withdraw the tight junctional component cingulin from the contracting circumferential tight junctions. Either in conjunction with or immediately after subducting, the mesodermal cells undergo an epithelial-to-mesenchymal transition. The mechanism by which epithelial cells release their apical junctions to become mesenchymal, without disrupting the integrity of the epithelium, remains mysterious, but this system should prove useful in understanding this process in a developmental context.

The dorsal marginal zone (DMZ) of an amphibian early gastrula is thought to consist of at least two distinct domains: the future head and trunk-tail organizers. We studied the mechanism by which the organizing activities of the lower half of the DMZ (LDMZ) of the urodelean (Cynops pyrrhogaster) embryo are changed. The uninvoluted LDMZ induces the notochord and then organizes the trunk-tail structures, whereas after cultivation in vitro or suramin treatment, the same LDMZ loses the notochord-inducing ability and organizes the head structures. A cell-lineage experiment indicated that the change in the organizing activity of the LDMZ was reflected in the transformation of the inductive ability: from notochord-inducing to neural-inducing activity. Using RT-PCR, we showed that the LDMZ expressed gsc, lim-1, chordin, and noggin, but not the mesoderm marker bra. In the sandwich assay, the LDMZ induced bra expression in the animal cap ectoderm, but the inductive activity was inhibited by cultivation or suramin treatment. The present study indicates that the change in the organizing activity of the LDMZ from trunk-tail to head is coupled with the loss of notochord-inducing activity. Based on these results, we suggest that this change is essential for the specification of the head and trunk-tail organizers during gastrulation., (Copyright 2002 Elsevier Science (USA).)

Previously, we found that the conversion of hemoglobins (Hbs) from the larval to the adult type occurred within a single erythroid cell population in a salamander, Hynobius retardatus ("Hb switching" model), whereas the transition involves replacement of red-blood-cell (RBC) populations ("RBC replacement" model) in many amphibians (M. Yamaguchi, H. Takahashi, and M. Wakahara, 2000, Dev. Gene Evol. 210, 180-189). To further characterize the Hb transition, developmental changes in the erythropoietic sites have been intensively analyzed using larval- and adult-specific globin antibodies and globin and GATA-3 RNA probes. Cells of the ventral blood island (VBI) and the dorsolateral plate (DLP) in embryos differentiate in situ to erythroid cells that contain larval globin mRNA, suggesting that both the VBI and the DLP contribute to "primitive" erythropoiesis. In contrast, the expression pattern of the GATA-3 gene suggests that cells of the DLP may contribute to "definitive" hematopoiesis. In order to determine whether it is possible to define a definitive erythropoiesis in H. retardatus or not, further experiments were done: (1) when metamorphosing larvae were treated with phenylhydrazine to induce anemia and then bled at the postmetamorphic stage after recovery from the anemia, a precocious Hb transition was observed in these animals; (2) an RBC population expressing only adult Hb was confirmed by subtracting the number of RBCs expressing larval Hb from the total number of RBCs during metamorphosis. All these results support the existence of a definitive erythroid cell population that contributes only adult RBCs in this species., (Copyright 2001 Academic Press.)

The response of various anuran and urodele eggs to a brief rotation prior to first cleavage were observed. In all instances, the dorsal lip formed on the side of the egg which opposed gravity during the rotation. The ability of egg rotation to prevent the series of developmental abnormalities associated with UV irradiation of the uncleaved egg was also examined. Anuran and urodele eggs responded to those manipulations in a similar fashion. These results should facilitate future studies on the mechanisms involved in the establishment of the dorsal/ventral polarity of the amphibian egg.

In order to study the transmission mechanism of neuralising signals during primary embryonic induction, the interacting components (competent newt gastrula ectoderm and dorsal lip tissues) were separated by filter membranes of varying pore size. Nuclepore filters with nominal pore size from 0.1 to 8 mum were employed and the neuralising effect was shown to traverse all of these membranes. Electron microscopic examination did not reveal any cytoplasmic processes in the pores and the authors conclude that the morphogenetic signals are carried by transmissable compounds rather than through direct cytoplasmic contacts.

Using transmission and scanning electron microscopy we were able to show that during primary embryonic induction in amphibians (Triturus alpestris) the interspace between the inducing chorda-mesoderm and the reacting ectoderm (presumptive medullary plate) of mid-gastrula stages is traversed by cell projections starting from cells of both tissue layers. In addition intimate membrane contacts between the main bodies of the ectodermal and chorda-mesodermal cells could be observed. It could be ruled out that cytoplasmic bridges (anastomosis) exist between cells of inducing chorda-mesoderm and reacting ectoderm, which would allow a free transfer of inducing substances without passing through membranes, as Eakin and Lehmann [1] have postulated. The possible role of cell to cell contact for neural induction is emphasized.