Talk:Axolotl Development

From Embryology
About Discussion Pages  
Mark Hill.jpg
On this website the Discussion Tab or "talk pages" for a topic has been used for several purposes:
  1. References - recent and historic that relates to the topic
  2. Additional topic information - currently prepared in draft format
  3. Links - to related webpages
  4. Topic page - an edit history as used on other Wiki sites
  5. Lecture/Practical - student feedback
  6. Student Projects - online project discussions.
Links: Pubmed Most Recent | Reference Tutorial | Journal Searches

Glossary Links

Glossary: A | B | C | D | E | F | G | H | I | J | K | L | M | N | O | P | Q | R | S | T | U | V | W | X | Y | Z | Numbers | Symbols | Term Link

Cite this page: Hill, M.A. (2021, April 16) Embryology Axolotl Development. Retrieved from


Vincent E, Villiard E, Sader F, Dhakal S, Kwok BH & Roy S. (2020). BMP signaling is essential for sustaining proximo-distal progression in regenerating axolotl limbs. Development , 147, . PMID: 32665245 DOI.

BMP signaling is essential for sustaining proximo-distal progression in regenerating axolotl limbs

Amputation of a salamander limb triggers a regeneration process that is perfect. A limited number of genes have been studied in this context and even fewer have been analyzed functionally. In this work, we use the BMP signaling inhibitor LDN193189 on Ambystoma mexicanum to explore the role of BMPs in regeneration. We find that BMP signaling is required for proper expression of various patterning genes and that its inhibition causes major defects in the regenerated limbs. Fgf8 is downregulated when BMP signaling is blocked, but ectopic injection of either human or axolotl protein did not rescue the defects. By administering LDN193189 treatments at different time points during regeneration, we show clearly that limb regeneration progresses in a proximal to distal fashion. This demonstrates that BMPs play a major role in patterning of regenerated limbs and that regeneration is a progressive process like development.


Rediscovering the Axolotl as a Model for Thyroid Hormone Dependent Development

Front Endocrinol (Lausanne). 2019 Apr 12;10:237. doi: 10.3389/fendo.2019.00237. eCollection 2019.

Crowner A1, Khatri S1, Blichmann D1, Voss SR1.

The Mexican axolotl (Ambystoma mexicanum) is an important model organism in biomedical research. Much current attention is focused on the axolotl's amazing ability to regenerate tissues and whole organs after injury. However, not forgotten is the axolotl's equally amazing ability to thwart aspects of tissue maturation and retain juvenile morphology into the adult phase of life. Unlike close tiger salamander relatives that undergo a thyroid hormone regulated metamorphosis, the axolotl does not typically undergo a metamorphosis. Instead, the axolotl exhibits a paedomorphic mode of development that enables a completely aquatic life cycle. The evolution of paedomorphosis allowed axolotls to exploit relatively permanent habitats in Mexico, and preadapted axolotls for domestication and laboratory study. In this perspective, we first introduce the axolotl and the various meanings of paedomorphosis, and then stress the need to move beyond endocrinology-guided approaches to understand the axolotl's hypothyroid state. With the recent completion of the axolotl genome assembly and established methods to manipulate gene functions, the axolotl is poised to provide new insights about paedomorphosis and the role of thyroid hormone in development and evolution. KEYWORDS: ambystoma; axolotl; metamorphosis; paedomorphosis; thyroid hormone PMID: 31031711 PMCID: PMC6473073 DOI: 10.3389/fendo.2019.00237


Biosystems. 2018 Nov;173:214-220. doi: 10.1016/j.biosystems.2018.10.006. Epub 2018 Oct 25. Acquisition and reconstruction of 4D surfaces of axolotl embryos with the flipping stage robotic microscope. Crawford-Young SJ1, Dittapongpitch S2, Gordon R3, Harrington KIS4. Author information Abstract We have designed and constructed a Flipping Stage for a light microscope that can view the whole exterior surface of a 2 mm diameter developing axolotl salamander embryo. It works by rapidly inverting the bottom-heavy embryo, imaging it as it rights itself. The images are then montaged to reconstruct the whole 3D surface versus time, for a full 4D record of the surface. Imaging early stage axolotl development will help discover how cell differentiation and movement takes place in the early embryo. For example, the switch from ectodermal to neural plate cells takes place on the top, animal surface portion the egg/embryo and can be observed using the flipping stage microscope. Detailed pictures of the whole surface need to be obtained so that cell tracking and event histories, such as cell divisions and participation in differentiation waves, of individual cells can be recorded. Imaging the whole exterior of the eggs/embryos will allow for the analysis of cell behavior and the forces the cells experience in their natural setting in the intact or manipulated embryo. This will give insights into embryogenesis, development, developmental disruptions, birth defects, cell differentiation and tissue engineering.

