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PubMed1976

Characterization of embryonic antigens in the plasma of developing chick embryos.

Two prominent alpha-fetoproteins are found in chick embryo plasma up to and slightly beyond hatching. Only antisera against chick embryo plasma resolve them. alpha3-fetoprotein is a lipoprotein, the first fetoprotein thus described. It probably functions to transport lipid from the yolk to the embryo. The other fetoprotein (alpha4) is a major glycoprotein. Both are made by the liver and yolk sac. From 9 days onward, albumin is the principal export protein of the liver; at 7 days, when it first appears in the plasma, the yolk sac, but not the liver, makes it. The reverse situation occurs at 9 days. These facts suggest a humoral relationship between the yolk sac and liver. The embryo synthesizes every plasma protein including fetoproteins and "adult" proteins (prealbumin, albumin, and transferrin). No "cold" or unlabeled proteins are seen.

Animals↗

PubMed1992

Histochemical identification and behavior of quail primordial germ cells injected into chick embryos by the intravascular route.

The behavior of quail primordial germ cells (PGC) after injection into chick embryos by the intravascular route was examined. The quail (donor) PGC, taken from the bloodstream of quail embryos (recipient) at stage 13-14, were injected into the vitelline vessels of chick embryos (recipient) at stage 15. In the recipient embryos, the PGC of the quail and the chick were histochemically distinguished by a double-staining technique involving a lectin, from Wistaria floribunda (WFA) and the PAS reaction. One day after injection, quail PGC appeared in the prospective gonadal region of recipient chick embryos, being localized among the recipient chick PGC. This result indicates that a staining technique specific for WFA lectin is useful for identification of quail PGC and that quail PGC can be transferred by a vascular route for the production of germline chimeras.

Animals↗

PubMed1989

Biochemical aspects of chick embryo retina development: the effects of glucocorticoids.

In chick embryo retina during development, DNA synthesis and the activities of DNA polymerase, thymidine kinase, thymidylate synthetase, and ornithine decarboxylase (ODC) declined in parallel from day 7 to 12. The administration in ovo of hydrocortisone reduced significantly, particularly at 8-10 days of incubation, both DNA synthesis and the four enzyme activities tested. The effect was dose dependent, reaching the maximum with 50-100 nmol of hydrocortisone, 8-16 h after treatment. The highest inhibition was found for ODC activity (70%), followed by thymidine kinase activity (62%) and DNA synthesis (45%), whereas activities of DNA polymerase and thymidylate synthetase were reduced only by 30%. The inhibitory effect was exerted by all the glucocorticoids tested, with dexamethasone and hydrocortisone being the most efficacious. The results support the view that glucocorticoids reduce the proliferative events in chick embryo retina, particularly at 8-10 days of embryonic life.

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PubMed1978

Scanning electron microscopy of primordial germ cells in early chick embryos.

Primordial germ cells (PGCs) of the early chick embryo were examined using scanning electron microscopy. Temporal changes in the form and distribution of surface projections were found to be correlated with migratory phases of PGCs. Non-migrating PGCs were spherical to ovoid with relatively smooth surfaces. Their transition to the migratory phase was first evidenced by a burst of membrane activity. Migrating PGCs became somewhat flattened against the underlying hypoblast (which serves as the substratum for their migration) and exhibited blebs and lamellar processes. The lamellar processes were most prominent at the leading edges of actively migrating PGCs. Overall results of the present study indicates that PGCs found in the germinal crescent area of early chick embryos actively migrate on the dorsal surface of the hypoblast towards posterior embryonic regions.

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PubMed1982

Effect of augmentation of nerve supply upon limb regeneration in the chick embryo.

Limb regeneration was induced in 4-day chick embryos by implanting stage 15 neural tube into the amputated stump of the limb. In control animals, amputation alone or amputation with implant of either notochord or somites resulted in development of proximal wing segments only. In experimental animals, more than one-fourth of the animals containing viable neural tube implants developed proximal, middle, and distal limb segments. These regenerated limb segments contained muscle groups, cartilage models, and nervous structures that approximated the normal situations. Since some new parts regenerated in more than one-fourth of the cases, it may be concluded that augmentation of nerve supply is an effective method of inducing regeneration in limbs of chick embryos. Volume measurements of the neural tube implant indicate that a "critical mass" of nerve tissue may be necessary for regeneration to occur.

