Frog Xenopus Laevis Research Articles

Quantification of underwater calling and foraging activities in the African clawed frog Xenopus laevis

Abstract Aquatic anuran species are difficult to detect and observe and this is a major limit to the study of their behaviour and ecology. This habit limits the direct monitoring of sexual and foraging activity, and the investigation of how environmental factors influence their expression as well as how individuals allocate time between competing activities. We investigated this issue in Xenopus laevis, a mostly aquatic frog that forages and emits calls underwater. This model species in biology has been extensively studied in the lab but its behaviour in nature remains poorly described. We carried out a study in a pond during the breeding season in the French invasive range. We recorded underwater vocal activity as a proxy for sexual activity using a hydrophone, set food-baited traps to quantify foraging activity, and recorded environmental conditions (moonlight intensity, temperature and rainfall) over two lunar cycles. We found that individuals engage in these two activities during the breeding season. At the peak of the breeding period, vocal activity was expressed during the day. The investment in reproduction (calling activity) may reduce the time allocated to foraging on a circadian scale. The two activities seem to be partitioned depending on moonlight intensity, with a stronger effect on males. Foraging activity decreased and vocal activity increased when moonlight intensity increased. We also observed a negative effect of temperature and a positive effect of rainfall on vocal activity only. Our method is promising to monitor the activity of other aquatic anurans.

Screening of Chemical Libraries Using Xenopus Embryos and Tadpoles for Phenotypic Drug Discovery.

Phenotypic drug discovery assesses the effect of small molecules on the phenotype of cells, tissues, or whole organisms without a priori knowledge of the target or pathway. Using vertebrate embryos instead of cell-based assays has the advantage that the screening of small molecules occurs in the context of the complex biology and physiology of the whole organism. Fish and amphibians are the only classes of vertebrates with free-living larvae amenable to high-throughput drug screening in multiwell dishes. For both animal classes, particularly zebrafish and Xenopus, husbandry requirements are straightforward, embryos can be obtained in large numbers, and they develop ex utero so their development can be monitored easily with a dissecting microscope. At 350 million years, the evolutionary distance between amphibians and humans is significantly shorter than that between fish and humans, which is estimated at 450 million years. This increases the likelihood that drugs discovered by screening in amphibian embryos will be active in humans. Here, we describe the basic protocol for the medium- to high-throughput screening of chemical libraries using embryos of the African clawed frog Xenopus laevis Bioactive compounds are identified by observing phenotypic changes in whole embryos and tadpoles. In addition to the discovery of compounds with novel bioactivities, the phenotypic screening protocol also allows for the identification of compounds with in vivo toxicity, eliminating early hits that are poor drug candidates. We also highlight important considerations for designing chemical screens, choosing chemical libraries, and performing secondary screens using whole mount in situ hybridization or immunostaining.

A monogenean parasite reveals the widespread translocation of the African clawed frog in its native range

Abstract The management of bio‐invasions relies upon the development of methods to trace their origin and expansion. Cointroduced parasites, especially monogenean flatworms, are ideal tags for the movement of their hosts due to their short generations, direct life cycles and host specificity. However, they are yet to be applied to trace the intraspecific movement of host lineages in their native ranges. As proof of this concept, we conducted a comparative phylogeographic analysis based upon two mitochondrial markers of a globally invasive frog Xenopus laevis and its monogenean parasite Protopolystoma xenopodis in its native range in southern Africa and invasive range in Europe. Translocation of lineages was largely masked in the frog's phylogeography. However, incongruent links between host and parasite phylogeography indicated host switches from one host lineage to another after these were brought into contact in the native range. Thus, past translocation of host lineages is revealed by the invasion success of its cointroduced parasite lineage. This study demonstrates that parasite data can serve as an independent line of evidence in invasion biology, also on the intraspecific level, shedding light on previously undetected invasion dynamics. Based upon the distribution of these invasive parasite lineages, we infer that there is widespread anthropogenic translocation of this frog, not only via official export routes, but also facilitated by the frog's use as live bait by angling communities. Synthesis and applications. Data from cointroduced, host‐specific parasites, as tags for translocation, can add value to investigations in invasion biology and conservation. A better understanding of the translocation history and resulting genetic mixing of host and parasite lineages in the native range can shed light on the genetic make‐up of parasite assemblages cointroduced to the invasive range. Knowledge of the intraspecific movement of different lineages of animals in their native ranges also has conservation implications, since contact between divergent lineages of hosts and parasites can facilitate host switches and altered parasite dynamics in both native and invasive populations. Therefore, we recommend the inclusion of parasite data as a more holistic approach to the invasion ecology of animals on the intraspecific level.