KEYWORDS: Amphibian embryogenesis; Cell tracking; Differentiation waves; Embryo imaging; Image montaging; Robotic microscopy PMID: 30554603 DOI: 10.1016/j.biosystems.2018.10.006

The axolotl genome and the evolution of key tissue formation regulators

Nature. 2018 Feb 1;554(7690):50-55. doi: 10.1038/nature25458. Epub 2018 Jan 24.

Nowoshilow S1,2,3, Schloissnig S4, Fei JF5, Dahl A3,6, Pang AWC7, Pippel M4, Winkler S1, Hastie AR7, Young G8, Roscito JG1,9,10, Falcon F11, Knapp D3, Powell S4, Cruz A11, Cao H7, Habermann B12, Hiller M1,9,10, Tanaka EM1,2,3, Myers EW1,10.

Abstract Salamanders serve as important tetrapod models for developmental, regeneration and evolutionary studies. An extensive molecular toolkit makes the Mexican axolotl (Ambystoma mexicanum) a key representative salamander for molecular investigations. Here we report the sequencing and assembly of the 32-gigabase-pair axolotl genome using an approach that combined long-read sequencing, optical mapping and development of a new genome assembler (MARVEL). We observed a size expansion of introns and intergenic regions, largely attributable to multiplication of long terminal repeat retroelements. We provide evidence that intron size in developmental genes is under constraint and that species-restricted genes may contribute to limb regeneration. The axolotl genome assembly does not contain the essential developmental gene Pax3. However, mutation of the axolotl Pax3 paralogue Pax7 resulted in an axolotl phenotype that was similar to those seen in Pax3-/- and Pax7-/- mutant mice. The axolotl genome provides a rich biological resource for developmental and evolutionary studies. PMID: 29364872 DOI: 10.1038/nature25458

Morphological and transcriptomic analyses reveal three discrete primary stages of postembryonic development in the common fire salamander, Salamandra salamandra

J Exp Zool B Mol Dev Evol. 2018 Mar 5. doi: 10.1002/jez.b.22792. [Epub ahead of print]

Sanchez E1, Küpfer E1, Goedbloed DJ1, Nolte AW2,3, Lüddecke T1, Schulz S4, Vences M1, Steinfartz S1.


The postembryonic development of amphibians has been characterized as divided into three predominant periods, hereafter named primary developmental stages: premetamorphosis (PreM), prometamorphosis (ProM), metamorphic climax (Meta), and completion of metamorphosis (PostM), largely based on examination of anuran development. Here, we categorized the postembryonic development of larvae of a poisonous fire salamander (Salamandra salamandra) by integrating morphology and gene expression (transcriptomic) data. Morphological analysis revealed three distinct clusters suggestive of PreM, ProM, and Meta, which were confirmed in parallel by microarray-derived gene expression analysis. In total, 3,510 probes targeted transcripts differentially expressed between the clusters we identified. Genes upregulated in PreM related to organogenesis, and those upregulated in Meta underlie structural proteins and related to development of anatomical structures and pigmentation. Biosynthesis pathways of pigments (pteridines and melanin) were upregulated during late ProM and Meta. Gas chromatographic analysis of alkaloids indicated the onset of steroidal alkaloid biosynthesis at ProM. When comparing gene expression in the fire salamander to that in other amphibians-three anurans, Xenopus laevis, X. tropicalis, and Michrohyla fissipes, and one caudate, Ambystoma mexicanum- we identified genes with conserved expression patterns involved in basic metamorphic processes such as skin restructuring and tail fin resorption. Our results support that primary stages of postembryonic development in caudates are homologous to those of anurans, and offer a baseline for the study of the evolution of developmental modes. KEYWORDS: amphibian metamorphosis evolution; gene expression; pigments (melanin and pteridines) biosynthesis; salamander steroidal alkaloids; skin restructuring; tail fin resorption PMID: 29504232 DOI: 10.1002/jez.b.22792


Dual embryonic origin and patterning of the pharyngeal skeleton in the axolotl (Ambystoma mexicanum)

Evol Dev. 2015 May-Jun;17(3):175-84. doi: 10.1111/ede.12124.