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PubMed1988

Partitioning behaviour in aqueous two-phase systems and fractionation by counter-current distribution of chick-embryo erythrocytes with numerical resolution of distribution curves.

The partition of chick-embryo and young-chick erythrocytes in dextran-poly-(ethylene glycol) two-phase systems depends on the interfacial tension and electrical potential differences between the phases. Counter-current distribution with charged 5% dextran-poly(ethylene glycol) systems has proved to be an adequate method for the separation of primitive and definitive erythrocytes present in chick embryos when a phase settling time of 20 min is used. The computer-aided numerical resolution of experimental curves has shown the existence of subpopulations which could not have been detected by using conventional methods.

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PubMed1984

Development of calcium reabsorption by the allantoic epithelium in chick embryos grown in shell-less culture.

The allantoic sac of the chick embryo functions as a primitive urinary bladder, storing and modifying the excretory fluid produced by the embryo. We have used chick embryos grown in shell-less culture to study the in situ handling of Ca2+ by the allantoic epithelium. Between Days 8 and 13 of incubation (38 degrees C, 5% CO2), the [Ca2+] of the allantoic sac fluid declines from about 1.5 mM to less than 0.3 mM, with most of this Ca2+ reabsorption occurring between Days 10 and 11. In 13-day-old embryos, the allantoic epithelium reabsorbs within 24 hr 85-92% of 45Ca2+ injected into the allantoic sac, while in 9-day-old embryos 45Ca2+ reabsorption is less than 40% by 24 hr. This is evidence for the developmental onset of a Ca2+ reabsorption process in the allantoic epithelium. The allantoic fluid Ca2+ is reabsorbed into the embryo's blood in which the serum [Ca2+] is about 1.5 mM. Also, electrical potential profiles reveal that the serosal (mesenchymal) side of the allantoic epithelium is 15-30 mV positive compared to the mucosal (luminal) side. Thus, by electrochemical criteria this reabsorption process appears to be active.

Absorption↗

PubMed1995

Evidence for calcium regulation of spinal cord motoneuron death in the chick embryo in vivo.

We have studied in living chick embryos the effects of an extracellular calcium load on the induction of apoptosis in spinal cord motoneurons. The action of a calcium ionophore, A23187, that does not raise extracellular calcium was also evaluated in order to explore the role of endogenous calcium in determining developmentally-regulated cell death of motoneurons. The application of a single dose of 50 microliters of 1.8 M CaCl2 onto the chorioallantoic membrane of E7 chick embryos produce a transient elevation of intraembryonic calcium concentration that was followed by a transitory rise in the number of apoptotic cells in the lateral motor column. Administration of 250 microM of the ionophore A23187 (100 microliters), also results in an increase in apoptosis of motoneurons in the lateral motor column on E6 and E7 but this effect is progressively lost following treatment at more advanced stages of development. Neither of these effects can be explained by unspecific calcium cytotoxicity since they can be inhibited by prior administration of the protein synthesis inhibitor cycloheximide or the neuromuscular blocking agent (+)-tubocurarine. After calcium loading, degenerating cells display similar ultrastructural characteristics as during physiologically occurring motoneuron death and exhibit histochemically detectable DNA fragmentation. Chronic administration of CaCl2 or A23187 does not reduce the total number of surviving motoneurons at the end of the normal period of naturally occurring motoneuron death (E10). It is suggested that calcium loading stimulates and accelerates the physiological degeneration of a restricted subpopulation of motoneurons which will undergo the process of natural cell death.

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PubMed1981

Lodish model and regulation of ribosomal protein synthesis by insulin-deficient chick embryo fibroblasts.

