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Calcium signalling during zebrafish embryonic development.

Calcium signals appear throughout the first 24 hours of zebrafish development. These begin at egg activation, then continue to be generated throughout the subsequent zygote, cleavage, blastula, gastrula, and segmentation periods. They are thus associated with the major phases of pattern formation: cell proliferation, cell differentiation, axis determination, the generation of primary germ layers, the emergence of rudimentary organ systems, and therefore the establishment of the basic vertebrate body plan. When signals need to be transmitted across significant distances they take the form of waves, either intracellular waves when the cell size is large, or later in development when the cell size is reduced, intercellular waves. We will consider both types of calcium signals and their integration into signalling networks, and discuss their possible functions and developmental significance with regard to pattern formation. BioEssays 22:113-123, 2000.

Animals↗

Specific pan-neural crest expression of zebrafish Crestin throughout embryonic development.

Zebrafish crestin was identified in a screen for genes dependent on cyclops function and is a member of a family of retroelements (Rubinstein et al. [2000] Genesis 26:86-97). We report here a detailed description of crestin mRNA expression during zebrafish embryogenesis. Crestin expression was first observed during the onset of somitogenesis in cells of the neural crest domain of the ectoderm. Crestin expression was subsequently observed in premigratory cranial and trunk neural crest cells and then in actively migrating crest cells. Cell counts of crestin-expressing premigratory trunk neural crest cells strongly suggest that crestin is expressed by all neural crest cells at this stage. Crestin expression co-localized with a battery of markers for premigratory neural crest cells, developmentally distinct neural crest-derived precursor sublineages, and overtly differentiated neural crest-derived cell types. Expression of crestin is gradually downregulated in overtly differentiated cells. Our results indicate that crestin is a specific pan-neural crest marker throughout zebrafish embryogenesis.

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The effect of cyproheptadine chlorhydrate on rat embryonic development.

The teratogenic effect of cyproheptadine chlorhydrate was studied in Wistar rats. Rats were given the substance by gastric intubation on days 6-15 of pregnancy in two experimental series. In the first series rats were treated at doses of 25 and 50 mg/kg/d and in the second one they received 15, 25 and 35 mg/kg/d. Controls received only an equivalent volume of water by the same route. Doses of 25 or more mg/kg/d are highly embryotoxic. At 15 mg/kg/d the effects were relatively minor. The major anomalies caused by the drug were edema and abnormal ossification of the ribs. Some other malformations were also found: craniorrhachischisis, cleft lip, cleft palate, hypoplastic limbs, micrognathy, micromelia, and vascular damage.

Abnormalities, Multiple↗

Culture in vitro as a means of analysing the effect of maternal diabetes on embryonic development in rats.

Rodents have been used by many investigators for studies of the effects that maternal diabetes during pregnancy may have on developing fetuses. In Wistar rats, the induction of mild chronic diabetes at the onset of pregnancy by alloxan or streptozotocin results in abnormalities of the nervous system and the heart, recognized in embryos at 11--13 days' gestation. Thus, early organogenesis is evidently affected in these embryos. With a method which enables rat embryos to be cultured in vitro in serum for the period of their early organogenesis, the growth and differentiation of embryos from normal and from diabetic rats can be observed in some detail. It is also possible to compare the effects of normal and of diabetic maternal serum on their development. The results reported here show that embryos from diabetic animals are more likely to be retarded or abnormal than those from non-diabetic animals when cultured in identical serum. The development of both types of embryo is more successful in diabetic than in non-diabetic serum, however, possibly because of the higher glucose content of diabetic serum. Cultures of fetal organs may also be used as test systems for the effects of diabetic maternal serum. Sacral vertebrae, some of which fail to ossify in fetuses from daibetic rats, are now being grown in media containing diabetic serum. It is planned to test the effects of insulin on these cultures.

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Timing of ovulation and early embryonic development in Myotis lucifugus (Chiroptera: Vespertilionidae) from northern central Ontario.

Female Myotis lucifugus lucifugus, obtained from a hibernaculum in northern central Ontario, were aroused during the week that normal departures from hibernation began, maintained in large cages at 28-30 degrees C, and fed mealworms daily. Some bats ovulated less than 24 hr after arousal, and with one exception, all bats with normal, mature follicles ovulated within 72 hr. Histological examination of reproductive tracts from ovulated bats indicated that oviductal transit was completed in 24-48 hr, and that blastocyst formation and implantation occurred 4 and 10 days, respectively, after fertilization. These time intervals were somewhat shorter than those reported in previous studies; however, the differences appeared to be due to the relative accuracy in determining the time of ovulation, not to differences in rates of development among different populations.