Sefton EM1, Piekarski N1, Hanken J1.


The impressive morphological diversification of vertebrates was achieved in part by innovation and modification of the pharyngeal skeleton. Extensive fate mapping in amniote models has revealed a primarily cranial neural crest derivation of the pharyngeal skeleton. Although comparable fate maps of amphibians produced over several decades have failed to document a neural crest derivation of ventromedial elements in these vertebrates, a recent report provides evidence of a mesodermal origin of one of these elements, basibranchial 2, in the axolotl. We used a transgenic labeling protocol and grafts of labeled cells between GFP+ and white embryos to derive a fate map that describes contributions of both cranial neural crest and mesoderm to the axolotl pharyngeal skeleton, and we conducted additional experiments that probe the mechanisms that underlie mesodermal patterning. Our fate map confirms a dual embryonic origin of the pharyngeal skeleton in urodeles, including derivation of basibranchial 2 from mesoderm closely associated with the second heart field. Additionally, heterotopic transplantation experiments reveal lineage restriction of mesodermal cells that contribute to pharyngeal cartilage. The mesoderm-derived component of the pharyngeal skeleton appears to be particularly sensitive to retinoic acid (RA): administration of exogenous RA leads to loss of the second basibranchial, but not the first. Neural crest was undoubtedly critical in the evolution of the vertebrate pharyngeal skeleton, but mesoderm may have played a central role in forming ventromedial elements, in particular. When and how many times during vertebrate phylogeny a mesodermal contribution to the pharyngeal skeleton evolved remain to be resolved. © 2015 Wiley Periodicals, Inc.

PMID 25963195


Stochastic specification of primordial germ cells from mesoderm precursors in axolotl embryos

Development. 2014 Jun;141(12):2429-40. doi: 10.1242/dev.105346.

Chatfield J1, O'Reilly MA1, Bachvarova RF2, Ferjentsik Z1, Redwood C1, Walmsley M3, Patient R3, Loose M1, Johnson AD4.


A common feature of development in most vertebrate models is the early segregation of the germ line from the soma. For example, in Xenopus and zebrafish embryos primordial germ cells (PGCs) are specified by germ plasm that is inherited from the egg; in mice, Blimp1 expression in the epiblast mediates the commitment of cells to the germ line. How these disparate mechanisms of PGC specification evolved is unknown. Here, in order to identify the ancestral mechanism of PGC specification in vertebrates, we studied PGC specification in embryos from the axolotl (Mexican salamander), a model for the tetrapod ancestor. In the axolotl, PGCs develop within mesoderm, and classic studies have reported their induction from primitive ectoderm (animal cap). We used an axolotl animal cap system to demonstrate that signalling through FGF and BMP4 induces PGCs. The role of FGF was then confirmed in vivo. We also showed PGC induction by Brachyury, in the presence of BMP4. These conditions induced pluripotent mesodermal precursors that give rise to a variety of somatic cell types, in addition to PGCs. Irreversible restriction of the germ line did not occur until the mid-tailbud stage, days after the somatic germ layers are established. Before this, germline potential was maintained by MAP kinase signalling. We propose that this stochastic mechanism of PGC specification, from mesodermal precursors, is conserved in vertebrates. © 2014. Published by The Company of Biologists Ltd. KEYWORDS: Axolotl; Evolution; Germ plasm; Mesoderm; PGC; Pluripotency; Primordial germ cell

PMID 24917499


Gain-of-function assays in the axolotl (Ambystoma mexicanum) to identify signaling pathways that induce and regulate limb regeneration

Methods Mol Biol. 2013;1037:401-17. doi: 10.1007/978-1-62703-505-7_23.

Lee J, Aguilar C, Gardiner D. Source Department of Developmental and Cell Biology, University of California Irvine, Irvine, CA, USA.


The adult salamander has been studied as a model for regeneration of complex tissues for many decades. Only recently with the development of gain-of-function assays for regeneration, has it been possible to screen for and assay the function of the multitude of signaling factors that have been identified in studies of embryonic development and tumorigenesis. Given the conservation of function of these regulatory pathways controlling growth and pattern formation, it is now possible to use the functional assays in the salamander to test the ability of endogenous as well as small-molecule signaling factors to induce a regenerative response.

PMID 24029949

Cranial muscle development in the model organism ambystoma mexicanum: implications for tetrapod and vertebrate comparative and evolutionary morphology and notes on ontogeny and phylogeny

Anat Rec (Hoboken). 2013 Jul;296(7):1031-48. doi: 10.1002/ar.22713. Epub 2013 May 6.