Functions of block of proliferation 1 during anterior development in Xenopus laevis.

Block of proliferation 1 (Bop1) is a nucleolar protein known to be necessary for the assembly of the 60S subunit of ribosomes. Here, we show a specific bop1 expression in the developing anterior tissue of the South African clawed frog Xenopus laevis. Morpholino oligonucleotide-mediated knockdown approaches demonstrated that Bop1 is required for proper development of the cranial cartilage, brain, and the eyes. Furthermore, we show that bop1 knockdown leads to impaired retinal lamination with disorganized cell layers. Expression of neural crest-, brain-, and eye-specific marker genes was disturbed. Apoptotic and proliferative processes, which are known to be affected during ribosomal biogenesis defects, are not hindered upon bop1 knockdown. Because early Xenopus embryos contain a large store of maternal ribosomes, we considered if Bop1 might have a role independent of de novo ribosomal biogenesis. At early embryonic stages, pax6 expression was strongly reduced in bop1 morphants and synergy experiments indicate a common signaling pathway of the two molecules, Bop1 and Pax6. Our studies imply a novel function of Bop1 independent of ribosomal biogenesis.

Aquatic Freshwater Vertebrate Models of Epilepsy Pathology: Past Discoveries and Future Directions for Therapeutic Discovery.

Epilepsy is an international public health concern that greatly affects patients’ health and lifestyle. About 30% of patients do not respond to available therapies, making new research models important for further drug discovery. Aquatic vertebrates present a promising avenue for improved seizure drug screening and discovery. Zebrafish (Danio rerio) and African clawed frogs (Xenopus laevis and tropicalis) are increasing in popularity for seizure research due to their cost-effective housing and rearing, similar genome to humans, ease of genetic manipulation, and simplicity of drug dosing. These organisms have demonstrated utility in a variety of seizure-induction models including chemical and genetic methods. Past studies with these methods have produced promising data and generated questions for further applications of these models to promote discovery of drug-resistant seizure pathology and lead to effective treatments for these patients.

Anesthesia for Echocardiography and Magnetic Resonance Imaging in the African Clawed Frog (Xenopus laevis).

This report describes an anesthesia technique that we used to study cardiovascular anatomy and physiology with echocardiography and cardiac magnetic resonance (CMR) in 46 African clawed frogs (Xenopus laevis) (n = 24 for electrocardiography and n = 22 for CMR). For administration of anesthesia, 3 holding tanks, one each for transportation, sedation, and recovery, were filled with filtered water, with 0.05% buffered tricaine methasulfonate solution (MS-222) added into the sedation tank. Fifteen minutes after the frog was placed in the sedation tank, a paper towel was soaked in MS-222 solution, and the frog was placed in a supine position and rolled 3 to 4 times in the soaked paper with the head and legs exposed. Vital signs were monitored and recorded throughout the procedure. After imagining, frogs were unrolled from the paper towel, placed in the recovery tank, and later returned to their home tank. Monitoring was discontinued when the frogs resumed typical activity. No mortality or complications were observed in frogs that underwent this procedure. Mean duration ±1 SD of anesthesia induction was 12 ± 5 min in the echocardiography group and 14 ± 6 min in the CMR group. The mean duration of anesthesia maintenance was 60 ± 18 min in the echocardiography group and 118 ± 37 min in the CMR group. An additional dose of anesthesia was necessary during maintenance for 9 of 24 (37%) frogs in the echocardiography group and 6 of 22 (27%) frogs in the CMR group. At the end of the procedure, the mean oxygen saturation was 66 ± 9% in the echocardiography group and 85 ± 6% in the CMR group, and heart rate was 48 ± 13 beats/min in the echocardiography group and 42 ± 7 beats/min in the CMR group. We conclude that the anesthesia technique of immersion in MS-222 is suitable for performing echocardiography and CMR imaging in this species without complications.