The production of ribosomal proteins in chick embryo fibroblasts that have been deprived of insulin is depressed in a much greater degree than that of most or all other cell proteins. Previous observations ruled out explanations for the preferential decrease in ribosomal protein formation that depend upon a selective destruction of ribosomal protein messages or a regulatory role for nascent ribosomal ribonucleic acid. The proposition has now been examined that ribosomal protein messenger ribonucleic acids (mRNAs) in the hormone-deficient chick embryo cells have a lower affinity for a limiting, early acting component of the initiating machinery than do most other cell messages and, in consequence, suffer from a translational disadvantage. The approach that was used depends upon the findings of Lodish and others that all mRNAs are not initiated with equal ease, that inhibitors of elongation favor the initiation of low-affinity mRNAs, and that agents that dampen an early step in initiation discriminate against the low-affinity messages. The idea was tested by comparing the effects of various inhibitors on the rates of synthesis of total cell protein and individual nonribosomal proteins, on the one hand, with those of individual ribosomal proteins, on the other. The results fit the Lodish model and are consistent with the conclusions that ribosomal protein mRNAs are more poorly initiated in the resulting fibroblasts than are most or all other cell messages and that this condition is largely or entirely responsible for the low rate of ribosomal protein formation.

Animals↗

PubMed1978

Further characterization of collagen galactosyltransferase from chick embryos.

Optimum extraction of collagen galactosyltransferase activity from chick embryos required relatively high concentrations of detergent and salt. The activity was inhibited by concanavalin A, and the enzyme had a high affinity for columns of this lectin coupled to agarose; these results suggest the presence of carbohydrate units in the enzyme molecule. Collagen galactosyltransferase was highly labile, and only 1% of the originally bound enzyme activity could be eluted from the concanavalin A-agarose column with a buffer containing methyl glucoside and ethylene glycol. The purification of the activity over the original supernatant of chick embryo homogenate was 250-300-fold, with the optimum reaction conditions for the purified transferase differing somewhat from those for crude enzyme preparations. The reaction was inhibited by glucose-free basement-membrane collagen, UDP and galactosylhydroxylsine, and also by Co2+ and a number of compounds resembling UDP-galactose. Hydroxylysine was also a weak inhibitor. Immobilized hydroxylysine and UDP-glucuronic acid did not bind the collagen galactosyltransferase, but the enzyme was retarded in a column of UDP-galacturonic acid linked to agarose.

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PubMed2000

Differential effects of the trophic factors BDNF, NT-4, GDNF, and IGF-I on the isthmo-optic nucleus in chick embryos.

The isthmo-optic nucleus (ION) of chick embryos is a model system for the study of retrograde trophic signaling in developing CNS neurons. The role of brain-derived neurotrophic factor (BDNF) is well established in this system. Recent work has implicated neurotrophin-4 (NT-4), glial cell line-derived neurotrophic factor (GDNF), and insulin-like growth factor I (IGF-I) as additional trophic factors for ION neurons. Here it was examined in vitro and in vivo whether these factors are target-derived trophic factors for the ION in 13- to 16-day-old chick embryos. Unlike BDNF, neither GDNF, NT-4, nor IGF-I increased the survival of ION neurons in dissociated cultures identified by retrograde labeling with the fluorescent tracer DiI. BDNF and IGF-I promoted neurite outgrowth from ION explants, whereas GDNF and NT-4 had no effect. Injections of NT-4, but not GDNF, in the retina decreased the survival of ION neurons and accelerated cell death in the ION. NT-4-like immunoreactivity was present in the retina and the ION. Exogenous, radiolabeled NT-4, but not GDNF or IGF-I, was retrogradely transported from the retina to the ION. NT-4 transport was significantly reduced by coinjection of excess cold nerve growth factor (NGF), indicating that the majority of NT-4 bound to p75 neurotrophin receptors during axonal transport. Binding of NT-4 to chick p75 receptors was confirmed in L-cells, which express chick p75 receptors. These data indicate that GDNF has no direct trophic effects on ION neurons. IGF-I may be an afferent trophic factor for the ION, and NT-4 may act as an antagonist to BDNF, either by competing with BDNF for p75 and/or trkB binding or by signaling cell death via p75.

Animals↗

PubMed1993

Tissue variation of two large chondroitin sulfate proteoglycans (PG-M/versican and PG-H/aggrecan) in chick embryos.

