Embryonic Development Of Xenopus Laevis Research Articles

Assessing the impact of antiviral drugs commonly utilized during the COVID-19 pandemic on the embryonic development of Xenopus laevis

The antiviral drugs favipiravir and oseltamivir are widely used to treat viral infections, including coronavirus 2019 (COVID-19), and their levels are expected to increase in the aquatic environment. In this study, the potential toxic and teratogenic effects of these drugs were evaluated using the frog embryo teratogenesis assay Xenopus (FETAX). In addition, glutathione S-transferase (GST), glutathione reductase (GR), catalase, carboxylesterase (CaE), and acetylcholinesterase (AChE) enzyme activities and malondialdehyde levels were measured as biochemical markers in embryos and tadpoles for comparative assessment of the sublethal effects of the test compounds. Prior to embryo exposure, drug concentrations in the exposure medium were measured with high-performance liquid chromatography. The 96-h median lethal concentration (LC50) was 137.9 and 32.3 mg/L for favipiravir and oseltamivir, respectively. The teratogenic index for favipiravir was 4.67. Both favipiravir and oseltamivir inhibited GR, CaE, and AChE activities in embryos, while favipiravir increased the GST and CaE activities in tadpoles. In conclusion, favipiravir, for which teratogenicity data are available in mammalian test organisms and human teratogenicity is controversial, inhibited Xenopus laevis embryo development and was teratogenic. In addition, sublethal concentrations of both drugs altered the biochemical responses in embryos and tadpoles, with differences between the developmental stages.

Investigation of the effects of metal oxide nanoparticle mixtures on Danio rerio and Xenopus laevis embryos

ABSTRACT In this study, the toxic effects of single TiO2, ZnO and Fe2O3 nanoparticles (NPs) and their binary and ternary combinations on the embryonic development of Xenopus laevis and Danio rerio were evaluated. It was aimed to determine the median lethal concentrations, growth retardation and developmental malformation effects and the effects of mixed nanoparticle exposure to selected enzyme biomarkers resulting from the exposure of embryos to NPs. The results showed that ZnO NPs, which are the most toxic according to the highest lethality level, cause developmental delay and malformations. The Fe2O3 and TiO2 NPs caused growth retardation only in D. rerio embryos. It was determined that double and triple combinations of NPs did not cause significant lethality in both species. Biochemical markers showed that mixed NPs could lead to greater toxicity compared to single exposures. The results also revealed that TiO2 or Fe2O3 NPs caused an increase in toxicity when co-existing with ZnO NP due to additive effects. Overall, the results showed that D. rerio was more susceptible to NP exposure in terms of growth and mortality than X. laevis. However, studies that are more comprehensive are needed to better understand the toxicity mechanism of metal oxide NP mixtures.

Impact of glyphosate-based herbicide on early embryonic development of the amphibian Xenopus laevis.

Worldwide, amphibian populations are declining drastically. One reason might be the use of pesticides including herbicides. The herbicide glyphosate is an inhibitor of the 5-enolpyruvylshikimate-3-phosphate (EPSP) synthase of the plant shikimate pathway, preventing the formation of aromatic amino acids and thus inducing plant death. Due to this specific action, GBH are considered nontoxic to non-target organisms. However, GBH impairs embryonic development of chickens, amphibians and fishes. So far, no detailed tissue- and organ-specific analysis of the effects of GBH during development in amphibians has been performed. We demonstrated that GBH Roundup® LB plus has a negative effect on embryonic development of the South African clawed frog Xenopus laevis. GBH treatment with sublethal concentrations resulted in a reduced body length and mobility of embryos. Furthermore, incubation with GBH led to smaller eyes, brains and cranial cartilages in comparison to untreated embryos. GBH incubation also resulted in shorter cranial nerves and had an effect on cardiac development including reduced heart rate and atrium size. On a molecular basis, GBH treatment led to reduced expression of marker genes in different tissues and developmental stages. GBH leads to disturbed embryonic development of Xenopus laevis.

Systematic mapping of rRNA 2'-O methylation during frog development and involvement of the methyltransferase Fibrillarin in eye and craniofacial development in Xenopus laevis.

