Distant wave interaction in the early embryogenesis of the loach Misgurnus fossilis L

Groups of loach (Misgurnus fossilis L.) embryos of different ages were kept in different quartz cuvettes for 20-24 h so that only optic contact between the groups was possible. Subsequent observations showed that parameters of their development deviated from those in the control groups. Wave-mediated biocorrection proved to have both positive and negative effects, depending on the developmental stages of the interacting groups. Changes in spectral characteristics and polarization of biological radiation affected the results of the experiments. Various developmental abnormalities caused by distant wave-mediated interactions of embryos and specific to each combination of developmental stages and conditions of optic communication are described.     

Ultraweak emissions of developing <Emphasis Type="Italic">Xenopus laevis</Emphasis> eggs and embryos

We measured ultraweak emissions of the Xenopus laevis eggs and embryos during normal development and under the influence of stress factors in a spectral range of 250 to 800 nm using a photomultiplier. The registered emissions were analyzed by several basic characteristics: mean intensity, histograms, kurtosis, linear trends, and Fourier spectra. We followed relationships between these parameters and developmental stage, as well as the number of individuals in optic contact with each other. The ultraweak emissions did not differ from the background at all developmental stages according to the mean intensity. But Fourier analysis revealed the reliable presence of a number of spectral lines of ultraweak emission, predominantly in the range of 10^?2–50 Hz, in the embryos at developmental stages 2 to 11. The intensity of ultraweak emissions reliably decreased within the first 10 min after egg activation and fertilization, as well as in the case of optic interaction between groups of embryos. Sharp cooling, increase in osmotic medium pressure, and transfer in a Ca²⁺ and Mg²⁺-free medium induced a short term (～1–5 min) increase in the mean intensity of ultraweak emission. We studied specific features of ultraweak emissions from different parts of the embryo. The intensity of emission from the animal part of early blastula exceeded those from the vegetal area and entire embryo. Separated fragments of the lateral ectoderm at the neurula stage had higher mean intensities of ultraweak emission than intact embryos at the same developmental stages.     

Ultraweak emissions of the developing Xenopus laevis eggs and embryos

We measured ultraweak emissions of the Xenopus laevis eggs and embryos during normal development and under the influence of stress factors in a spectral range of 250 to 800 nm using a photomultiplier. The registered emissions were analyzed by several basic characteristics: mean intensity, histograms, kurtosis, linear trends, and Fourier spectra. We followed relationships between these parameters and developmental stage, as well as the number of individuals in optic contact with each other. The ultraweak emissions did not differ from the background at all developmental stages according to the mean intensity. But Fourier analysis revealed the reliable presence of a number of spectral lines of ultraweak emission, predominantly in the ranges of 10-20 and 30-40 Hz, in the embryos at developmental stages 2 to 11. The intensity of ultraweak emissions reliably decreased within the first 10 min after egg activation and fertilization, as well as in the case of optic interaction between groups of embryos. Sharp cooling, increase in osmotic medium pressure, and transfer in a Ca(2+)- and Mg(2+)-free medium induced a short term (approximately 1-5 min) increase in the mean intensity of ultraweak emission. We studied specific features of ultraweak emissions from different parts of the embryo. The intensity of emission from the animal part of early blastula exceeded those from the vegetal area and entire embryo. Separated fragments of the lateral ectoderm at the neurula stage had higher mean intensities of ultraweak emission than intact embryos at the same developmental stages.     

Statistical and frequency-amplitude characteristics of ultra-weak emissions of loach eggs and embryos under normal conditions and on their optic interactions. II. Changes in characteristics of ultra-weak emissions upon optic interaction of groups of embryos of different ages.

We compared the characteristics of ultraweak emissions from groups of loach embryos of different ages in the presence or absence of optic interaction. The percentage of zero values of emission gradually increased during the first hour of optic interaction. The number and height of rare big pulses estimated by the value of kurtosis increased in parallel. In addition, the correlation between the Fourier spectra of optically interacting samples decreased at a higher rate than in the absence of optical contact. Just after the 1-hour optic interaction was terminated, the number of high pulses decreased in a younger interacting group and increased in the older one and the farther away the partner groups were in developmental stages, the more pronounced these differences were. Measurements of the Fourier spectra after long-term (12-22-hour) optic interactions have shown that an "exchange" of autocorrelation characteristics of the spectra took place among the samples: the sums of autocorrelation coefficients were inverted in the vast majority of cases, often with an "overshoot" or, at least, were smoothed over with reference to the control samples. We conclude that the previously described effects of optic interactions between groups of loach embryos of different ages could be due to changes in the frequency spectra of their ultraweak emissions.     

