Vitamin A modification of the positional information of blastema cells during limb regeneration in the axolotl Ambystoma mexicanum

1988 ◽  
Vol 66 (9) ◽  
pp. 2065-2070 ◽  
Author(s):  
Steven R. Scadding
Keyword(s):  
Vitamin A ◽  
Limb Regeneration ◽  
Normal Structure ◽  
Positional Information ◽  
Ambystoma Mexicanum ◽  
Limb Amputation ◽  
Temporary Change ◽  
Cellular Changes ◽  
Regeneration Blastema

Vitamin A is known to cause proximodistal duplication of parts of the limb during limb regeneration in amphibians. The objective of this study was to investigate the nature and location of the cellular changes induced by vitamin A when it causes this duplication in the axolotl Ambystoma mexicanum. When axolotls were treated with retinol palmitate by immersion for 14 days before limb amputation, proximodistal duplications were still observed in subsequent regenerates of limbs amputated after vitamin A treatment was discontinued. This observation suggests that some characteristic of the cells is changed by vitamin A and that exogenous vitamin A need not be present while the limb is regenerating. When a limb that was induced to undergo proximodistal duplication by vitamin A was reamputated 49 days later through the original mid radius–ulna amputation plane, it regenerated a limb of normal structure. A regeneration blastema transplanted from a vitamin A treated axolotl to an untreated axolotl regenerated on the host limb stump, producing a limb with proximodistal duplication; this indicates that the blastema cells underwent some change by the early to mid cone stage, which was expressed later when the blastema redifferentiated into a new limb. Conversely, when an untreated blastema was transplanted onto a vitamin A treated axolotl from which the forelimb blastema had been removed, proximodistal duplications developed. This result is interpreted to mean that the stump cells, although morphologically of the radius–ulna level, were proximalized by the prior vitamin A treatment and still displayed proximal positional values, leading to intercalation of missing proximodistal structures. These results are consistent with the hypothesis that vitamin A brings about a temporary change in the positional information of the limb stump and blastema cells, and that when the vitamin A treatment is discontinued, there is a gradual return to normal positional values over a period of several weeks.

Symmetrical vascularization patterns in normal and retinoic acid treated limb regenerates of the axolotl, Ambystoma mexicanum

1998 ◽  
Vol 76 (9) ◽  
pp. 1795-1796 ◽  
Author(s):  
Steven R Scadding ◽  
Andrew Burns
Keyword(s):  
Retinoic Acid ◽  
Pattern Formation ◽  
Vascular System ◽  
Limb Regeneration ◽  
Ambystoma Mexicanum ◽  
Limb Pattern ◽  
Regeneration Blastema

The purpose of this investigation was to determine whether there were any asymmetries in the vascularization of the limb-regeneration blastema in the axolotl, Ambystoma mexicanum, that might be related to pattern formation, and to determine if retinoic acid could modify the vascular patterns of the blastema. We used acrylic casts of the vascular system of the limbs to assess the pattern of vascularization. We observed a very regular symmetrical arrangement of capillaries in the limb-regeneration blastema that did not appear to be modified by doses of retinoic acid sufficient to modify the limb pattern.

Skeletal patterns in the autopodium of native and regenerated limbs of the larval axolotl, Ambystoma mexicanum

1991 ◽  
Vol 69 (1) ◽  
pp. 1-6 ◽  
Author(s):  
Steven R. Scadding
Keyword(s):  
Vitamin A ◽  
Significant Variation ◽  
Limb Regeneration ◽  
Ambystoma Mexicanum ◽  
Skeletal Patterning ◽  
Skeletal Pattern ◽  
Limb Pattern ◽  
Salamander Species

The purpose of this investigation was to study the autopodial skeletal patterns that are observed in native (never regenerated) and regenerated limbs of the larval axolotl, Ambystoma mexicanum. The axolotl is used widely in limb regeneration studies, and in the regenerating axolotl limb mesopodial patterns can be modified by such factors as vitamin A administration. It is also known that other salamander species show significant variation in autopodial skeletal patterning. Hence, it seemed important to determine the type and frequency of autopodial variants in both native limbs and those that have regenerated after amputation at either the stylopodial and zeugopodial levels. The results showed that native limbs exhibited a complete skeletal pattern in the majority of cases, but that variants involving loss of a phalange or reduction in the number of carpals or tarsals occurred frequently. Regenerated limb patterns were more variable than those seen in native limbs, and limbs regenerating from zeugopodial level amputations were more variable than those regenerating from stylopodial level amputation. The significance of these observations for the development and regeneration of limb pattern is discussed.