PG-M and PG-H, chick large chondroitin sulfate proteoglycans corresponding to versican (fibroblast-type proteoglycan) and aggrecan (cartilage-characteristic proteoglycan), respectively, which are found in mammals, have been characterized in various tissues of chick embryos. Their distribution and the compositions of the core molecules were analyzed by immunofluorescence staining and immunoblotting, respectively, using various monospecific antibodies. Molecules reactive to a monoclonal antibody to the PG-M core protein (designated MY-174) were distributed in various tissues, including aorta, lung, cornea, brain, skeletal muscle and dermis. Immunoblotting with MY-174 of the chondroitinase ABC-digested tissue extracts showed a tissue variation of MY-174-reactive core molecules (550-kD, 500-kD, 450-kD, and 350-300-kD). In contrast, PG-H, besides massive occurrence in cartilage, was only found in a few tissues such as aorta and brain. In addition, PG-H in aorta, cornea, and skin was atypical in structure, because it lacked keratan sulfate. The expression of PG-M in developing chick embryos was then examined. PG-M was found in some developmentally active areas, such as the perinotochordal mesenchyme between notochord and neural tube, the basement membranes facing neuroepithelial cells, and condensing mesenchymal cells in limb buds, suggesting some functions distinctive of the developing tissues.

Aggrecans↗

PubMed2005

RNA interference is ineffective as a routine method for gene silencing in chick embryos as monitored by fgf8 silencing.

The in vivo accessibility of the chick embryo makes it a favoured model system for experimental developmental biology. Although the range of available techniques now extends to miss-expression of genes through in ovo electroporation, it remains difficult to knock out individual gene expression. Recently, the possibility of silencing gene expression by RNAi in chick embryos has been reported. However, published studies show only discrete quantitative differences in the expression of the endogenous targeted genes and unclear morphological alterations. To elucidate whether the tools currently available are adequate to silence gene expression sufficiently to produce a clear and specific null-like mutant phenotype, we have performed several experiments with different molecules that trigger RNAi: dsRNA, siRNA, and shRNA produced from a plasmid coexpressing green fluorescent protein as an internal marker. Focussing on fgf8 expression in the developing isthmus, we show that no morphological defects are observed, and that fgf8 expression is neither silenced in embryos microinjected with dsRNA nor in embryos microinjected and electroporated with a pool of siRNAs. Moreover, fgf8 expression was not significantly silenced in most isthmic cells transformed with a plasmid producing engineered shRNAs to fgf8. We also show that siRNA molecules do not spread significantly from cell to cell as reported for invertebrates, suggesting the existence of molecular differences between different model systems that may explain the different responses to RNAi. Although our results are basically in agreement with previously reported studies, we suggest, in contrast to them, that with currently available tools and techniques the number of cells in which fgf8 gene expression is decreased, if any, is not sufficient to generate a detectable mutant phenotype, thus making RNAi useless as a routine method for functional gene analysis in chick embryos.

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PubMed1977

Utilization of analogues of riboflavin by the riboflavin-deficient chick embryo.

Detailed studies of the biological activity of several analogues of riboflavin in the riboflavin-deficient chick embryo were undertaken to extend our knowledge of the comparative biochemistry of these flavins. The riboflavin-deficient eggs were produced by hens homozygous for an autosomal recessive gene (rdrd). 7-Ethyl-8-methyl-flavin was found to be an adequate replacement for riboflavin in che chick embryo, a finding which mimics its effect in the riboflavin-deficient rat. 7,8-Diethyl-flavin was found to be an antagonist in the chick embryo as had been found to be true in the rat. 7-Methyl-8-ethyl-flavin can not be used as a replacement for riboflavin in the chick embryo and further, it inhibits the utilization of riboflavin in normal eggs. 7-Methyl-8-ethyl-flavin had been found to be an adequate replacement for riboflavin in the riboflavin-deficient rat. The classes mammalia and aves are clearly different in respect to the utilization of this flavin.