Studies on the embryogenesis of Aulocara elliotti (Thomas) (Orthoptera, Acrididae). I. External morphogenesis

The growth and morphogenesis of embryos of Aulocara elliotti (Thomas) at constant temperature are described in terms of 27 discrete morphological stages with four stages designating blastokinesis. The developmental variability of two series of embryos reared from a single wild population in two different years is compared. A bibliography to studies on other embryos of the Acrididae is included.     

Methoprene Photolytic Compounds Disrupt Zebrafish Development,Producing Phenocopies of Mutants in the Sonic Hedgehog Signaling Pathway

Environmental chemicals have been proposed to impact endocrine or retinoid pathways, causing developmental abnormalities in humans and other vertebrates. Presented evidence shows that exposure of zebrafish embryos to sunlight-induced photolytic products of the pesticide methoprene results in developmental defects in the head, heart, pectoral fins, and somites, and in spinal motor and optic nerve axons. Exposed embryos are phenocopies of zebrafish you-type mutants and, as in the mutant sonic-you, show underexpression of the signaling protein sonic hedgehog. Reduced expression of sonic hedgehog is also displayed in embryos treated with the retinoic acid synthesis inhibitor citral. This study identifies citral-related compounds as embryonic signaling disruptors of potential environmental concern.     

Analysis of the development of the lizard, Calotes versicolor. I. A series of normal stages in the embryonic development

It has been shown that the external parameters of eggs of the garden lizard, Calotes versicolor, are not suitable for assessing the exact developmental stages of embryos. In order to make use of this lizard's embryos for experimental work, a series of developmental stages has been characterized, using various morphological features.     

Comparative study of reserve lipid accumulation during somatic and zygotic <Emphasis Type="Italic">Acca sellowiana</Emphasis> (O. Berg.) Burret embryogenesis

Somatic embryogenesis is an in vitro morphogenetic route in which isolated cells or a small group of somatic cells give rise to bipolar structures resembling zygotic embryos. Lipids, carbohydrates, and proteins are major compounds in plant and animal metabolism. Comparative analysis along different developmental stages of Acca sellowiana (Myrtaceae) zygotic and somatic embryos, revealed a progressive increase in levels of total lipids. A high degree of similarity could be found in the total lipids composition between A. sellowiana somatic and zygotic embryos. High lipid levels were found in zygotic embryos in the torpedo and cotyledonary stages, and these levels increased according to the progression in the developmental stages. Somatic embryos obtained through direct embryogenesis route showed higher levels of lipids than in indirect somatic embryogenesis. The compounds most frequently were linoleic acid (C18:2), palmitic (C16:0) and oleic (C18:1). These results indicate a high similarity degree of accumulation of total lipids, regardless of zygotic or somatic embryogenesis.     

Statistical and Frequency-Amplitude Characteristics of Ultraweak Emissions of the Loach Eggs and Embryos under the Normal Conditions and upon Their Optic Interactions. 2. Changes in Characteristics of Ultraweak Emissions upon Optic Interaction of Groups of Embryos of Different Ages

We compared the characteristics of ultraweak emissions from groups of loach embryos of different ages in the presence or absence of optic interaction. The percentage of zero values of emission gradually increased during the first hour of optic interaction. The number and height of rare big pulses estimated by the value of kurtosis increased in parallel. In addition, the correlation between the Fourier spectra of optically interacting samples decreased at a higher rate than in the absence of optical contact. Just after the 1-hour optic interaction was terminated, the number of high pulses decreased in a younger interacting group and increased in the older one and the farther away the partner groups were in developmental stages, the more pronounced these differences were. Measurements of the Fourier spectra after long-term (12–22-hour) optic interactions have shown that an exchange of autocorrelation characteristics of the spectra took place among the samples: the sums of autocorrelation coefficients were inverted in the vast majority of cases, often with an overshoot or, at least, were smoothed over with reference to the control samples. We conclude that the previously described effects of optic interactions between groups of loach embryos of different ages could be due to changes in the frequency spectra of their ultraweak emissions.     