Ribosomes are essential nanomachines responsible for protein production. Although ribosomes are present in every living cell, ribosome biogenesis dysfunction diseases, called ribosomopathies, impact particular tissues specifically. Here, we evaluate the importance of the box C/D snoRNA-associated ribosomal RNA methyltransferase fibrillarin (Fbl) in the early embryonic development of Xenopus laevis. We report that in developing embryos, the neural plate, neural crest cells (NCCs), and NCC derivatives are rich in fbl transcripts. Fbl knockdown leads to striking morphological defects affecting the eyes and craniofacial skeleton, due to lack of NCC survival caused by massive p53-dependent apoptosis. Fbl is required for efficient pre-rRNA processing and 18S rRNA production, which explains the early developmental defects. Using RiboMethSeq, we systematically reinvestigated ribosomal RNA 2'-O methylation in X. laevis, confirming all 89 previously mapped sites and identifying 15 novel putative positions in 18S and 28S rRNA. Twenty-three positions, including 10 of the new ones, were validated orthogonally by low dNTP primer extension. Bioinformatic screening of the X. laevis transcriptome revealed candidate box C/D snoRNAs for all methylated positions. Mapping of 2'-O methylation at six developmental stages in individual embryos indicated a trend towards reduced methylation at specific positions during development. We conclude that fibrillarin knockdown in early Xenopus embryos causes reduced production of functional ribosomal subunits, thus impairing NCC formation and migration.

Physiological Functions of Thiol Peroxidases (Gpx1 and Prdx2) during Xenopus laevis Embryonic Development.

Glutathione peroxidase 1 (Gpx1) and peroxiredoxin 2 (Prdx2) belong to the thiol peroxidase family of antioxidants, and have been studied for their antioxidant functions and roles in cancers. However, the physiological significance of Gpx1 and Prdx2 during vertebrate embryogenesis are lacking. Currently, we investigated the functional roles of Gpx1 and Prdx2 during vertebrate embryogenesis using Xenopus laevis as a vertebrate model. Our investigations revealed the zygotic nature of gpx1 having its localization in the eye region of developing embryos, whereas prdx2 exhibited a maternal nature and were localized in embryonic ventral blood islands. Furthermore, the gpx1-morphants exhibited malformed eyes with incompletely detached lenses. However, the depletion of prdx2 has not established its involvement with embryogenesis. A molecular analysis of gpx1-depleted embryos revealed the perturbed expression of a cryba1-lens-specific marker and also exhibited reactive oxygen species (ROS) accumulation in the eye regions of gpx1-morphants. Additionally, transcriptomics analysis of gpx1-knockout embryos demonstrated the involvement of Wnt, cadherin, and integrin signaling pathways in the development of malformed eyes. Conclusively, our findings indicate the association of gpx1 with a complex network of embryonic developmental pathways and ROS responses, but detailed investigation is a prerequisite in order to pinpoint the mechanistic details of these interactions.

Eya1 protein distribution during embryonic development of Xenopus laevis

Eya1 and other Eya proteins are important regulators of progenitor proliferation, cell differentiation and morphogenesis in all three germ layers. At present, most of our knowledge of Eya1 distribution is based on in situ hybridization for Eya1 mRNA. However, to begin to dissect the mechanisms underlying Eya1 functions, we need a better understanding of the spatiotemporal distribution of Eya1 proteins during embryonic development, their subcellular localization and their levels of expression in various tissues. Here we report the localization of Eya1 protein throughout embryonic development from neural plate stages to tadpole stages of Xenopus laevis using a specific antibody for Xenopus Eya1. Our study confirms the expression of Eya1 protein in cranial placodes, placodally derived sensory primordia (olfactory epithelium, otic vesicle, lateral line primordia) and cranial ganglia, as well as in somites, secondary heart field and pharyngeal endoderm. In addition, we report here a novel expression of Eya1 proteins in scattered epidermal cells in Xenopus. Our findings also reveal that, while being predominantly expressed in nuclei in most expression domains, Eya1 protein is also localized to the cytoplasm, in particular in the early preplacodal ectoderm, some placode-derived ganglia and a subset of epidermal cells. While some cytoplasmic roles of Eya1 have been previously described in other contexts, the functions of cytoplasmic Eya1 in the preplacodal ectoderm, cranial ganglia and epidermal cells remain to be investigated.