Normal Table of Xenopus development: a new graphical resource.

ABSTRACTNormal tables of development are essential for studies of embryogenesis, serving as an important resource for model organisms, including the frog Xenopus laevis. Xenopus has long been used to study developmental and cell biology, and is an increasingly important model for human birth defects and disease, genomics, proteomics and toxicology. Scientists utilize Nieuwkoop and Faber's classic ‘Normal Table of Xenopus laevis (Daudin)’ and accompanying illustrations to enable experimental reproducibility and reuse the illustrations in new publications and teaching. However, it is no longer possible to obtain permission for these copyrighted illustrations. We present 133 new, high-quality illustrations of X. laevis development from fertilization to metamorphosis, with additional views that were not available in the original collection. All the images are available on Xenbase, the Xenopus knowledgebase (http://www.xenbase.org/entry/zahn.do), for download and reuse under an attributable, non-commercial creative commons license. Additionally, we have compiled a ‘Landmarks Table’ of key morphological features and marker gene expression that can be used to distinguish stages quickly and reliably (https://www.xenbase.org/entry/landmarks-table.do). This new open-access resource will facilitate Xenopus research and teaching in the decades to come.

Phosphate position is key in mediating transmembrane ion channel TMEM16A–phosphatidylinositol 4,5-bisphosphate interaction

TransMEMbrane 16A (TMEM16A) is a Ca2+-activated Cl− channel that plays critical roles in regulating diverse physiologic processes, including vascular tone, sensory signal transduction, and mucosal secretion. In addition to Ca2+, TMEM16A activation requires the membrane lipid phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2). However, the structural determinants mediating this interaction are not clear. Here, we interrogated the parts of the PI(4,5)P2 head group that mediate its interaction with TMEM16A by using patch- and two-electrode voltage-clamp recordings on oocytes from the African clawed frog Xenopus laevis, which endogenously express TMEM16A channels. During continuous application of Ca2+ to excised inside–out patches, we found that TMEM16A-conducted currents decayed shortly after patch excision. Following this rundown, we show that the application of a synthetic PI(4,5)P2 analog produced current recovery. Furthermore, inducible dephosphorylation of PI(4,5)P2 reduces TMEM16A-conducted currents. Application of PIP2 analogs with different phosphate orientations yielded distinct amounts of current recovery, and only lipids that include a phosphate at the 4′ position effectively recovered TMEM16A currents. Taken together, these findings improve our understanding of how PI(4,5)P2 binds to and potentiates TMEM16A channels.

Patterns of tubb2b Promoter-Driven Fluorescence in the Forebrain of Larval Xenopus laevis.

Microtubules are essential components of the cytoskeleton of all eukaryotic cells and consist of α- and β-tubulin heterodimers. Several tissue-specific isotypes of α- and β-tubulins, encoded by distinct genes, have been described in vertebrates. In the African clawed frog (Xenopus laevis), class II β-tubulin (tubb2b) is expressed exclusively in neurons, and its promoter is used to establish different transgenic frog lines. However, a thorough investigation of the expression pattern of tubb2b has not been carried out yet. In this study, we describe the expression of tubb2b-dependent Katushka fluorescence in the forebrain of premetamorphic Xenopus laevis at cellular resolution. To determine the exact location of Katushka-positive neurons in the forebrain nuclei and to verify the extent of neuronal Katushka expression, we used a transgenic frog line and performed several additional antibody stainings. We found tubb2b-dependent fluorescence throughout the Xenopus forebrain, but not in all neurons. In the olfactory bulb, tubb2b-dependent fluorescence is present in axonal projections from the olfactory epithelium, cells in the mitral cell layer, and fibers of the extrabulbar system, but not in interneurons. We also detected tubb2b-dependent fluorescence in parts of the basal ganglia, the amygdaloid complex, the pallium, the optic nerve, the preoptic area, and the hypothalamus. In the diencephalon, tubb2b-dependent fluorescence occurred mainly in the prethalamus and thalamus. As in the olfactory system, not all neurons of these forebrain regions exhibited tubb2b-dependent fluorescence. Together, our results present a detailed overview of the distribution of tubb2b-dependent fluorescence in neurons of the forebrain of larval Xenopus laevis and clearly show that tubb2b-dependent fluorescence cannot be used as a pan-neuronal marker.