Differential proteomic analysis of developmental stages of <Emphasis Type="Italic">Acca sellowiana</Emphasis> somatic embryos

Feijoa (Acca sellowiana, Myrtaceae), a native fruit species from southern Brazil and northern Uruguay, is considered to constitute a reference system for somatic embryogenesis in woody dicots. This in vitro regenerative pathway is an efficient micropropagation method, and a suitable model system for studies in plant developmental physiology. This study attempts to detect and identify proteins that are expressed during the different developmental stages of somatic embryos of A. sellowiana. Using high resolution two-dimensional polyacrylamide gel electrophoresis (2-DE), a high degree of similarity between protein profiles of the assayed somatic embryos was observed. Of the 74 different protein spots extracted for analysis, 60 were identified by means of 2-DE/MALDI-TOF/MS. Twelve proteins were expressed in all the assayed stages. Ten proteins were expressed in the initial stages and 22 proteins were expressed in the mature developmental stages of somatic embryos. Only one protein was expressed exclusively in the torpedo stage, whereas four were expressed in the pre-cotyledonary, and none in the cotyledonary stage. The proteins identified were involved in the synthesis of phenylalanine ammonia-lyase, a conspicuous polyphenol present in the induction of feijoa embryogenic cultures. The presence of essential proteins of nitrogen metabolism, such as the cytosolic glutamine synthetase protein, was also observed. The physiological implications of these findings are discussed.     

Improved genetic identification of acipenseriform embryos with application to the endangered pallid sturgeon Scaphirhynchus albus

We produced pallid sturgeon Scaphirhynchus albus embryos at five pre-hatch developmental stages and isolated and quantified genomic DNA from four of the stages using four commercial DNA isolation kits. Genomic DNA prepared using the kit that produced the largest yields and concentrations were used for microsatellite DNA analyses of 10–20 embryos at each of the five developmental stages. We attempted to genotype the hatchery-produced embryos at 19 microsatellite loci and confirmed reliable genotyping by comparing the microsatellite genotypes to those of known parents. Embryos at stages 5 and 8 did not produce reliable genotyping while those at stages 14, 24 and 33 did. We used the same DNA isolation method on 262 wild-caught acipenseriform embryos collected from the lower Yellowstone River. A total of 200 of the wild embryos were successfully identified to stages 8 to 34 and the rest could not be staged. Using a combination of single nucleotide polymorphism and microsatellite markers, 249 of the wild-caught embryos were genetically identified as paddlefish Polyodon spathula, five were identified as shovelnose sturgeon Scaphirhynchus platorynchus and eight failed to amplify. None were identified as pallid sturgeon. This study demonstrates that early-stage wild-spawned acipenseriform embryos can be genetically identified less than 24 h post-spawn. This methodology will be useful for recovery efforts for endangered pallid sturgeon and can be applied to other acipenseriform species.     

Visuospatial information in the retinotectal system of xenopus before correct image formation by the developing eye

The retinotectal pathway of Xenopus laevis is a well-established experimental model for studying activity-dependent processes during visual system development. Such processes can be guided by stimulus-evoked activity patterns, which depend on the refractive characteristics of the eye. Previous work has shown that many animals are hyperopic at early developmental stages due to immature refractive properties. Whether this is also the case for Xenopus laevis is unknown. Here, we measure the focal length of the lens and the size of the eye of embryos at different stages and find that Xenopus laevis exhibits a similar shift from hyperopia to emmetropia. At early stages, immediately after innervation of the tectum by the optic nerve, Xenopus embryos are hyperopic. Soon afterwards the focal length of the lens decreases and the eye converges to a state of emmetropia. Despite being hyperopic we find that some visuospatial information is available to the young circuit. Calculations based on the optical properties of the eye show that even when the animals are hyperopic the blurred retinal image provides a crude spatial resolution. Furthermore, using whole-cell recordings in the optic tectum combined with visual stimulation through the intact eye, we show that tectal neurons in hyperopic embryos have spatially restricted glutamatergic receptive fields. Our data demonstrate that Xenopus laevis eyes undergo a process of developmental emmetropization, and suggest that despite an initial stage of suboptimal image formation there is potentially enough information to guide activity-dependent refinements of the retinotectal pathway from the onset of vision.     

Somatic embryony patterns and plant regeneration inIpomoea batatas poir

Summary Patterns of somatic embryo development from callus were elucidated inIpomoea batatas Poir. (sweet potato). Embryos at three stages of development were able to grow into plants. They included embryos arrested at the late torpedo and cotyledonary stages, and an arrested torpedo embryo that resulted from precocious hypocotyl expansion. Early torpedo-stage embryos rooted on reculture but did not form shoots. A diversity of other embryo stages were also produced which upon reculture formed only adventitious embryos and roots. Morphologic variants with similar growth potential were noted among embryos at similar stages of development. Plasticity in somatic embryo developmental patterns led to different forms that were mature enough to produce plants. This work has been funded in part by an IFAS/Gas Research Institute cooperative grant. Florida Agricultural Experiment Station Journal Series No. 8789.     