Ziermann JM, Diogo R. Source Department of Anatomy, Howard University College of Medicine, Washington DC 20059, USA.


There is still confusion about the homology of several cranial muscles in salamanders with those of other vertebrates. This is true, in part, because of the fact that many muscles present in early ontogeny of amphibians disappear during development and specifically during metamorphosis. Resolving this confusion is important for the understanding of the comparative and evolutionary morphology of vertebrates and tetrapods because amphibians are the phylogenetically most plesiomorphic tetrapods, concerning for example their myology, and include two often used model organisms, Xenopus laevis (anuran) and Ambystoma mexicanum (urodele). Here we provide the first detailed report of the cranial muscle development in axolotl from early ontogenetic stages to the adult stage. We describe different and complementary types of general muscle morphogenetic gradients in the head: from anterior to posterior, from lateral to medial, and from origin to insertion. Furthermore, even during the development of neotenic salamanders such as axolotls, various larval muscles become indistinct, contradicting the commonly accepted view that during ontogeny the tendency is mostly toward the differentiation of muscles. We provide an updated comparison between these muscles and the muscles of other vertebrates, a discussion of the homologies and evolution, and show that the order in which the muscles appear during axolotl ontogeny is in general similar to their appearance in phylogeny (e.g. differentiation of adductor mandibulae muscles from one anlage to four muscles), with only a few remarkable exceptions, as for example the dilatator laryngis that appears evolutionary later but in the development before the intermandibularis. Copyright © 2013 Wiley Periodicals, Inc.

PMID 23650269


Regeneration of Limb Joints in the Axolotl (Ambystoma mexicanum)

PLoS One. 2012;7(11):e50615. doi: 10.1371/journal.pone.0050615. Epub 2012 Nov 21.

Lee J, Gardiner DM. Source Department of Developmental and Cell Biology, University of California Irvine, Irvine, California, United States of America ; The Developmental Biology Center, University of California Irvine, Irvine, California, United States of America.


In spite of numerous investigations of regenerating salamander limbs, little attention has been paid to the details of how joints are reformed. An understanding of the process and mechanisms of joint regeneration in this model system for tetrapod limb regeneration would provide insights into developing novel therapies for inducing joint regeneration in humans. To this end, we have used the axolotl (Mexican Salamander) model of limb regeneration to describe the morphology and the expression patterns of marker genes during joint regeneration in response to limb amputation. These data are consistent with the hypothesis that the mechanisms of joint formation whether it be development or regeneration are conserved. We also have determined that defects in the epiphyseal region of both forelimbs and hind limbs in the axolotl are regenerated only when the defect is small. As is the case with defects in the diaphysis, there is a critical size above which the endogenous regenerative response is not sufficient to regenerate the joint. This non-regenerative response in an animal that has the ability to regenerate perfectly provides the opportunity to screen for the signaling pathways to induce regeneration of articular cartilage and joints.

PMID 23185640


Dual epithelial origin of vertebrate oral teeth

Nature. 2008 Oct 9;455(7214):795-8. doi: 10.1038/nature07304. Epub 2008 Sep 14.

Soukup V, Epperlein HH, Horácek I, Cerny R. Source Department of Zoology, Charles University in Prague, Vinicna 7, 128 44 Prague, Czech Republic.


The oral cavity of vertebrates is generally thought to arise as an ectodermal invagination. Consistent with this, oral teeth are proposed to arise exclusively from ectoderm, contributing to tooth enamel epithelium, and from neural crest derived mesenchyme, contributing to dentin and pulp. Yet in many vertebrate groups, teeth are not restricted only to the oral cavity, but extend posteriorly as pharyngeal teeth that could be derived either directly from the endodermal epithelium, or from the ectodermal epithelium that reached this location through the mouth or through the pharyngeal slits. However, when the oropharyngeal membrane, which forms a sharp ecto/endodermal border, is broken, the fate of these cells is poorly known. Here, using transgenic axolotls with a combination of fate-mapping approaches, we present reliable evidence of oral teeth derived from both the ectoderm and endoderm and, moreover, demonstrate teeth with a mixed ecto/endodermal origin. Despite the enamel epithelia having a different embryonic source, oral teeth in the axolotl display striking developmental uniformities and are otherwise identical. This suggests a dominant role for the neural crest mesenchyme over epithelia in tooth initiation and, from an evolutionary point of view, that an essential factor in teeth evolution was the odontogenic capacity of neural crest cells, regardless of possible 'outside-in' or 'inside-out' influx of the epithelium. Comment in Developmental biology: Teeth in double trouble. [Nature. 2008]

PMID 18794902


Transforming growth factor: beta signaling is essential for limb regeneration in axolotls

PLoS One. 2007 Nov 28;2(11):e1227.

