Young Axolotls Research Articles

Natural and induced apoptosis during lymphocyte development in the axolotl

Lymphocytes apoptosis was characterized in a urodele amphibian, the axolotl, by morphology using electron microscopy and by flow cytometry after propidium iodide staining, as well as by biochemical criteria with the detection of DNA ladders after glucocorticoid treatment. The morphological and biochemical features observed in treated axolotls are in accordance with the criteria of apoptosis found in different models of mammalian lymphocyte programmed cell death. The onset of natural apoptosis was then detected by DNA fragmentation in thymus and in spleen during lymphocyte development and ontogenesis. A typical DNA ladder characteristic of apoptosis is detectable in the thymus as early as 5 months; apoptosis increases and peaks at 8 months, and is no longer detected by 10 months or thereafter. The ability of a superantigen, Staphylococcus aureus enterotoxin B (SEB), to induce T lymphocyte apoptosis in larvae was investigated as well. In vivo exposure of young axolotl larvae to SEB induces, as in mammals, thymocyte apoptosis as indicated by the enhancement of DNA fragmentation. These last results, natural programmed cell death and SEB induced apoptosis during thymic ontogeny, are discussed in correlation with what is known during mammalian thymic selection and apoptosis.

Activation by mitogens and superantigens of axolotl lymphocytes: functional characterization and ontogenic study.

Urodele amphibians have weak and slow immune responses compared to mammals and anuran amphibians. Using new culture conditions, we tested the ability of lymphocytes of a well-studied salamander, the Mexican axolotl (Ambystoma mexicanum) to proliferate in vitro with diverse mitogenic agents. We demonstrated that the axolotl has a population of B lymphocytes that proliferate specifically and with a high stimulation index to the lipopolysaccharide (LPS) known as a B-cell mitogen in mammals. This proliferative capacity is observed without significant changes throughout ontogenesis. In the presence of LPS, axolotl B lymphocytes are able to synthesize and secrete both isotopes of immunoglobulin described in this species, IgM and IgY. Moreover, a distinct lymphocyte subpopulation is able to poliferate significantly in response to the mitogens usually known as T-cell specific in mammals, phytohaemagglutinin (PHA) and concanavalin A (Con A). The activated cells are T lymphocytes, as shown by depletion experiments performed in vitro with monoclonal antibodies, and in vivo by thymectomy. Splenic T lymphocytes of young axolotls (before 10 months) do not have this functional ability, which suggests maturation and/or migration phenomena during T-cell ontogenesis in this species. Axolotl lymphocytes are able to proliferate in vitro with a significant stimulation index to staphylococcal enterotoxins A and B (SEA and SEB). These products act on mammalian lymphocytes as superantigens: in combination with products of the major histocompatibility complex (MHC), they bind T-cell receptors with particular V beta elements. The fact that these superantigens are able to activate lymphocytes of a primitive vertebrate suggests a striking conservation of molecular structures implied in superantigen presentation and recognition.

Blastema cell proliferation in vitro: Effects of limb amputation on the mitogenic activity of spinal cord extracts

Primary cultures of mesenchymal cells of axolotl limb blastemas provide a very sensitive in vitro bioassay for studying nerve dependence of newt regeneration. These cells can be stimulated by crude spinal cord extracts of non-amputated animals in a dose-dependent manner up to 60 micrograms protein/ml of culture medium; at this concentration the mitotic index is increased 4-fold. Spinal cord extracts of axolotls 14 days after forelimb amputation (i.e., late bud stage) are more efficient in stimulating blastema cell proliferation (+50%) than extracts of axolotls 7 days after forelimb amputation (i.e., early bud stage) or of axolotls without amputation. In a similar manner, spinal cord extracts of young axolotls 14 days after forelimb amputation, are more stimulatory than older axolotls 14 d after forelimb amputation which regenerate only a very small blastema during the same time. It appears that spinal cord mitogenic activity is enhanced after limb amputation, probably in correlation with blastema cell requirements for limb regeneration.

The regeneration of axolotl limbs covered by frog skin

Forearm skin of Stage XXIV Rana pipiens, which cannot regenerate limbs, was removed and placed upon the skinned forearms of young axolotls. The axolotl limbs were amputated immediately through the level of the grafts. Frog epidermis migrated to cover the amputation surface. Dedifferentiation and early blastema formation occurred beneath the frog wound epidermis. Limb regeneration continued, but in time axolotl epidermis overgrew the frog epidermis. The experiment shows that epidermis from nonregenerating frog limbs is still capable of supporting typical epimorphic regeneration.

Regeneration of reversed aneurogenic arms of the axolotl

Aneurogenic arms of young axolotls were implanted into the flank as heterotopic autografts with reversed proximo-distal orientation. The formerly proximal ends of such arms regressed to a variable extent, and then either regenerated or could do so following a second amputation. The regenerate always contained a complete sequence of skeletal elements between the adjacent stump skeleton and terminal digits, being a mirror image of the implanted arms with identical transverse axes but an opposed proximo-distal axis. Many reversed arms also regenerated fingers from the implanted hand. Identical results were obtained from reversed arms of control larvae, confirming previous studies on reversed well-innervated arms. Nerves are not required for the establishment of a new proximo-distal axis, therefore, and probably have no influence on the determination of any limb axis. Morphogenesis of the regenerate is clearly related to the position along the proximo-distal axis where the blastema originates. Although this axis is reasonably envisaged in terms of a gradient, its polarity is ignored during regeneration.