Optogenetic Control of the Canonical Wnt Signaling Pathway During Xenopus laevis Embryonic Development

Optogenetics uses light-inducible protein-protein interactions to precisely control the timing, localization, and intensity of signaling activity. The precise spatial and temporal resolution of this emerging technology has proven extremely attractive to the study of embryonic development, a program faithfully replicated to form the same organism from a single cell. We have previously performed a comparative study for optogenetic activation of receptor tyrosine kinases, where we found that the cytoplasm-to-membrane translocation-based optogenetic systems outperform the membrane-anchored dimerization systems in activating the receptor tyrosine kinase signaling in live Xenopus embryos. Here, we determine if this engineering strategy can be generalized to other signaling pathways involving membrane-bound receptors. As a proof of concept, we demonstrate that the cytoplasm-to-membrane translocation of the low-density lipoprotein receptor-related protein-6 (LRP6), a membrane-bound coreceptor for the canonical Wnt pathway, triggers Wnt activity. Optogenetic activation of LRP6 leads to axis duplication in developing Xenopus embryos, indicating that the cytoplasm-to-membrane translocation of the membrane-bound receptor could be a generalizable strategy for the construction of optogenetic systems.

Role of TrkA signaling during tadpole tail regeneration and early embryonic development in Xenopus laevis.

Neurotrophic signaling regulates neural cell behaviors in development and physiology, although its role in regeneration has not been fully investigated. Here, we examined the role of neurotrophic signaling in Xenopus laevis tadpole tail regeneration. After the tadpole tails were amputated, the expression of neurotrophin ligand family genes, especially ngf and bdnf, was up-regulated as regeneration proceeded. Moreover, notochordal expression of the NGF receptor gene TrkA, but not that of other neurotrophin receptor genes TrkB and TrkC, became prominent in the regeneration bud, a structure arising from the tail stump after tail amputation. The regenerated tail length was significantly shortened by the pan-Trk inhibitor K252a or the TrkA inhibitor GW-441756, but not by the TrkB inhibitor ANA-12, suggesting that TrkA signaling is involved in elongation of regenerating tails. Furthermore, during Xenopus laevis embryonic development, TrkA expression was detected in the dorsal mesoderm at the gastrula stage and in the notochord at the neurula stage, and its knockdown led to gastrulation defects with subsequent shortening of the body axis length. These results suggest that Xenopus laevis TrkA signaling, which can act in the mesoderm/notochord, plays a key role in body axis elongation during embryogenesis as well as tail elongation during tadpole regeneration.

Molecular characterization of wdr68 gene in embryonic development of Xenopus laevis

WDR68, also known as DCAF7, is a WD40 repeated domain protein highly conserved in eukaryotic organisms in both plants and animals. This protein participates in numerous cellular processes and exerts its function through interaction with other proteins. In the present work, we isolated, sequenced and characterized cDNA corresponding to the wdr68 gene in embryos of the amphibian Xenopus laevis. Syntenic analysis revealed high conservation of the genomic region containing the WDR68 locus in amniotes. Nevertheless, in fishes and amphibians, we observed that the tandem genes surrounding wdr68 undergoes certain rearrangements with respect to the organization found in amniotes. We also defined the temporal and spatial expression pattern of the wdr68 gene in the development of Xenopus laevis through whole mount in situ hybridization and RT-PCR techniques. We observed that wdr68 is ubiquitously expressed during early embryonic development but, during the neurula stage, it undergoes a strong expression in the neural tube and in the migratory cephalic streams of the neural crest. At the tailbud stages, it is strongly expressed in the cephalic region, particularly in otic and optic vesicles, in addition to branchial arches. In contrast, wdr68 transcripts are localized in the somitic mesoderm in the trunk. The expression area that includes the migratory neural crest of the head and the branchial arches suggest that this gene would be involved in jaws formation, probably through a hierarchical relationship with the component genes of the endothelin-1/endothelin receptor type A cell signaling pathway.