Effect of embryo developmental stage and culture conditions on number and quality of ovine in vitro produced blastocysts

This study evaluated the final output and quality of in vitro produced blastocysts derived from in vivo recovered sheep embryos cultured at various early developmental stages to blastocyst. A total of 270 embryos were recovered from the oviduct, at different days of the early luteal phase, and were classified into three different developmental stages: 2- to 4-cell (n = 93); 5- to 8-cell (n = 92) and 9- to 12-cell (n = 85). The effect of culture conditions was studied, at the same time, by randomly allocating the embryos to one of four groups: three groups of culture with fresh oviduct monolayers (2, 4 and 5 days old) and a fourth group with 2-day monolayers derived from frozen-thawed oviduct cells. Two control groups were established: first, embryos cultured in semi-defined medium (n = 29) and, second, blastocysts obtained in vivo and cryopreserved (n = 43). Influence on blastocyst yield of embryo developmental stage at the start of culture was statistically significant (p < 0.001). Two- to four-cell embryos showed a significantly lower developmental rate (67.7%) than the 5- to 8-cell (83.6%; p < 0.001) and 9- to 12-cell groups (90.5%; p < 0.0001) and lower quality in terms of blastocyst cryotolerance (56.0 vs. 83.7%; p < 0.005). There were no detected effects relating to the age or handling of the monolayer on the embryo developmental rate, but the day of blastocyst appearance was different between embryos cultured on monolayers derived from fresh or frozen-thawed cells (p < 0.0001); the main influence was on the group of 9- to 12-cell embryos (p < 0.0001). Current results confirm the temporal sensitivities of sheep embryos to in vitro culture, regardless of the culture conditions.     

ULTRASTRUCTURE OF ARRESTED EMBRYOS FROM LETHAL MUTANTS OF ARABIDOPSIS THALIANA

The purpose of this study was to examine the extent of cellular differentiation in arrested embryos from lethal mutants of Arabidopsis thaliana. The question to be addressed was whether arrested embryos in heterozygous siliques resembled mature wild-type embryos at the cellular level. Protein bodies were chosen as developmental markers because they appear only during the final stages of embryogenesis. Both the hypocotyl and cotyledons of wild-type embryos contained protein bodies that became filled with storage protein during the cotyledonary stages of development. Some mutant embryos (emb30) contained normal protein bodies and resembled mature wild-type embryos at the cellular level. Other mutant embryos (emb22) contained only immature protein bodies and were therefore blocked in both morphogenesis and cellular differentiation. The formation of protein bodies in emb31 was normal in the hypocotyl but delayed in the cotyledons. In this case the mutant gene appears to disrupt the timing of both morphogenesis and differentiation. This ultrastructural view of arrested embryos has provided additional information on the nature of developmental arrest that should facilitate the classification of embryonic lethals and the identification of mutants with defects in developmental rather than housekeeping functions.     

Studies on the embryogenesis of Aulocara elliotti (Thomas) (Orthoptera, Acrididae). II. Developmental variability and the effects of maternal age and environment

The stages of diapause embryos of A. elliotti reared in the laboratory were described, and the lengths of embryos of stage 19 were compared with a second group of diapause embryos of advanced ages. Collections of diapause embryos of the same species were made from wild populations during the autumn and winter of two consecutive years. The stages of embryos present in these samples were found to vary widely. The mean lengths of embryos in stage 19 from different populations were compared and found to differ with statistical significance. The numbers of retarded embryos in different populations were found to vary significantly between populations of diapause embryos. In two populations, mean lengths of embryos of stage 19 were found to differ from one year to the next. Post-diapause embryos were collected from wild populations in the spring of two consecutive years and their stages described. No significant difference was found in the incidence of retardation in post-diapause samples. Results of these studies and those of other authors indicate that there may be wide developmental variation between populations of acridid embryos as well as in one population in two different years. Rearing adult A. elliotti in varying numbers under two different environmental conditions was found to produce significant effects upon the rate of development and variability of their embryonic progeny. Embryos laid by young females were found to be more variable, in general, and to develop more slowly than those laid by older females.     