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Tissue-specific expression of type XII collagen during mouse embryonic development.

Polyclonal antibodies were raised in rabbits against a fusion peptide representing a portion of the amino-terminal non-triple-helical domain of mouse type XII collagen. The antibodies reacted with bands of 220 and 350 kDa on Western blots of mouse tissue extracts. Immunohistochemical analyses of mouse embryos demonstrated that type XII collagen is expressed mainly in dense connective tissues of tendons, ligaments, dermis, cornea, blood vessel walls, meninges, and developing membranous bones. Comparison of skin extracts and medium of cultured mouse skin fibroblasts by Western blotting showed that while tissue contain short 220 kDa type XII collagen polypeptides as well as the long form, cultured cells produce mainly the long form with 350 kDa polypeptides.

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Pattern regulation in the chick autopodium at advanced stages of embryonic development.

In previous studies we have observed that the interdigital tissue of the chick embryo leg bud during the stages previous to interdigital cell death exhibits a considerable chondrogenic potentiality both in vivo and in vitro. In the present investigation we have carried out a variety of experimental manipulations of the chick leg bud at stage 29 to discover possible mechanisms accounting for interdigital ectopic chondrogenesis and extradigit formation. Our results show that the interdigital tissue is capable of forming an extradigit when temporarily isolated microsurgically and regrafted in its original location and after deletion of one of the adjacent digital primordia, suggesting that developing phalangeal cartilages exercise an inhibitory effect on chondrification in adjoining tissues. Furthermore, and of greater importance, ablation of the primordium of a digit is followed by normal development of the definitive digit if the wound surfaces are suitably apposed. These results reveal a considerable regulatory potential in the autopodium at advanced stages of development.

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Stages of embryonic development of the zebrafish.

We describe a series of stages for development of the embryo of the zebrafish, Danio (Brachydanio) rerio. We define seven broad periods of embryogenesis--the zygote, cleavage, blastula, gastrula, segmentation, pharyngula, and hatching periods. These divisions highlight the changing spectrum of major developmental processes that occur during the first 3 days after fertilization, and we review some of what is known about morphogenesis and other significant events that occur during each of the periods. Stages subdivide the periods. Stages are named, not numbered as in most other series, providing for flexibility and continued evolution of the staging series as we learn more about development in this species. The stages, and their names, are based on morphological features, generally readily identified by examination of the live embryo with the dissecting stereomicroscope. The descriptions also fully utilize the optical transparancy of the live embryo, which provides for visibility of even very deep structures when the embryo is examined with the compound microscope and Nomarski interference contrast illumination. Photomicrographs and composite camera lucida line drawings characterize the stages pictorially. Other figures chart the development of distinctive characters used as staging aid signposts.

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The embryonic development of theproctodeal gland of Coturnix coturnix japonica (Aves).

The embryogenesis of the proctodeal gland and development of the connective tissue of the associated lamina propria in the dorsal wall of the proctodeum of Common Coturnix (Coturnix c. japonica) were studied on embryos collected at 12-hour intervals from day 7 of incubation through hatching. Gland development began at 9.5 days from ectodermal-derived epithelial buds developing in the craniolateral region of the dorsal proctodeal epithelium. At 11 days, the epithelial buds had become solid convoluted epithelial cords which, by 11.5 days branced to form solid individual glandular units. Lumina of the unit were formed by degeneration and vacuolization of the central cells of these solid units. Squamous epithelial caps, several layers thick, separated the vacuolizing units from the proctodeal cavity at day 13; these caps subsequently thinned, bulged into the proctodeal cavity and, by day 16, began to disappear opening the lumen of the units to the proctodeal cavity. Sequential formation of epithelial buds, by region, was: craniolateral at 9.5 days, craniomedial expansion at 10.5 days followed, beginning at 11 days, by caudal expansion along entire dorsal proctodeal wall. Sequence of appearance of gland-associated connective tissue fibers was: 9.5 days, reticular fibers already present; 11 days, collagen fibers; 11.5 days elastic fibers. Serially reconstructed models of an individual glandular unit at hatching enabled classification of the unit as a simple branched alveolar gland.

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Ultrastructural study of the embryonic development in the rat pineal gland.