A multichannel computer-driven system to raise aquatic embryos under selectable hypoxic conditions.

The formation of a functional cardiovascular system is an essential step in the early vertebrate embryo. Nevertheless, the effect of hypoxia on the developmental program of organisms was studied rarely. In particular, this holds true for vertebrate embryos that depend on a functional placenta for proper development and had not been studied in this respect due to the obvious limitation. We established a protocol to culture aquatic embryos, which enabled us to culture a high number of Xenopus embryos until tadpole stage under defined hypoxic conditions in four hypoxia chambers simultaneously, employing a computerized system. In general, our results show that hypoxia results in delayed development and, in particular, we could show that oxygen availability was most crucial during gastrulation and organogenesis (early tailbud) phases during embryonic development of Xenopus laevis.

Specific inhibition of GPCR-independent G protein signaling by a rationally engineered protein

Activation of heterotrimeric G proteins by cytoplasmic nonreceptor proteins is an alternative to the classical mechanism via G protein-coupled receptors (GPCRs). A subset of nonreceptor G protein activators is characterized by a conserved sequence named the Gα-binding and activating (GBA) motif, which confers guanine nucleotide exchange factor (GEF) activity in vitro and promotes G protein-dependent signaling in cells. GBA proteins have important roles in physiology and disease but remain greatly understudied. This is due, in part, to the lack of efficient tools that specifically disrupt GBA motif function in the context of the large multifunctional proteins in which they are embedded. This hindrance to the study of alternative mechanisms of G protein activation contrasts with the wealth of convenient chemical and genetic tools to manipulate GPCR-dependent activation. Here, we describe the rational design and implementation of a genetically encoded protein that specifically inhibits GBA motifs: GBA inhibitor (GBAi). GBAi was engineered by introducing modifications in Gαi that preclude coupling to every known major binding partner [GPCRs, Gβγ, effectors, guanine nucleotide dissociation inhibitors (GDIs), GTPase-activating proteins (GAPs), or the chaperone/GEF Ric-8A], while favoring high-affinity binding to all known GBA motifs. We demonstrate that GBAi does not interfere with canonical GPCR-G protein signaling but blocks GBA-dependent signaling in cancer cells. Furthermore, by implementing GBAi in vivo, we show that GBA-dependent signaling modulates phenotypes during Xenopus laevis embryonic development. In summary, GBAi is a selective, efficient, and convenient tool to dissect the biological processes controlled by a GPCR-independent mechanism of G protein activation mediated by cytoplasmic factors.

Retinoid-like compounds produced by phytoplankton affect embryonic development of Xenopus laevis

Teratogenic effects, which were remarkably similar to those induced by retinoic acids, have been seen in wild frogs indicating possible source of retinoids in the environment. Recent studies indicate that some cyanobacterial species can contain teratogenic retinoic acids (RAs) and their analogues. Retinoids are known to regulate important processes such as differentiation, development, and embryogenesis. The study investigated the effects of exudates (extracellular compounds) of two cyanobacteria species with retinoic-like activity and one algae species on embryonic development of amphibians. The retinoid-like activity determined by in vitro reporter gene assay reached 528ng retinoid equivalents (REQ)/L and 1000ng REQ/L in exudates of Cylindrospermopsis raciborskii and Microcystis aeruginosa, respectively, while algal exudates showed no detectable activity. Total mean of retinoid-like copounds into exudate was 35.6ng ATRA/mil.cells for M.aeruginosa and 6.71ng ATRA/mil.cells for C.raciborskii, respectively. Toxicity tests with amphibian embryos up to 96h of development were carried out according to the standard guide for the Frog Embryo Teratogenesis Assay Xenopus. Lowest observed effect concentrations (LOEC) of malformations (2.5–2.6µg/L REQ) were two times lower than LOEC for ATRA (5µg/L). The exudates of both cyanobacteria were indeed provoking diverse teratogenic effects (e.g. tail, gut and eyes deformation) and interference with growth in frogs embryos, while such effects were not observed for the algae. Xenopus embryos were also exposed to all-trans retinoic acid (ATRA) in concentration range (1−40µg/L) equivalent to the REQs detected in cyanobacterial exudates. ATRA (10µg/L) caused similar teratogenic phenotypes at corresponding REQs as cyanobacterial exudates. The study confirms the ability of some species of cyanobacteria to produce retinoids naturally and excrete them directly into the environment at concentrations which might have adverse influence on the development of amphibians.