Activities of acetylcholinesterase, choline acetyltransferase, and catecholamine production in the spinal cord of the axolotl Ambystoma mexicanum during forelimb regeneration

1994 ◽  
Vol 72 (5-6) ◽  
pp. 188-194 ◽  
Author(s):  
Patrick Scaps ◽  
François Bernet ◽  
Jean Gautron ◽  
Bénoni Boilly
Keyword(s):  
Spinal Cord ◽  
Molecular Weight ◽  
Choline Acetyltransferase ◽  
Control Level ◽  
Limb Regeneration ◽  
Nerve Fibers ◽  
Ambystoma Mexicanum ◽  
Limb Amputation ◽  
Metabolic Activities ◽  
Molecular Weight Form

Amputation of an axolotl limb causes severance of the brachial nerves, followed by their regeneration into a blastema. It is known that these nerves provide a neurotophic factor to blastemal cells. To approach the problem of the response of spinal cord nerve centers to forelimb amputation, we have studied biosynthetic activities in the nerve centers involved in axonal injury during limb regeneration. We report that the acetylcholinesterase (AChE) activity in the spinal cord is elevated 2 days (+ 69%) and 7 days (+ 28%) after limb amputation compared with levels in unamputated control animals, but is not significantly elevated at 3 h or 15 days. The percentages of slow (3.6 S and 6.0 S) and fast (18 S) sedimenting forms of AChE progressively decrease 2 and 7 days after amputation, while those of intermediate sedimenting forms (10.5 S and 14.0 S) increase. Fifteen days after amputation, lower molecular weight forms return to the control level, but the heavy molecular weight form of AChE is absent as at 7 days; consequently intermediate molecular weight forms are in a greater proportion than the other two forms. Choline acetyltransferase activity was measured only 2 days after amputation (when AChE was at its highest level). It increases by about 34% with regard to the controls. Adrenaline is higher than controls 2 days after amputation, while noradrenaline is not significantly modified. The metabolic changes observed in the spinal cord during limb regeneration probably are the result of a general reaction to the stress of amputation (transection of brachial nerves) and regeneration of nerve fibers, since similar metabolic activities were observed after a simple denervation of the two unamputated forelimbs.Key words: acetylcholinesterase, choline acetyltransferase, catecholamines, regeneration, axolotl.

Actinomycin D, cycloheximide, and tunicamycin inhibit vitamin A induced proximodistal duplication during limb regeneration in the axolotl Ambystoma mexicanum

1988 ◽  
Vol 66 (4) ◽  
pp. 879-884 ◽  
Author(s):  
Steven R. Scadding
Keyword(s):  
Protein Synthesis ◽  
Vitamin A ◽  
Actinomycin D ◽  
Limb Regeneration ◽  
Ambystoma Mexicanum ◽  
Glycoprotein Synthesis ◽  
Left Limb ◽  
Site Of Action ◽  
The Right

Vitamin A is known to cause proximodistal duplication of parts of the limb during limb regeneration in amphibians. The objective of this study was to investigate the cellular site of action of vitamin A when it causes this duplication in the axolotl Ambystoma mexicanum. Both forelimbs of larval axolotls were amputated through the radius–ulna, and treated with retinol palmitate by immersion for 10 days. Actinomycin D, cycloheximide, and tunicamycin were each applied to the right forelimb by implantation of a silastin block containing one of these antibiotics on days 2 and 6 postamputation. The left limb received a control silastin block at the same time. Actinomycin D and cycloheximide completely blocked the proximodistal duplication caused by retinol palmitate, and tunicamycin substantially inhibited the development of these duplications. These results suggest that retinol palmitate (i) induces duplication by switching on a gene (as it is antagonized by actinomycin D), (ii) requires protein synthesis (as it is antagonized by cycloheximide), and (iii) may effect glycoprotein synthesis (as it is antagonized by tunicamycin).

Gap junctions in the limb regeneration blastema of the axolotl, Ambystoma mexicanum, are not distributed uniformly and are regulated by retinoic acid

1999 ◽  
Vol 77 (6) ◽  
pp. 902-909
Author(s):  
Leigh-Anne D Miller ◽  
Melissa L Farquhar ◽  
John S Greenwood ◽  
Steven R Scadding
Keyword(s):  
Retinoic Acid ◽  
Gap Junctions ◽  
Limb Regeneration ◽  
Ambystoma Mexicanum ◽  
Limb Bud ◽  
Gap Junctional Communication ◽  
Junctional Communication ◽  
Gap Junctional ◽  
Regeneration Blastema