Patterning in synaptic knobs which connect with Mauthner's cell (Ambystoma mexicanum).

AbstractThe Mauthner's cells (M‐cells) of the young axolotl larva receive a rich afferent supply of synaptic knobs (club and bulb synapses). The knobs may be classified into at least five types according to their ultrastructure after direct fixation in OsO4. The different types are non‐randomly arranged on the M‐cell surface such that groups of knobs of the same type are frequently observed to form a cluster or patch. The M‐cell surface pattern is thus a mosaic of such clusters. Different types of knobs may be restricted to different locations on the M‐cell; one type is found almost exclusively with the M‐cell lateral dendrites, and another two types with the axon hillock and axonal initial segment. The dorsal perikaryon is nearly devoid of knobs. This pattern is at least in part related to the pattern of fasciculation in the adjacent neural tissue, and it is likely that all or some of the distinctive types of knobs originate from different kinds of neurons.The pattern of distribution of the synaptic knobs appears to increase in complexity during development, and this can be related to the asymmetrical growth of the M‐cell during its maturation. These observations are discussed with reference to theories of neuronal specificity.

The response of the cornea of the adult axolotl ( Siredon mexicanum) to X-radiation

The anterior portion of the head was irradiated in 70 adult axolotls (Mexican salamanders of the species Siredon mexicanum), 4–5 years old, using 1000 r in 9 animals, 3000 r in 20, 6000 r in 10, 8000 r in 11, and 10,000 r in 20; 5 animals served as untreated controls. Specimens fixed at various intervals following irradiation were studied histologically. After irradiation with 1000 r, the substantia propria of the cornea was almost normal. The corneal epithelium, however, became much thicker, and melanophores and Leydig cells developed in it. After irradiation with 3000 r, the substantia propria was much thicker than normal, and underwent stratification; cavities and pigment cells developed in it. In the epithelium, melanophores and Leydig cells developed, and sometimes cavities formed between the cells, and vacuoles appeared in the cytoplasm. After irradiation with 6000–10,000 r, the substantia propria underwent thickening and stratification, and blood vessels penetrated into it. The epithelium became much thicker, giant cells developed, and melanophores and Leydig cells were formed. Sometimes the corneal epithelium disappeared and was replaced by skin epithelium. Some phenomena typical of the response of young axolotls to radiation (e.g., the formation of abnormal outgrowths on the cornea) were not observed in adult animals. On the other hand, the development of melanophores and blood vessels in the substantia propria occurred in the adults, but not in the young animals. The development of melanophores and blood vessels in areas of the irradiated cornea where they do not occur in the nonirradiated cornea may be considered a result of X-ray stimulation. Most of the histopathological changes that were observed in the irradiated cornea in axolotls are not peculiar to amphibians; they have been reported by various investigators to occur in mammals, including man.

Successive Changes in the Cornea of Young Axolotl (Siredon mexicanum) after X-Irradiation

The heads of 287 small axolotls were irradiated locally with 9000, 6000, 3000, 1000, 500, or 250 r. Fifty animals served as untreated controls. A histologic study of the specimens fixed at short intervals of irradiated and control corneas was made. After irradiation with 9000 r, the first change was noted 8 days later, and it progressed gradually until 18 days when the last surviving animal was fixed. The outer surface of the cornea became uneven, the cells eniarged, and melanophores appeared in the epithelium. The isolated projections of separate cells could be noted. The cornea gradually became completely disorganized and completely destroyed. After irradiation with 6000 r, the first changes in the corneal epithelium could be noted 18 days after treatment. Thirty days after irradiation, many very great compact melanophores andd small pigment granules were observed in the corneal epithelium. The number of cells were greatly reduced, and some of them were of giant size. Isolated projections from the epithelium were also observed. After irradiation with 3000 r, definite morphological changes of the epithelium were observed 22 days after irradiation. Thirty-five days after irradiation, disorganization of the corneal epithelium was observed. Only isolated giant epithelial cells were seen. Aftermore » complete destruction of the corneal epithelium, it was replaced by a typical skin epithelium with many Leydig cells. After irradiation with 1000 r and 500 r, the typical primary changes were observed at 21 days. In some cases, the corneal epithelium completely recovered, and in others it completely disappeared and was replaced with skin epithelium. After irradiation with 250 r, destruction of the corneal epithelium was not observed, but in some cases a very thick corneal epithelium was seen which was three to four times as thick as the epithelium of control animals. The development of melanophores in the cornea is a result of irradiation. Normally, the cornea has no melanophores. In control animals they were never observed in the cornea. The peculiar resistance to irradiation of eye pigment cells in the chloroid is emphasized and contrasts greatly with sensitivity of melanophores in other regions of the body of amphibians and mammals, e.g., graying of hairs after irradiation. Resistance and the progressive development of melanophores in new regions such as epithelium or cornea were observed. (auth)« less

