Analysis of retinal cell development in chick embryo by immunohistochemistry and in ovo electroporation techniques

Retinal,cell aggregates develop in culture in a pattern similar to the in ovo retina, forming neurites first and then synapses. In the present study, we continuously exposed chick retinal cell aggregates to a high concentration (1 mM) of carbamylcholine (carbachol), an acetylcholine (ACh) analog that resists hydrolysis by acetylcholinesterase (AChE). This situation is similar to organophosphorus anticholinesterase poisoning in which the ACh level is elevated at synaptic junctions due to inhibition of AChE, Our objective was to determine whether continuous carbachol exposure either damaged cholino- ceptive neurites, cell bodies, and synaptic elements of the aggregates or influenced (hastened or retarded) their development.The retinal tissue was isolated aseptically from 11 day embryonic White Leghorn chicks and then enzymatically (trypsin) and mechanically (trituration) dissociated into single cells. After washing the cells by repeated suspension and low (about 200 x G) centrifugation twice, aggregate cell cultures (about l0 cells/culture) were initiated in 1.5 ml medium (BME, GIBCO) in 35 mm sterile culture dishes and maintained as experimental (containing 10-3 M carbachol) and control specimens.

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GnRH Neuronal Migration and Olfactory Bulb Neurite Outgrowth Are Dependent on FGF Receptor 1 Signaling, Specifically via the PI3K p110α Isoform in Chick Embryo

Endocrinology ◽

10.1210/en.2012-1555 ◽

2013 ◽

Vol 154 (1) ◽

pp. 388-399 ◽

Cited By ~ 33

Author(s):

Youli Hu ◽

Subathra Poopalasundaram ◽

Anthony Graham ◽

Pierre-Marc Bouloux

Keyword(s):

Chick Embryo ◽

Olfactory Bulb ◽

Neurite Outgrowth ◽

Neuronal Migration ◽

Glycogen Synthase ◽

Kallmann Syndrome ◽

Pi3k Inhibitor ◽

Fgf Signaling ◽

In Ovo ◽

Fgf Receptor

Fibroblast growth factor (FGF) signaling is essential for both olfactory bulb (OB) morphogenesis and the specification, migration, and maturation of the GnRH-secreting neurons. Disruption of FGF signaling contributes to Kallmann syndrome characterized by both anosmia and sexual immaturity. However, several unanswered questions remain as to which specific FGF receptor (FGFR)-1 signaling pathways are necessary for OB and GnRH neuronal development. Here, using pharmacological phosphatidylinositol 3-kinase (PI3K) isoform-specific inhibitors, we demonstrate a central role for the PI3K p110α isoform as a downstream effector of FGFR1 signaling for both GnRH neuronal migration and OB development. We show that signaling via the PI3K p110α isoform is required for GnRH neuronal migration in explant cultures of embryonic day (E) 4 chick olfactory placodes. We also show that in ovo administration of LY294002, a global PI3K inhibitor as well as an inhibitor to the PI3K p110α isoform into the olfactory placode of E3 chick embryo impairs GnRH neuronal migration toward the forebrain. In contrast, in ovo PI3K inhibitor treatment produced no obvious defects on primary olfactory sensory neuron axonal targeting and bundle formation. We also demonstrate that anosmin-1 and FGF2 induced neuronal migration of immortalized human embryonic GnRH neuroblast cells (FNC-B4-hTERT) is mediated by modulating FGFR1 signaling via the PI3K p110α isoform, specifically through phosphorylation of the PI3K downstream effectors, Akt and glycogen synthase kinase-3β. Finally, we show that neurite outgrowth and elongation of OB neurons in E10 chick OB explants are also dependent on the PI3K p110α isoform downstream of FGFR1. This study provides mechanistic insight into the etiology of Kallmann syndrome.

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Mesoblastic Cells in the Early Phase of Primitive Streak Formation in Chick Embryos. Observations by Scanning Electron Microscopy in ovo and time-Lapse Cinematography in vitro.. (chick embryo/primitive streak/mesoblastic cells/cell movement/blebs)

Development Growth & Differentiation ◽

10.1111/j.1440-169x.1984.00235.x ◽

1984 ◽

Vol 26 (3) ◽

pp. 235-247 ◽

Cited By ~ 8

Author(s):

SHIGEO TAKEUCHI

Keyword(s):

Electron Microscopy ◽

Scanning Electron Microscopy ◽

Chick Embryo ◽

Cell Movement ◽

Primitive Streak ◽

Time Lapse ◽

Chick Embryos ◽

In Ovo ◽

Scanning Electron

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Fate of brachial muscles of the chick embryo innervated by inappropriate nerves: structural, functional and histochemical analyses

Development ◽

10.1242/dev.95.1.147 ◽

1986 ◽

Vol 95 (1) ◽

pp. 147-168

Author(s):

Jane Butler ◽

Peter Cauwenbergs ◽

Ethel Cosmos

Keyword(s):

Chick Embryo ◽

Muscle Growth ◽

Extended Period ◽

Differential Rate ◽

Innervation Pattern ◽

In Ovo ◽

Intramuscular Nerve ◽

Quantitative Analyses ◽

Wing Bud ◽

Actual Loss

The extent of interaction between brachial muscles and foreign (thoracic) nerves of the chick embryo was determined during an extended period of development in ovo from the perspectives of innervation pattern, function (motility analyses), muscle growth (quantitative analyses of muscle volume) and fibre-type expression (myosin-ATPase profiles). Results indicated that according to all parameters analysed, initially a compatible union existed between the foreign nerves and their muscle targets. During subsequent stages of development, deterioration of the once compatible relationship emerged, until eventually denervation of muscles, i.e. an actual loss of intramuscular nerve branches, was observed. The process of denervation, which proceeded at a differential rate among individual muscles, however was restricted to brachial muscles derived from the premuscle masses of the wing bud. In contrast, brachial muscles of myotomal origin were spared the fate of wing-bud-derived muscles and maintained a successful union with the foreign nerves.