Neofunctionalization of a Noncoding Portion of a DNA Transposon in the Coding Region of the Chimerical Sex-Determining Gene dm-W in Xenopus Frogs.

Most vertebrate sex-determining genes (SDGs) emerge as neofunctionalized genes through duplication and/or mutation of ancestral genes that are involved with sexual differentiation. We previously demonstrated dm-W to be the SDG in the African clawed frog Xenopus laevis and found that a portion of this gene emerged from the masculinization gene dmrt1 after allotetraploidization by interspecific hybridization between two ancestral species around 17–18 Ma. dm-W has four exons consisting of a noncoding exon 1, dmrt1-derived exons 2 and 3, and an orphan exon 4 (Ex4) of unknown origin that includes coding sequence (CDS). In this study, we searched for the origin of Ex4 and investigated the function of the CDS of this exon. We found that the Ex4-CDS is derived from a noncoding portion of the hAT-10 family of DNA transposon. Evolutionary analysis of transposons and determination of the Ex4 sequences from three other species indicated that Ex4 was generated before the diversification of most or all extant allotetraploid species in subgenus Xenopus, during which time we hypothesize that transposase activity of this hAT superfamily was active. Using DNA–protein binding and transfection assays, we further demonstrate that the Ex4-encoded amino acid sequence increases the DNA-binding ability and transrepression activity of DM-W. These findings suggest that the conversion of the noncoding transposon sequence to the CDS of dm-W contributed to neofunctionalization of a new chimeric SDG in the ancestor of the allotetraploid Xenopus species, offering new insights into de novo origin and functional evolution of chimerical genes.

An in vivo study of the toxic effects of triclosan on Xenopus laevis (Daudin, 1802) frog: Assessment of viability, tissue damage and mitochondrial dysfunction

The present study describes the in vivo effect of triclosan on the frog Xenopus laevis (Daudin, 1802). We have found a dose-dependence of the effect of triclosan on the survival of frogs. At a dose of 2 mg/L, the death of frogs was observed already on the 4th day of the experiment, while at a concentration of 0.5 mg/L, the frogs remained viable for 11 days. Triclosan caused damage to the liver tissue, which was expressed in an increase in the area of hemorrhage and the number of melanomacrophage centers. 0.5 mg/L of this agent did not affect the number of frog red blood cells, but reduced their osmotic resistance. Keeping animals in water containing triclosan (0.5 mg/L for 96 h) led to the suppression of the state 3 respiration rate of frog liver mitochondria. This effect was accompanied by suppression of the combined activity of complexes II and III of the mitochondrial respiratory chain. In parallel with this, we observed a reduction in the Ca2+ retention capacity of frog liver mitochondria, indicating a decrease in the resistance of organelles to mitochondrial permeability transition pore opening. The paper discusses the effects of triclosan on aquatic organisms.

A Review on Structure - Activity Relationship of Antimicrobial Peptide Magainin 2

The rapid development of antibacterial resistance has increased the need for the discovery of novel antimicrobial agents. In this context, an emerging class of molecules called antimicrobial peptides (AMPs) has been considered an excellent material for introducing novel antibiotics. Among the wide variety of AMPs, magainin 2 is a 23 residue (GIGKFLHSAKKFGKAFVGEIMNS) peptide obtained from the African clawed frog Xenopus laevis. Different mutants of magainin 2 have been studied to investigate the structure-function relationship. In the present study, we recapitulate the effects of modifying several structural parameters of magainin 2 such as peptide length, amino acid composition, net charge, alpha helicity, hydrophobicity, hydrophobic moment and amphiphilicity on its antibacterial activity. Moreover, we summarized the hemolytic activity regarding these modifications. An optimum change of these parameters would provide desired antibacterial activity with minimum hemolysis. This structurefunction relationship study could be used as a template in designing novel functional antibacterial agents. Dhaka Univ. J. Pharm. Sci. 20(3): 427-434, 2022 (June) Centennial Special Issue