Animals↗

PubMed1983

Functional development of the hypothalamic-adenohypophyseal-testicular (HAT) axis in the chick embryo.

Plasma testosterone concentrations in intact and surgically partially decapitated (hypothalamo-hypophyseoprivic) chick embryos were determined by radioimmunoassay for the incubation interval, Days 7.5-17.5. "Hypophysectomy" of male chick embryos at 33-38 hr of incubation resulted in plasma testosterone levels which were significantly lower than those of intact embryos on Days 13.5, 15.5, and 17.5. The addition of luteinizing hormone (LH) to the chorioallantoic membrane (CAM) of decapitated embryos increased plasma testosterone concentrations to normal levels by Day 13.5, while LH and pituitary transplants to the CAM elevated plasma testosterone to above normal values by Day 15.5. These observations, together with other findings, are interpreted as demonstrating that in the chick embryo the adenohypophyseal-testicular axis is functional by Day 13.5 of incubation. There are also indications that the hypothalamic-adenohypophyseal-testicular complex is functional at this same embryonic time.

Age Factors↗

PubMed1985

Induction of chick embryo feather malformations by an influenza C virus.

The effect of influenza C virus, strain JJ/50, on the development of chicken embryos infected at 10 or 12 days was documented by microscopic techniques, as well as by gross observations of embryos or chicks at hatching. The infected, newly hatched chicks displayed marked abnormalities in their feathering. Such abnormalities were observed neither in mock-infected embryos nor in embryos injected with virus which had been previously treated with specific influenza C virus antibody. At a microscopic level, the abnormalities apparently are a result of hypertrophy and/or hyperplasia of the developing barb and barbule cells. Further, the additional development of integumental necrotic foci was correlated with the development of relatively high viral titers (greater than 256) as measured by hemagglutination (HA). Embryos infected after 12 instead of 10 days incubation showed normal feathering at hatching. Infection at 12 days, however, was correlated with the development of relatively low viral titers (HA = 4) and limited degeneration of the respiratory epithelium. The relationship of teratogenic effects to the site of viral replication in rapidly differentiating tissue is discussed.

Animals↗

PubMed1991

[Chick embryo culture using duck egg shell--first successful hatch ].

The suitability of duck egg shell (DES) for chick embryo culture was investigated. Chick embryos were transferred into DESs with all egg contents after 3 days of normal incubation and cultured. The vessels made of polyethylene cling film were used for shell-less control. Among 35 embryos cultured in DESs, 21 survived until 16 days of incubation (13 days after transfer) and finally 3 newly hatched chicks were obtained at 22 days of incubation. One of them died 4 days later, but remaining two became full-grown cocks showing normal body weight and production of fertile sperms. Among 37 embryos cultured in polyethylene vessels, none survived over the period of 19 days of incubation. It is suggested that DES culture system may be useful for the various experiments using chick embryos.

Animals↗

PubMed1982

Embryotoxicity of 3,3',4,4'-tetrachloroazobenzene and 3,3',4,4'-tetrachloroazoxybenzene in the chick embryo.

The toxicity of 3,3',4,4'-tetrachloroazobenzene (TCAB) and 3,3',4,4'-tetrachloroazoxybenzene (TCAOB) to chick embryos was examined. TCAB or TCAOB was dissolved in corn oil and injected into the air cell of fertile chicken eggs. The time of injection had a major effect on embryo mortality as eggs injected with TCAB or TCAOB on the fourth day of incubation had a higher incidence of embryo mortality than eggs injected on days 11-13. Both TCAB and TCAOB were more toxic than all other chemicals that have been tested in the chick embryo with the exception of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). Comparing the potency of the two compounds, TCAOB was more potent than TCAB in the chick embryo with an estimated LD50 of 12 ng and 44 ng respectively. Rump edema was the major abnormality observed in embryos treated with either TCAB or TCAOB. Other malformations included altered feather pattern and lack of down, hemorrhage, external viscera, reduced body size, failure to withdraw the yolk sac, beak malformation, dilation of blood vessels, and monomicropthalmia. The results of this investigation suggest that both TCAB and TCAOB are teratogenic in the chick embryo.

Animals↗