Prepatterning and patterning of the thalamus along embryonic development of Xenopus laevis.

Previous developmental studies of the thalamus (alar part of the diencephalic prosomere p2) have defined the molecular basis for the acquisition of the thalamic competence (preparttening), the subsequent formation of the secondary organizer in the zona limitans intrathalamica, and the early specification of two anteroposterior domains (rostral and caudal progenitor domains) in response to inducing activities and that are shared in birds and mammals. In the present study we have analyzed the embryonic development of the thalamus in the anuran Xenopus laevis to determine conserved or specific features in the amphibian diencephalon. From early embryonic stages to the beginning of the larval period, the expression patterns of 22 markers were analyzed by means of combined In situ hybridization (ISH) and immunohistochemical techniques. The early genoarchitecture observed in the diencephalon allowed us to discern the boundaries of the thalamus with the prethalamus, pretectum, and epithalamus. Common molecular features were observed in the thalamic prepatterning among vertebrates in which Wnt3a, Fez, Pax6 and Xiro1 expression were of particular importance in Xenopus. The formation of the zona limitans intrathalamica was observed, as in other vertebrates, by the progressive expression of Shh. The largely conserved expressions of Nkx2.2 in the rostral thalamic domain vs. Gbx2 and Ngn2 (among others) in the caudal domain strongly suggest the role of Shh as morphogen in the amphibian thalamus. All these data showed that the molecular characteristics observed during preparttening and patterning in the thalamus of the anuran Xenopus (anamniote) share many features with those described during thalamic development in amniotes (common patterns in tetrapods) but also with zebrafish, strengthening the idea of a basic organization of this diencephalic region across vertebrates.

The effect of 900 and 1800 MHz GSM-like radiofrequency irradiation and nicotine sulfate administration on the embryonic development of Xenopus laevis

The aim of this study was to investigate the effects of GSM-like radiofrequency electromagnetic radiation (RF EMR) and nicotine sulfate (NS) exposure on Xenopus embryonic development.The developmental effects of GSM-like RF-EMR (900–1800MHz, at a SAR value of 1W/kg and NS on Xenopus laevis embryos were investigated). Following the application of radiofrequency radiation and/or NS administration, the embryos were closely examined in order to determine their possible teratogenic effects. Xenopus frogs obtained from the Department of Physiology of the Cukurova University, in accordance described by the Standard Guide of the American Society for Testing and Materials (ASTM). Following the exposure of Xenopus embryos to RF-EMR at 900 and 1800MHz (1.0W/kg) for 4, 6 and 8h; the whole body specific energy absorption rate (SAR) of the embryos was calculated. With the exception of irradiation at 1800MHz no dramatic developmental anomalies were observed in the Xenopus embryos in association with RF-EMR applications. Combined RF-EMR and NS applications resulted in dramatic abnormalities and death among the Xenopus embryos. The study results indicated that GSM-like RF-EMR (e.g. radiation from cell phones) was not as harmful to Xenopus embryos as might have been expected. However, the combined effects of GSM-like RF-EMR and NS on Xenopus embryos were more severe than the effect of RF-EMR or NS alone. In conclusion, the study results appear to suggest that the combined use of nicotine and cell phones might result in more pronounced detrimental effects on the health of smokers.

Changes in acetyl CoA levels during the early embryonic development of Xenopus laevis.