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Regulation of hepatic haem metabolism. Disparate mechanisms of induction of haem oxygenase by drugs and metals

Biochemical Journal ◽

10.1042/bj2500189 ◽

1988 ◽

Vol 250 (1) ◽

pp. 189-196 ◽

Cited By ~ 43

Author(s):

B C Lincoln ◽

J F Healey ◽

H L Bonkovsky

Keyword(s):

Chick Embryo ◽

Primary Culture ◽

In Ovo ◽

Oxygenase Activity ◽

Haem Oxygenase ◽

Cytochrome P 450

We studied drug- and metal-mediated increases in activity of haem oxygenase, the rate-controlling enzyme for haem breakdown, in chick-embryo hepatocytes in ovo and in primary culture. Phenobarbitone and phenobarbitone-like drugs (glutethimide, mephenytoin), which are known to increase concentrations of an isoform of cytochrome P-450 in chick-embryo hepatocytes, were found to increase activities of haem oxygenase as well. In contrast, 20-methylcholanthrene, which increases the concentration of a different isoform of cytochrome P-450, had no effect on activity of haem oxygenase. Inhibitors of haem synthesis, 4,6-dioxoheptanoic acid or desferrioxamine, prevented drug-mediated induction of both cytochrome P-450 and haem oxygenase in embryo hepatocytes in ovo or in culture. Addition of haem restored induction of both enzymes. These results are interpreted to indicate that phenobarbitone and its congeners induce haem oxygenase by increasing hepatic haem formation. In contrast, increases in haem oxygenase activity by metals such as cobalt, cadmium and iron were not dependent on increased haem synthesis and were not inhibited by 4,6-dioxoheptanoic acid. We conclude that (1) induction of hepatic haem oxygenase activity by phenobarbitone-type drugs is due to increased haem formation, and (2) induction of haem oxygenase by drugs and metals occurs by different mechanisms.

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In Ovo Electroporation of miRNA Plasmids to Silence Genes in a Temporally and Spatially Controlled Manner

MicroRNA Technologies - Neuromethods ◽

10.1007/7657_2016_5 ◽

2016 ◽

pp. 161-181

Author(s):

Nicole H. Wilson ◽

Esther T. Stoeckli

Keyword(s):

In Ovo ◽

In Ovo Electroporation

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Special Problems of Experimenting in ovo on the Early Chick Embryo, and a Solution

Development ◽

10.1242/dev.8.4.369 ◽

1960 ◽

Vol 8 (4) ◽

pp. 369-375

Author(s):

P. H. S. Silver

Keyword(s):

Chick Embryo ◽

Short Term ◽

Stages Of Development ◽

In Ovo ◽

Technical Problems ◽

Early Chick Embryo ◽

In Vitro Methods ◽

Early Stages

It seems to be generally accepted that experimenting in ovo on the chick during the early stages of development (up to about 48 hours) is fraught with the greatest difficulty. After about this time no serious technical problems arise and a high proportion of successful results can be expected. It is natural to ask why there should be this change-over from extreme difficulty to reasonable simplicity. New (1955) attributed to this ‘inaccessibility of the chick embryo in the egg’ the invention of his own and many other in vitro methods during the last 30 years. There is no doubt that, when short-term experiments only are required, in vitro methods will probably always be preferred. But all in vitro methods suffer from the disadvantage that the embryo cannot be expected to survive for more than 48 hours or so after explantation.

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In ovo Experiments Concerning the Eye, the Orbit, and Certain Juxta-Orbital Structures, in the Chick Embryo

Development ◽

10.1242/dev.10.3.423 ◽

1962 ◽

Vol 10 (3) ◽

pp. 423-450

Author(s):

P. H. S. Silver

Keyword(s):

Chick Embryo ◽

Gallus Domesticus ◽

Otic Capsule ◽

Present Communication ◽

In Ovo ◽

Nasal Process ◽

Optic Vesicles

The present communication concerns the influence in the chick (Gallus domesticus) of the growing eyes on various structures which form the wall of the orbits, or which lie in their vicinity. Features of the development of the trabeculae cranii, the fronto-nasal process, the nasal capsules, the maxillary ‘process’, the otic capsule, and the cephalic flexure will be described as well as certain growth relationships of the upper and lower jaws. The methods employed include (1) removal of one or both primary optic vesicles, (2) the grafting of an additional primary vesicle into the head mesenchyme so that two eyes develop in one orbit, and (3) the isolation in the coelom of various head components. The importance of the trabeculae to our present study stems from the fact that they contribute to the boundaries of the orbits on their medial sides.

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