The Effect of Total-Body X-Irradiation on the Adult Axolotl (Siredon mexicanum)

The importance of age as a factor in the radiosensitivity of animals is a wellestablished fact. In general, the older the organism, the less is the radiosensitivity. According to Abrams (1), however, mice most sensitive to X-rays are about 30 days old, whereas younger ones are less sensitive. The difference in radiosensitivity of young axolotls of various ages was shown by Sheremetjeva (2, 3). According to this author, 150 r is the lethal dose for freshly hatched axolotls, all of which died within one year. Some animals that were irradiated with the same dose only 5 days later (6 days after hatching) lived 3 years. The lethal dose for 100 % of animals irradiated 20 days after hatching is 300 r, and for animals irradiated 30 days after hatching it is 450 r. The sensitivity of adult axolotls to total-body X-irradiation has not been reported, and this is the purpose of the present study. The first specific problem considered is the comparison of the lethal effects of various doses on survival time of adult exolotls. The second specific problem is the correlation of tissue damage with lethality. The third specific problem is the comparison of the reactions to irradiation of adult axolotls with reactions of young animals.

Investigation of the Lethal Effect of X-Irradiation on the Young Axolotl (Siredon mexicanum)

The extensive literature concerning the lethal effect of X-irradiation consists mainly of investigations of this problem in mammals. For instance, the papers of Ellinger and Barnett (1, 2), Henshaw (3), Quastler (4), and Tullis and co-workers (5) may be mentioned. In order to understand the basic biological actions of X-irradiation it is important to investigate the lethal effect in various animal groups. The use of amphibia in these investigations is desirable because they are convenient laboratory animals very suitable for histological investigation. The most important contributions concerning this problem in amphibia are the works of Rugh (6-10), Sheremetieva (11), and Steamer (12). In spite of the fact that these papers give some very important data, the problem of lethality of irradiation is not yet completely resolved, and investigations in this direction are important. The first specific problem considered in this paper is the comparison of the effects of total-body and partial-body irradiation. The second specific problem is correlation of tissue damage with lethality.

The effect of local x-ray irradiation upon the teeth and surrounding tissues in young axolotls (Siredon mexicanum).

The effect of local x-ray irradiation upon the teeth and surrounding tissues in young axolotls (Siredon mexicanum).

Influence of local x-ray treatment on development of jaws in young axolotl (Siredon mexicanum).

SummaryLower or upper jaws of young axolotls (age 20 to 60 days) were locally irradiated with 2000-4000 r. Suppression of growth of treated jaws was observed in all cases. The irradiated jaw is a miniature copy of the normal jaw. Tissue reaction of irradiated jaw was evident 23 days after treatment with 4000 r (primary acute reaction). The treated jaw at this time showed a typical degenerating giant cell epithelium. The enamel epithelium disappeared and resorption of teeth was in progress. Thirty-four days after irradiation there remained small remnants of teeth in parts of treated jaws. At this time, when the resorption of teeth is almost completed, only a small retardation of growth of the irradiated jaw was observed. The degenerating giant cell epithelium has disappeared and been replaced with normal-appearing epithelium, but this epithelium was unable to produce new enamel organs. Fifty-eight days after treatment the retardation of growth of the treated jaws was clearly evident, and all remnants of te...

The development of secondary tails in young axolotls after local x‐ray ibradiation

Journal of MorphologyVolume 89, Issue 1 p. 111-133 Article The development of secondary tails in young axolotls after local x-ray ibradiation† V. V. Brunst, V. V. Brunst Department of Anatomy, University of Maryland, School of Medicine, BaltimoreSearch for more papers by this authorFrank H. J. Figge, Frank H. J. Figge Department of Anatomy, University of Maryland, School of Medicine, BaltimoreSearch for more papers by this author V. V. Brunst, V. V. Brunst Department of Anatomy, University of Maryland, School of Medicine, BaltimoreSearch for more papers by this authorFrank H. J. Figge, Frank H. J. Figge Department of Anatomy, University of Maryland, School of Medicine, BaltimoreSearch for more papers by this author First published: July 1951 https://doi.org/10.1002/jmor.1050890108Citations: 7 † This work was supported by grants from the National Cancer Institute of U.S.P.H.S. and the Anna Fuller Fund. AboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onFacebookTwitterLinked InRedditWechat Citing Literature Volume89, Issue1July 1951Pages 111-133 RelatedInformation

Influence of local x‐ray treatment on the development of extremities of the young axolotl (Siredon mexicanum)

Influence of local x‐ray treatment on the development of extremities of the young axolotl (Siredon mexicanum)

The effects of local x‐ray irradiation on the tail development of young axolotls

The effects of local x‐ray irradiation on the tail development of young axolotls