Biochemical and osmoregulatory responses of the African clawed frog experimentally exposed to salt and pesticide

Salinization and pollution are two main environmental stressors leading deterioration to water quality and degradation of aquatic ecosystems. Amphibians are a highly sensitive group of vertebrates to environmental disturbance of aquatic ecosystems. However, studies on the combined effect of salinization and pollution on the physiology of amphibians are limited. In this study, we measured the standard metabolic rate (SMR) and biochemical parameters of adult males of the invasive frog Xenopus laevis after 45 days of exposure to contrasting salinity environments (400 and 150 mOsm NaCl) with either 1.0 μg/L of the organophosphate pesticide chlorpyrifos (CPF) or pesticide-free medium. Our results revealed a decrease in SMR of animals exposed to the pesticide and in the ability to concentrate the plasma in animals exposed simultaneously to both stressors. The lack of ability to increase plasma concentration in animals exposed to both salt water and CPF, suggests that osmoregulatory response is decreased by pesticide exposure. In addition, we found an increase of liver citrate synthase activity in response to salt stress. Likewise, the liver acetylcholinesterase (AChE) activity decreased by 50% in frogs exposed to salt water and CPF and 40% in those exposed only to CPF, which suggest an additive effect of salinity on inhibition of AChE. Finally, oxidative stress increased as shown by the higher lipid peroxidation and concentration of aqueous peroxides found in the group exposed to salt water and pesticide. Thus, our results revealed that X. laevis physiology is compromised by salinization and pesticide exposure to both environmental stressors join.

Sequence of chondrocranial development in basal anurans\u2014Let\u2019s make a cranium

BackgroundThe craniofacial skeleton is an evolutionary innovation of vertebrates. Due to its complexity and importance to protect the brain and aid in essential functions (e.g., feeding), its development requires a precisely tuned sequence of chondrification and/or ossification events. The comparison of sequential patterns of cartilage formation bears important insights into the evolution of development. Discoglossus scovazzi is a basal anuran species. The comparison of its chondrocranium (cartilaginous neuro- & viscerocranium) development with other basal anurans (Xenopus laevis, Bombina orientalis) will help establishing the ancestral pattern of chondrification sequences in anurans and will serve as basis for further studies to reconstruct ancestral conditions in amphibians, tetrapods, and vertebrates. Furthermore, evolutionary patterns in anurans can be studied in the light of adaptations once the ancestral sequence is established.ResultsWe present a comprehensive overview on the chondrocranium development of D. scovazzi. With clearing and staining, histology and 3D reconstructions we tracked the chondrification of 44 elements from the first mesenchymal Anlagen to the premetamorphic cartilaginous head skeleton and illustrate the sequential changes of the skull. We identified several anuran and discoglossoid traits of cartilage development. In D. scovazzi the mandibular, hyoid, and first branchial arch Anlagen develop first followed by stepwise addition of the branchial arches II, III, and IV. Nonetheless, there is no strict anterior to posterior chondrification pattern within the viscerocranium of D. scovazzi. Single hyoid arch elements chondrify after elements of the branchial arch and mandibular arch elements chondrify after elements of the branchial arch I.ConclusionsIn Osteichthyes, neurocranial elements develop in anterior to posterior direction. In the anurans investigated so far, as well as in D. scovazzi, the posterior parts of the neurocranium extend anteriorly, while the anterior parts of the neurocranium, extend posteriorly until both parts meet and fuse. Anuran cartilaginous development differs in at least two crucial traits from other gnathostomes which further supports the urgent need for more developmental investigations among this clade to understand the evolution of cartilage development in vertebrates.