The ultrastructure of the albino rat embryo pineal gland was studied from day 13 of development through birth. In the first stages (13-16.5 days of development) the pineal evagination presents a barely differentiated epithelium. From 17 days onward the transformation of the pineal gland from a tubular evagination into a compact organ occurs. The obliteration of the recess takes place by means of two mechanisms: (a) multiple foldings of the epithelium which determine an approximation and fusion of the walls of the recess, and (b) occupation of the lumen by cells extruded from the pineal epithelium. Embryos of 18-21 days of gestation still show remains of the pineal recess. From day 16.5 onward elements of the pineal parenchyma have been found outside the pineal epithelium contour. They contact with the mesenchymal cells without a basal lamina separating both elements. Day 20 marks the beginning of recognizable differentiation of pineal cellular types. However, in the newborn rat these types are not yet clearly established.

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Embryonic development of the posterolateral structures of the knee.

The development of the structures in the posterolateral corner of the knee was studied in fifteen human embryos and seventeen fetuses by means of serial sections in the coronal and sagittal planes. The attachments of the lateral meniscus and fibular head to the popliteal tendon and muscle are formed during the process of cavitation that forms the bursa. The connection between the tendon and the posterior border of the lateral meniscus forms obliquely to follow the direction of the muscle and tendon. The edge of the bursa is limited by attachments that connect the tendon to the lateral meniscus and fibular head. In this study, no direct ligamentous continuity was found to exist between the posterolateral femur and tibia during development. The only ligament that indirectly connects the lateral femur posteriorly to the proximal segment of the tibia, which has never been reported in human development, is the one that links the popliteal tendon to the fibular head. We have designated this the popliteo fibular ligament which was also found in adult specimens. This ligament should not be confused with the arcuate popliteal ligament.

Fibula↗

Expression of cVg1 mRNA during chicken embryonic development.

Using degenerated PCR-primers to identify known and novel BMPs that are expressed in the developing chicken heart, we identified not only BMP2, -4, and -7 mRNA, but also the TGFbeta superfamily member cVg1. The expression pattern of cVg1 mRNA was determined during chicken development from HH4 to HH44. In early developmental stages, cVg1 mRNA is expressed in the primitive streak, paraxial mesoderm, developing somites, and developing neural tube. Subsequently, cVg1 mRNA is expressed in the developing central and peripheral nervous system, retina, auditory vesicle, notochord, lung alveoli, and olfactory mucosa. In the heart, cVg1 is initially expressed through the linear heart tube, but becomes restricted to the forming chamber myocardium, in an expression domain similar to that of atrial natriuretic factor (ANF) mRNA.

Animals↗

Innervation of avian latissimus dorsi muscles and axonal outgrowth pattern in the posterior latissimus dorsi motor nerve during embryonic development.

The distribution of the innervation to the anterior latissimus dorsi (ALD) and posterior latissimus dorsi (PLD) muscles of the chicken are described on the day of hatching and 6 weeks later using electron microscopy. In the ALD muscle, there are 5,000 muscle fibres and 374,000 endplates supplied by about 169 skeletomotor axons; in the PLD muscle, there are 12,000 focally innervated muscle fibers supplied by about 20 skeletomotor axons. On the cell surface of the muscle fibers the mean total subsynaptic area contacted by each motor axon is comparable in the ALD and PLD muscles. The growth pattern of the axons in the PLD motor nerve was described from the ninth day in ovo up to 6 weeks after hatching. The axons arrive in the PLD muscle in two successive waves: first, the large somatic axons which are already present before the ninth day in ovo and second, the small autonomic axons which continue to accumulate until hatching. The total number of somatic axons decreases from the ninth day until the hatching day when it reaches its definitive value. This decrease takes place during a period when the numbers of myofibers and of endplates dramatically increase, and it coincides with the axonal segregation by the Schwann cells. The myelination of the axons starts on the 15th day in ovo and is essentially complete upon hatching. Despite the decreasing number of somatic axons in the PLD nerve, the decrease in number of nerve endings per PLD endplate and the increasing number of PLD endplates per PLD muscle, it was found that between the 16th day in ovo and 6 weeks after hatching the mean number of axonal branches per PLD motor axon does not decrease.

Aging↗

Light and electron microscopic studies of the basilar papilla in the duck, Anas platyrhynchos. II. Embryonic development.