Coenzyme A (CoA) is a ubiquitous and fundamental intracellular cofactor. CoA acts as a carrier of metabolically important carboxylic acids in the form of CoA thioesters and is an obligatory component of a multitude of catabolic and anabolic reactions. Acetyl CoA is a CoA thioester derived from catabolism of all major carbon fuels. This metabolite is at a metabolic crossroads, either being further metabolised as an energy source or used as a building block for biosynthesis of lipids and cholesterol. In addition, acetyl CoA serves as the acetyl donor in protein acetylation reactions, linking metabolism to protein post-translational modifications. Recent studies in yeast and cultured mammalian cells have suggested that the intracellular level of acetyl CoA may play a role in the regulation of cell growth, proliferation and apoptosis, by affecting protein acetylation reactions. Yet, how the levels of this metabolite change in vivo during the development of a vertebrate is not known. We measured levels of acetyl CoA, free CoA and total short chain CoA esters during the early embryonic development of Xenopus laevis using HPLC. Acetyl CoA and total short chain CoA esters start to increase around midblastula transition (MBT) and continue to increase through stages of gastrulation, neurulation and early organogenesis. Pre-MBT embryos contain more free CoA relative to acetyl CoA but there is a shift in the ratio of acetyl CoA to CoA after MBT, suggesting a metabolic transition that results in net accumulation of acetyl CoA. At the whole-embryo level, there is an apparent correlation between the levels of acetyl CoA and levels of acetylation of a number of proteins including histones H3 and H2B. This suggests the level of acetyl CoA may be a factor, which determines the degree of acetylation of these proteins, hence may play a role in the regulation of embryogenesis.

Evidence and uptake routes for Zinc oxide nanoparticles through the gastrointestinal barrier in Xenopus laevis

The developmental toxicity of nanostructured materials, as well as their impact on the biological barriers, represents a crucial aspect to be assessed in a nanosafety policy framework. Nanosized metal oxides have been demonstrated to affect Xenopus laevis embryonic development, with nZnO specifically targeting the digestive system. To study the mechanisms of the nZnO-induced intestinal lesions, we tested two different nominally sized ZnO nanoparticles (NPs) at effective concentrations. Advanced microscopy techniques and molecular marker analyses were applied in order to describe the NP-epithelial cell interactions and the mechanisms driving NP toxicity and translocation through the intestinal barrier. We attributed the toxicity to NP-induced cell oxidative damage, the small-sized NPs being the more effective. This outcome is sustained by a marked increase in anti-oxidant genes' expression and high lipid peroxidation level in the enterocytes, where disarrangement of the cytoskeleton and cell junctions' integrity were evidenced. These events led to diffuse necrotic changes in the intestinal barrier, and trans- and paracellular NP permeation through the mucosa. The uptake routes, leading NPs to cross the intestinal barrier and reach secondary target tissues, have been documented. nZnOs embryotoxicity was confirmed to be crucially mediated by the NPs' reactivity rather than their dissolved ions. The ZnO NPs' ability to overwhelm the intestinal barrier must be taken into high consideration for a future design of safer ZnO NPs.

Role of the hypoxia response pathway in lens formation during embryonic development of Xenopus laevis

The RING finger ubiquitin ligase seven in absentia homolog 2 (Siah2) was identified in the R7 photoreceptor cells of Drosophila melanogaster, and it regulates the stability of prolyl hydroxylase domains (PHDs), with a concomitant effect on HIF-1α availability in the hypoxia response pathway. We previously reported that the hypoxia response pathway contributes to eye development during the embryonic development of Xenopus laevis. In this paper, the role of Siah2-mediated hypoxia response pathway in eye development of X. laevis embryos was further characterized. Xenopus Siah2 (xSiah2) mRNA was detected in lens tissue and xSiah2 overexpression caused a thickened lens placode, leading to loss of the optic lens. In embryos overexpressing xSiah2, lens marker gene transcription was reduced, suggesting that xSiah2 contributes to lens formation. xSiah2 overexpression decreased Xenopus PHD accumulation and increased Xenopus HIF-1α (xHIF-1α) accumulation. xHIF-1α degeneration with resveratrol restored the optical abnormality caused by xSiah2 overexpression, suggesting that the xSiah2-mediated hypoxia response pathway contributes to lens formation. Moreover, xSiah2 overexpression decreased endothelial–mesenchymal transition (EMT)-related Notch signaling-responsive genes transcription during the invasion of the lens placode. Our results suggest that the hypoxia response pathway plays an important role in the regulation of the EMT via the Notch signaling pathway during lens formation.