Cell-Lineage Guided Mass Spectrometry Proteomics in the Developing (Frog) Embryo.

Characterization of molecular events as cells give rise to tissues and organs raises a potential to better understand normal development and design efficient remedies for diseases. Technologies enabling accurate identification and quantification of diverse types and large numbers of proteins would provide still missing information on molecular mechanisms orchestrating tissue and organism development in space and time. Here, we present a mass spectrometry-based protocol that enables the measurement of thousands of proteins in identified cell lineages in Xenopus laevis (frog) embryos. The approach builds on reproducible cell-fate maps and established methods to identify, fluorescently label, track, and sample cells and their progeny (clones) from this model of vertebrate development. After collecting cellular contents using microsampling or isolating cells by dissection or fluorescence-activated cell sorting, proteins are extracted and processed for bottom-up proteomic analysis. Liquid chromatography and capillary electrophoresis are used to provide scalable separation for protein detection and quantification with high-resolution mass spectrometry (HRMS). Representative examples are provided for the proteomic characterization of neural-tissue fated cells. Cell-lineage-guided HRMS proteomics is adaptable to different tissues and organisms. It is sufficiently sensitive, specific, and quantitative to peer into the spatio-temporal dynamics of the proteome during vertebrate development.

Cell-Lineage Guided Mass Spectrometry Proteomics in the Developing (Frog) Embryo

Characterization of molecular events as cells give rise to tissues and organs raises a potential to better understand normal development and design efficient remedies for diseases. Technologies enabling accurate identification and quantification of diverse types and large numbers of proteins would provide still missing information on molecular mechanisms orchestrating tissue and organism development in space and time. Here, we present a mass spectrometry-based protocol that enables the measurement of thousands of proteins in identified cell lineages in Xenopus laevis (frog) embryos. The approach builds on reproducible cell-fate maps and established methods to identify, fluorescently label, track, and sample cells and their progeny (clones) from this model of vertebrate development. After collecting cellular contents using microsampling or isolating cells by dissection or fluorescence-activated cell sorting, proteins are extracted and processed for bottom-up proteomic analysis. Liquid chromatography and capillary electrophoresis are used to provide scalable separation for protein detection and quantification with high-resolution mass spectrometry (HRMS). Representative examples are provided for the proteomic characterization of neural-tissue fated cells. Cell-lineage-guided HRMS proteomics is adaptable to different tissues and organisms. It is sufficiently sensitive, specific, and quantitative to peer into the spatio-temporal dynamics of the proteome during vertebrate development.

Capillary Electrophoresis Mass Spectrometry for Scalable Single-Cell Proteomics.

Understanding the biochemistry of the cell requires measurement of all the molecules it produces. Single-cell proteomics recently became possible through advances in microanalytical sample preparation, separation by nano-flow liquid chromatography (nanoLC) and capillary electrophoresis (CE), and detection using electrospray ionization (ESI) high-resolution mass spectrometry (HRMS). Here, we demonstrate capillary microsampling CE-ESI-HRMS to be scalable to proteomics across broad cellular dimensions. This study established proof-of-principle using giant, ∼250-µm-diameter cells from embryos of the frog Xenopus laevis and small, ∼35-µm-diameter neurons in culture from the mouse hippocampus. From ∼18 ng, or ∼0.2% of the total cellular proteome, subcellular analysis of the ventral-animal midline (V11) and equatorial (V12) cells identified 1,133 different proteins in a 16-cell embryo. CE-HRMS achieved ∼20-times higher sensitivity and doubled the speed of instrumental measurements compared to nanoLC, the closest neighboring single-cell technology of choice. Microanalysis was scalable to 722 proteins groups from ∼5 ng of cellular protein digest from identified left dorsal-animal midline cell (D11), supporting sensitivity for smaller cells. Capillary microsampling enabled the isolation and transfer of individual neurons from the culture, identifying 37 proteins between three different cells. A total of 224 proteins were detected from 500 pg of neuronal protein digest, which estimates to a single neuron. Serial dilution returned 157 proteins from sample amounts estimating to about half a cell (250 pg protein) and 70 proteins from ca. a quarter of a neuron (125 pg protein), suggesting sufficient sensitivity for subcellular proteomics. CE-ESI-HRMS complements nanoLC proteomics with scalability, sensitivity, and speed across broad cellular dimensions.