Efficient embryonic culture method for the Japanese striped snake,Elaphe quadrivirgata,and its early developmental stages

The morphogenesis of snake embryos is an elusive yet fascinating research target for developmental biologists. However, few data exist on development of early snake embryo due to limited availability of pregnant snakes, and the need to harvest early stage embryos directly from pregnant snakes before oviposition without knowing the date of fertilization. We established an ex vivo culture method for early snake embryos using the Japanese striped snake, Elaphe quadrivirgata. This method, which we named “sausage‐style (SS) culture”, allows us to harvest snake embryos at specific stages for each experiment. Using this SS culture system, we calculated somite formation rate at early stages before oviposition. The average somite formation rate between 6/7 and 12/13 somite stages was 145.9 min, between 60/70 and 80/91 somite stages 42.4 min, and between 113–115 and 126/127 somite stages 71 min. Thus, somite formation rate that we observed during early snake embryogenesis was changed over time. We also describe a developmental staging series for E. quadrivirgata. This is the first report of a developmental series of early snake embryogenesis prior to oviposition by full‐color images with high‐resolution. We propose that the SS culture system is an easy method for treating early snake embryos ex vivo.     

Somatic embryogenesis and plantlet development in Pinus sylvestris and Pinus pinaster on medium with and without growth regulators

To initiate somatic embryogenesis in Pinus sylvestris and Pinus pinaster, immature seeds were collected from June to August and the developmental stage of the zygotic embryos was determined. Four developmental stages were distinguished and the response of the zygotic embryos at each of the four developmental stages was compared intra- and inter-species. For this study, modified Litvay's medium (LM), with or without growth regulators, was chosen. Somatic embryogenesis was initiated and maintained on both media but the two species displayed different propensities. In P. sylvestris, the highest initiation frequency was obtained with intact megagametophytes containing embryos at the four-cell stage to the stage of cleavage polyembryony (up to 22 and 9%, respectively). The culture medium had no significant effect on the initiation and proliferation of embryogenic cultures. In P. pinaster, however, the best response occurred from excised zygotic embryos at the stage prior to elongation of cotyledon primordia (up to 40% explants responded), on medium with growth regulators. Another characteristic distinguishing the two species in culture was that in some embryogenic cell lines of P. sylvestris, somatic embryos matured spontaneously when initiated and maintained on medium without growth regulators. Some of these embryos developed into plantlets on the same medium at the frequency of 40%. Therefore, in P. sylvestris all the stages of somatic embryogenesis were achieved on the medium without growth regulators. However, in both species, maturation of a large number of somatic embryos was greatly improved on medium containing high concentration of gellan gum (Gelrite 10 g l^?1) and abscisic acid (60 μM). Cotyledonary somatic embryos subsequently germinated (72 and 80% for P. sylvestris and P. pinaster, respectively) and developed into plantlets (48 and 29%, for P. sylvestris and P. pinaster, respectively). This represents a significant improvement in plantlet recovery from somatic embryos of both species.     

Lectin-induced blockage of developmental processes in preimplantation mouse embryos in vitro

This study attempts to assess the developmental importance of cell surface glycoconjugates of preimplantation mouse embryos. This was done by incubating early embryos in various lectins and analyzing subsequent development. If specific cell surface glycoconjugates (lectin receptors) are linked to specific developmental processes, such as cell division, compaction, and blastocyst formation, then different lectins should block these different developmental processes. The results show that wheat-germ agglutinin (WGA; N-acetyl-D-glucosamine-specific) at 50 μg/ml prevents the cell division of four-cell embryos. However, this effect of WGA occurs only in embryos with intact zonae pellucidae. Concanavalin A (Con A; α-D-glucose and α-D-mannose-specific) treatment, 20 μg/ml, of four-cell or early eight-cell embryos prevents compaction, the first major change in cell shape in early mouse embryogenesis. Divalent succinly Con A does not affect development, suggesting that the Con A effect is due to crosslinking of cell surface glycoconjugates. Exposure of four-cell or early eight-cell embryos to 10 μg/ml Lotus Tetragonolobus puprureas agglutinin (LTA; α-L-fucose-specific) or 25 μg/ml Limulus polyphemus agglutinin (LPA; sialic acid-specific) allows compaction or development to the morula stage, but blocks blastocyst formation. All lectins tested retard cell division to some extent. Late morulae and early blastocysts are more resistant than earlier stages to all of the lectins studied. This study demonstrates that very low concentrations of these lectins affect different developmental processes, presumably based upon their sugar specificities.