Expression of the tetraspanin family members Tspan3, Tspan4, Tspan5 and Tspan7 during Xenopus laevis embryonic development

Tetraspanins comprise a large family of integral membrane proteins involved in the regulation of cell adhesion, migration and fusion. In humans it consists of 33 members divided in four subfamilies. Here, we examined the spatial and temporal gene expression of four related tetraspanins during the embryonic development of Xenopus laevis by quantitative real-time PCR and in situ hybridization: Tspan3 (encoded by the gene Tm4sf8 gene) Tspan4 (encoded by the gene Tm4sf7), Tspan5 (encoded by the gene Tm4sf9) and Tspan7 (encoded by the gene Tm4sf2). These genes appeared first in the vertebrates during the evolution and are conserved across different species. In humans, they were associated with several diseases such as sclerosis, mental retardation and cancer; however their physiological role remained unclear. This work provides a comprehensive comparative analysis of the expression of these tetraspanins during the development of X. laevis. The more closely related tetraspanins Tspan3, Tspan4 and Tspan7 exhibited very similar spatial expression patterns, albeit differing in their temporal occurrence. The corresponding transcripts were found in the dorsal animal ectoderm at blastula stage. At early tailbud stages (stage 26) the genes were expressed in the migrating cranial neural crest located in the somites, developing eye, brain, and in otic vesicles. In contrast, Tspan5 appeared first at later stages of development and was detected prominently in the notochord. These data support close relatedness of Tspan3, Tspan4 and Tspan7. The expression of these tetraspanins in the cells with a high migratory potential, e.g. neural crest cells, suggests their role in the regulation of migration processes, characteristic for tetraspanin family members, during development. Similarity of the expression profiles might indicate at least partial functional redundancy, which is in concordance with earlier findings of tissue-limited or absent phenotypes in the knock-down studies of tetraspanins family members performed.

The embryonic development of Xenopus laevis under a low frequency electric field

The aim of this study was to determine the effects of a low frequency electric field on the early embryonic development of frogs. The embryos of African clawed toads, Xenopus laevis, were exposed to a 20-μA electric current during the cleavage stages. The developmental processes of embryos during and after electric field exposure were monitored for teratogenic effects. All the embryos continuously exposed to the electric field died without undergoing any developmental processes. However, when the embryos were exposed to the electric field for 20-min periods (four times/over 2 d), the embryos developed into both normal tadpoles (70 %) and malformed tadpoles with light edema, reduced pigmentation, or axial anomalies, such as crooked tails. After exposure, the control embryos were at development stage 35.5 (2 d 2 h), while the normal embryos of the assay group were at developmental stage 41(3 d 4 h). There was a 1 d 2 h difference between the two developmental stages, revealing the importance of that time period for embryogenesis. In conclusion, the effects of electric current on Xenopus embryos are dependent on the initial developmental stage and the duration of exposure.

Altered development ofXenopusembryos in a hypogeomagnetic field

The hypogeomagnetic field (HGMF; magnetic fields <200 nT) is one of the fundamental environmental factors of space. However, the effect of HGMF exposure on living systems remains unclear. In this article, we examine the biological effects of HGMF on the embryonic development of Xenopus laevis (African clawed frog). A decrease in horizontal third cleavage furrows and abnormal morphogenesis were observed in Xenopus embryos growing in the HGMF. HGMF exposure at the two-cell stage, but no later than the four-cell stage, is enough to alter the third cleavage geometry pattern. Immunofluorescent staining for α-tubulin showed reorientation of the spindle of four-cell stage blastomeres. These results indicate that a brief (2-h) exposure to HGMF is sufficient to interfere with the development of Xenopus embryos at cleavage stages. Also, the mitotic spindle could be an early sensor to the deprivation of the geomagnetic field, which provides a clue to the molecular mechanism underlying the morphological and other changes observed in the developing and/or developed embryos.