Gap junctions are thought to play a role in pattern formation during limb development and regeneration by controlling the movement of small regulatory molecules between cells. An anteroposterior gradient of gap junctional communication that is higher posteriorly has been reported in the developing chick limb bud. In both the developing chick limb bud and the amphibian regenerating limb, an anteroposterior retinoic acid gradient is present, and this is also higher posteriorly. On the basis of these observations, we decided to examine the role of gap junctional communication in the regenerating amphibian limb. Gap junctions were observed in both the axolotl, Ambystoma mexicanum, limb regeneration blastema and cardiac tissue (as a positive control), using immunohistochemical labelling and laser scanning confocal microscopy. The scrape-loading/dye transfer technique for tracing the movement of a gap junction permeable dye, Lucifer yellow, showed that in blastemal epidermis there were nonuniform distributions of gap junctions in both the dorsoventral and anteroposterior axes of the blastema. Retinoic acid was found to increase gap junctional permeability in blastemal epidermis 48 h after injection and in blastemal mesenchyme 76 h after injection. The potential role of gap junctions during pattern formation in limb regeneration is discussed based on these results.

Histological effects of vitamin A on limb regeneration in the larval axolotl, Ambystoma mexicanum

1990 ◽  
Vol 68 (1) ◽  
pp. 159-167 ◽  
Author(s):  
Steven R. Scadding
Keyword(s):  
Electron Microscope ◽  
Vitamin A ◽  
Limb Regeneration ◽  
Ambystoma Mexicanum ◽  
Ciliated Cells ◽  
Posterior Edge ◽  
Histological Changes ◽  
Transmission Electron ◽  
Retinoid Treatment ◽  
Scanning Electron

Vitamin A causes profound changes in the development of pattern during amphibian limb regeneration. Vitamin A effects include the induction of duplications of skeletal structures in the anteroposterior, proximodistal, and dorsoventral axes. The purpose of this investigation was to study the underlying histological changes that are induced in the regenerating limb of the larval axolotl, Ambystoma mexicanum, by treatment with vitamin A. Axolotl larvae (7–10 cm in length) had forelimbs amputated through the radius and ulna and were then immersed in 75 IU/mL retinol palmitate for 14 days. Limbs were removed and fixed at intervals over the period of regeneration, both during and beyond the period of retinoid treatment. They were then examined in the light microscope, scanning electron microscope, or transmission electron microscope. Compared with the controls, the retinol palmitate treated regenerating limbs exhibited the development of an eccentric epidermal cap which was always displaced towards the posterior edge of the limb. Beneath this epidermal cap, the density of the cells of the blastema was greater than that of the cells towards the anterior edge of the developing blastema where the cells were much less densely arranged. Epidermal changes induced by retinol palmitate included the development of a very uneven and creviced surface, with a great deal of variation in cell size, and the development of ciliated cells in the surface layer of the epidermis. The significance of these observations for pattern modification by vitamin A are discussed.

scholarly journals Comparison of Gene Expression During Early Phases of Limb Regeneration Between Regeneration-permissive Neotenic and Regeneration-deficient Metamorphic Axolotl

2019 ◽  
Author(s):  
Mustafa Sibai ◽  
Ebru Altuntaş ◽  
Barış Ethem Süzek ◽  
Betül Şahin ◽  
Cüneyd Parlayan ◽  
...  
Keyword(s):  
Immune System ◽  
Limb Regeneration ◽  
Principal Component ◽  
Ambystoma Mexicanum ◽  
Functional Enrichment ◽  
Limb Amputation ◽  
Regenerative Capacity ◽  
Ecm Remodeling ◽  
Blastema Formation ◽  
Early Phases

ABSTRACTThe axolotl (Ambystoma Mexicanum) salamander, an urodele amphibian, has an exceptional regenerative capacity to fully restore an amputated limb throughout the life-long lasting neoteny. By contrast, when axolotls are experimentally induced to metamorphosis, attenuation of the limb’s regenerative competence is noticeable. Here, we sought to discern the proteomic profiles of the early stages of blastema formation of neotenic and metamorphic axolotls after limb amputation by means of LC-MS/MS technology. We quantified a total of 714 proteins having an adjusted p < 0.01 with FC greater or equal to 2 between two conditions. Principal component analysis revealed a conspicuous clustering between neotenic and metamorphic samples at 7 days post-amputation. Different set of proteins was identified as differentially expressed at all of the time points (1, 4, and 7 days post-amputations against day0) for neotenic and metamorphic stages. Although functional enrichment analyses underline the presence of common pathways between regenerative and nonregenerative stages, cell proliferation and its regulation associated pathways, immune system related pathways and muscle tissue and ECM remodeling and degradation pathways were represented at different rate between both stages. To validate the proteomics results and provide evidence for the putative link between immune system activity and regenerative potential, qRT-PCR for selected genes was performed.

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