Microplastics ingestion induces plasticity in digestive morphology in larvae of Xenopus laevis

Global changes in temperature, predator introductions, and pollution might challenge animals by altering food conditions. A fast-growing source of environmental pollution are microplastics. If ingested with the natural food source, microplastics act as artificial fibers that reduce food quality by decreasing nutrient and energy density with possible ramifications for growth and development. Animals might cope with altered food conditions with digestive plasticity. We examined experimentally whether larvae of the African clawed frog (Xenopus laevis) exhibit digestive morphology plasticity (i.e., gut length, mass, and diameter) in response to microplastics ingestion. As natural systems contain non-digestible particles similar in size and shape to microplastics, we included cellulose as a natural fiber control group. Gut length and mass increased in response to microplastics and cellulose ingestion indicating that both types of fibers induced digestive plasticity. Body mass and body condition were similar across experimental groups, indicating that larvae fully compensated for low nutrient and energy density by developing longer intestines. The ability of a species to respond plastically to environmental variation, as X. laevis responded, indicates that this species might have the potential to cope with new conditions during global change, although it is uncertain whether this potential may be reduced in a multi-stressor environment.

Meat storage under acoustic freezing conditions and assessment of its biotoxicity (part 2)

The necessity of developing new technologies for refrigerated storage, providing improved quality and safety of meat products, is shown. A critical analysis of cold storage methods has been carried out and the choice of the acoustic freezing method has been substantiated. As a result of the joint work of the students of school No. 2083, “Acoustic Freezing” company and the Institute of Biotechnology and Fisheries, Moscow State University of Technology and Management named after K.G. Razumovsky, studies were carried out to establish the degree of toxicity of pork subjected to refrigeration storage under conditions of shock and acoustic freezing. Histological studies of fresh meat and meat after acoustic and shock freezing have shown that shock freezing leads to a violation of the structure of striated muscles due to the formation of large ice crystals during freezing. The method of fractal analysis of proteins in fresh meat and meat after acoustic freezing showed that the ice crystallization in muscle tissue leads to the formation of different types of fractals. Fractal analysis can be used to select physical factors when improving the technology of acoustic freezing. Toxicological studies were carried out on juvenile clawed frogs Xenopus laevis fed with chironomid larvae, California worms Eisenia fetida (control), fresh meat and meat after acoustic freezing. Studies have shown that there are no significant deviations in the cytological parameters of red blood in frogs eating natural food (worms) and meat after acoustic freezing. Meat after shock freezing is eaten poorly by frogs, which indicates a significant loss of its quality compared to fresh meat. Comparative studies on the effects of feeding fresh meat and meat after acoustic freezing, carried out on the blood of laboratory fish Danio rerio, demonstrated that fresh meat and meat after acoustic freezing do not cause harmful effects on the red blood of fish. The work was carried out within the framework of the program for the introduction of project methods of school education in the secondary school of the Moscow State Budgetary Educational Institution “School No. 2083”.

Rejeneratif tıpta model organizma; Aksolotl (Ambystoma Mexicanum)

The axolotl has an extraordinary capacity to regenerate damaged and lost structures, especially the nervous system, limbs, organs such as the eye and heart, without causing scarring. It has become a very important model organism by attracting the attention of scientists working in both developmental biology and regenerative medicine and stem cell biology. The axolotl, which is amphibian and tetrapod, is a more preferred model due to its ease of maintenance and reproduction compared to other organisms such as African clawed frog (Xenopus laevis) or zebrafish (Danio rerio), which are relatively difficult to study. The main purposes of this review are the definition and origin of the axolotl, its taxonomy, anatomy, reproduction, nutrition, habitat, to give a perspective to scientists who want to work on this model organism by giving examples to the scientific data and study fields of the last 20 years by addressing issues such as how it contributes to scientific studies as a model organism.