Lévesque M, Gatien S, Finnson K, Desmeules S, Villiard E, Pilote M, Philip A, Roy S.

Source Department of Biochemistry, Université de Montréal, Montréal, Québec, Canada.


Axolotls (urodele amphibians) have the unique ability, among vertebrates, to perfectly regenerate many parts of their body including limbs, tail, jaw and spinal cord following injury or amputation. The axolotl limb is the most widely used structure as an experimental model to study tissue regeneration. The process is well characterized, requiring multiple cellular and molecular mechanisms. The preparation phase represents the first part of the regeneration process which includes wound healing, cellular migration, dedifferentiation and proliferation. The redevelopment phase represents the second part when dedifferentiated cells stop proliferating and redifferentiate to give rise to all missing structures. In the axolotl, when a limb is amputated, the missing or wounded part is regenerated perfectly without scar formation between the stump and the regenerated structure. Multiple authors have recently highlighted the similarities between the early phases of mammalian wound healing and urodele limb regeneration. In mammals, one very important family of growth factors implicated in the control of almost all aspects of wound healing is the transforming growth factor-beta family (TGF-beta). In the present study, the full length sequence of the axolotl TGF-beta1 cDNA was isolated. The spatio-temporal expression pattern of TGF-beta1 in regenerating limbs shows that this gene is up-regulated during the preparation phase of regeneration. Our results also demonstrate the presence of multiple components of the TGF-beta signaling machinery in axolotl cells. By using a specific pharmacological inhibitor of TGF-beta type I receptor, SB-431542, we show that TGF-beta signaling is required for axolotl limb regeneration. Treatment of regenerating limbs with SB-431542 reveals that cellular proliferation during limb regeneration as well as the expression of genes directly dependent on TGF-beta signaling are down-regulated. These data directly implicate TGF-beta signaling in the initiation and control of the regeneration process in axolotls.

PMID 18043735

The axolotl limb: a model for bone development, regeneration and fracture healing

Bone. 2007 Jan;40(1):45-56. Epub 2006 Aug 21.

Hutchison C, Pilote M, Roy S. Source Department of Biochemistry, Université de Montréal, Québec, Canada.


Among vertebrates, urodele amphibians (e.g., axolotls) have the unique ability to perfectly regenerate complex body parts after amputation. The limb has been the most widely studied due to the presence of three defined axes and its ease of manipulation. Hence, the limb has been chosen as a model to study the process of skeletogenesis during axolotl development, regeneration and to analyze this animal's ability to heal bone fractures. Extensive studies have allowed researchers to gain some knowledge of the mechanisms controlling growth and pattern formation in regenerating and developing limbs, offering an insight into how vertebrates are able to regenerate tissues. In this study, we report the cloning and characterization of two axolotl genes; Cbfa-1, a transcription factor that controls the remodeling of cartilage into bone and PTHrP, known for its involvement in the differentiation and maturation of chondrocytes. Whole-mount in situ hybridization and immunohistochemistry results show that Cbfa-1, PTHrP and type II collagen are expressed during limb development and regeneration. These genes are expressed during specific stages of limb development and regeneration which are consistent with the appearance of skeletal elements. The expression pattern for Cbfa-1 in late limb development was similar to the expression pattern found in the late stages of limb regeneration (i.e. re-development phase) and it did not overlap with the expression of type II collagen. It has been reported that the molecular mechanisms involved in the re-development phase of limb regeneration are a recapitulation of those used in developing limbs; therefore the detection of Cbfa-1 expression during regeneration supports this assertion. Conversely, PTHrP expression pattern was different during limb development and regeneration, by its intensity and by the localization of the signal. Finally, despite its unsurpassed abilities to regenerate, we tested whether the axolotl was able to regenerate non-union bone fractures. We show that while the axolotl is able to heal a non-stabilized union fracture, like other vertebrates, it is incapable of healing a bone gap of critical dimension. These results suggest that the axolotl does not use the regeneration process to repair bone fractures.

PMID 16920050