The intent of this study was to describe certain morphological aspects of cytodifferentiation of the basilar papilla in the duck, Anas platyrhynchos. The duck, with a longer incubation period than that of the chicken, has proven useful in identifying the earliest morphological changes during cyto-differentiation of the basilar papilla. Most of these changes occur first at the neural margin of the presumptive genu, at the site of initial contact by cochlear nerve fibers. A wave of differentiation of hair cells radiates abneurally, proximally, and distally, concurrent with the establishment of afferent innervation. Extracellular channels form in the papilla between cell bases just before the nerve fibers penetrate the basal lamina. Nerve fibers abut hair cell bases in the genu at the time the hair cells separate from the basal lamina on day 9. Hair cell bases contain granular vesicles and lie among the nuclei of supporting cells on day 10. Synaptic bodies are present opposite nerve endings on day 11. Efferent nerve endings appear on short hair cells in the genu on day 15 and on tall hair cells on day 17.

Animals↗

Embryonic development of the rat cerebellum. II. Translocation and regional distribution of the deep neurons.

In thymidine radiograms and plastic-embedded sections, the migration of cerebellar deep neurons was traced from their germinal source to their final settling sites. The route proved to be roundabout and three developmental events could be distinguished during the process. First, between days E14 and E16, transversely oriented cells of the nuclear transitory zone move in an arc from the ventrolateral neuroepithelium of the lateral cerebellar primordium in a medial direction. Second, between days E16 and E18, the cells of the rostral component of the nuclear transitory zone assume a longitudinal orientation. We postulated that this is the period of axonogenesis, the longitudinally oriented cells issuing efferents that join the superior cerebellar peduncle ipsilaterally and the transversely oriented cells (representing the neurons of the caudal fastigial nucleus) sending decussating fibers to the uncinate fasciculus (the hook bundle of Russell). Third, between days E18 and E21, the earlier-produced superficial cells of the nuclear transitory zone and the later-produced deep cells of the cortical transitory zone (the young Purkinje cells) exchange positions. The descent of the deep neurons is in the direction of the fibers of the inferior cerebellar peduncle, which becomes distributed throughout the cerebellum on day E17. The ascent of the Purkinje cells is in the direction of the external germinal layer, which begins to spread from caudal to rostral on day E17. The three deep nuclei, the lateral (dentate), interpositus, and medial (fastigial), can be distinguished before their descent into the depth of the cerebellum, and by day E22 a small-celled and a large-celled subdivision is identifiable in each nucleus.

Animals↗

Embryonic development of the rat cerebellum. III. Regional differences in the time of origin, migration, and settling of Purkinje cells.

The time of origin, site of origin, migratory path and settling pattern of the Purkinje cells of the cerebellar hemispheres, anterior vermis, and posterior vermis were investigated in thymidine radiograms and plastic-embedded materials from rat embryos ranging in age from 15 to 22 days. In the hemispheres there is a rostral-to-caudal cytogenetic gradient: the Purkinje cells of lobulus simplex, crus I, and crus II are produced earlier than the Purkinje cells of the paramedian lobule and paraflocculus, followed by the Purkinje cells of the flocculus. The Purkinje cells of the vermis, in general, are generated later than those of the hemispheres, and with a reverse gradient from caudal to rostral: the Purkinje cells of the posterior vermis (lobules X-VI) being produced ahead of the Purkinje cells of the anterior posteriorly directed wedge of early-produced Purkinje cells through the vermis. Evidence was obtained that the Purkinje cells of the hemispheres derive from the lateral cerebellar primordium capping the lateral recess of the fourth ventricle anteriorly. The Purkinje cells of the anterior vermis originate from the subisthmal cerebellar primordium medially lining the isthmal canal. The Purkinje cells of the posterior vermis originate in the postisthmal cerebellar primordium overlying the tela choroidea caudally. The young Purkinje cells migrate from the neuroepithelium to the surface of the cerebellum in a strictly caudal-to-rostral order, paralleling the spread of the EGL superficially from posteroventral to anterodorsal. This pattern is independent of the time of origin of Purkinje cells. In the posterior vermis the earliest-settling Purkinje cells of the uvula follow a short radial course, and a discrete Purkinje layer is formed 3 days after they are generated. In the anterior vermis the Purkinje cells of lobulus centralis, which follow an anterodorsal migratory course, are still settling on day E22, 7 days after their production, presumably awaiting the fusion of the cerebellar base anteriorly. The fissura prima forms medially at the interface region of Purkinje cells derived from the postisthmal and subisthmal cerebellar primordia. For 1-2 days after their settling, the Purkinje cells of the newly forming lobules can be distinguished by certain cytological criteria from the Purkinje cells in the more caudally-situated, earlier-settled lobules.

Animals↗