Chicken Embryo Spinal Cord Slice Culture Protocol

Tubby, Kristina C.; Norval, Dee; Price, Stephen R.

doi:10.3791/50295

Cited by 2 publications

(5 citation statements)

References 22 publications

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“…Here, we established a method combining in ovo electroporation and the culture of organotypic tissue slices to study exogenous gene functions in the CNS during chicken embryo development. Some of these methods have been established for the CNS tissue slice culture during chicken embryo development . Compared with the established methods, we have combined in ovo electroporation with tissue slice culture to study the function of exogenous genes in the CNS during chicken embryo development.…”

Section: Discussionmentioning

confidence: 99%

“…Some of these methods have been established for the CNS tissue slice culture during chicken embryo development. 18,19 Compared with the established methods, we have combined in ovo electroporation with tissue slice culture to study the function of exogenous genes in the CNS during chicken embryo development.…”

Section: Comparison Of Iba1 and Gfap Expression In Vivo And In Cultur...mentioning

confidence: 99%

“…Today, organotypic brain slice culture is mainly applied to mouse and rat tissue, as well as for disease modelling, such as with transgenic mice . However, there are few reports related to the study of organotypic slice culture using chicken embryos . The chicken embryo is a good animal model, used for both developmental biology and neurobiology studies.…”

Section: Introductionmentioning

confidence: 99%

“…[10][11][12][13][14][15][16][17] However, there are few reports related to the study of organotypic slice culture using chicken embryos. 18,19 The chicken embryo is a good animal model, used for both developmental biology and neurobiology studies. Particularly for the study of embryonic CNS development, the chicken embryo has the advantages of being abundant, easy to manipulate and easy to access to collect material from.…”

Section: Introductionmentioning

confidence: 99%

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Organotypic slice culture based on in ovo electroporation for chicken embryonic central nervous system

Yang

et al. 2018

J Cellular Molecular Medi

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Organotypic slice culture is a living cell research technique which blends features of both in vivo and in vitro techniques. While organotypic brain slice culture techniques have been well established in rodents, there are few reports on the study of organotypic slice culture, especially of the central nervous system (CNS), in chicken embryos. We established a combined in ovo electroporation and organotypic slice culture method to study exogenous genes functions in the CNS during chicken embryo development. We performed in ovo electroporation in the spinal cord or optic tectum prior to slice culture. When embryonic development reached a specific stage, green fluorescent protein (GFP)‐positive embryos were selected and fluorescent expression sites were cut under stereo fluorescence microscopy. Selected tissues were embedded in 4% agar. Tissues were sectioned on a vibratory microtome and 300 μm thick sections were mounted on a membrane of millicell cell culture insert. The insert was placed in a 30‐mm culture dish and 1 ml of slice culture media was added. We show that during serum‐free medium culture, the slice loses its original structure and propensity to be strictly regulated, which are the characteristics of the CNS. However, after adding serum, the histological structure of cultured‐tissue slices was able to be well maintained and neuronal axons were significantly longer than that those of serum‐free medium cultured‐tissue slices. As the structure of a complete single neuron can be observed from a slice culture, this is a suitable way of studying single neuronal dynamics. As such, we present an effective method to study axon formation and migration of single neurons in vitro.

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Section: Discussionmentioning

confidence: 99%

Section: Comparison Of Iba1 and Gfap Expression In Vivo And In Cultur...mentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Organotypic slice culture based on in ovo electroporation for chicken embryonic central nervous system

Yang

et al. 2018

J Cellular Molecular Medi

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“…Tubby et al ( 2013 ) recently established a slice culture protocol to examine motor neuron development in embryonic chick spinal cord. Using this method they found conserved transcription factor domains, normal motor neuron survival rates and the migration of motor neurons to appropriate positions in the spinal cord after 24 h in culture.…”

Section: Introductionmentioning

confidence: 99%

A method to investigate radial glia cell behavior using two-photon time-lapse microscopy in an ex vivo model of spinal cord development

Pakan

McDermott

2014

Front. Neuroanat.

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The mammalian central nervous system (CNS) develops from multipotent progenitor cells, which proliferate and differentiate into the various cell types of the brain and spinal cord. Despite the wealth of knowledge from progenitor cell culture studies, there is a significant lack of understanding regarding dynamic progenitor cell behavior over the course of development. This is in part due to shortcomings in the techniques available to study these processes in living tissues as they are occurring. In order to investigate cell behavior under physiologically relevant conditions we established an ex vivo model of the developing rat spinal cord. This method allows us to directly observe specific populations of cells ex vivo in real time and over extended developmental periods as they undergo proliferation, migration, and differentiation in the CNS. Previous investigations of progenitor cell behavior have been limited in either spatial or temporal resolution (or both) due to the necessity of preserving tissue viability and avoiding phototoxic effects of fluorescent imaging. The method described here overcomes these obstacles. Using two-photon and confocal microscopy and transfected organotypic spinal cord slice cultures we have undertaken detailed imaging of a unique population of neural progenitors, radial glial cells. This method uniquely enables analysis of large populations as well as individual cells; ultimately resulting in a 4D dataset of progenitor cell behavior for up to 7 days during embryonic development. This approach can be adapted to study a variety of cell populations at different stages of development using appropriate promoter driven fluorescent protein expression. The ability to control the tissue micro-environment makes this ex vivo method a powerful tool to elucidate the underlying molecular mechanisms regulating cell behavior during embryonic development.

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Chicken Embryo Spinal Cord Slice Culture Protocol

Cited by 2 publications

References 22 publications

Organotypic slice culture based on in ovo electroporation for chicken embryonic central nervous system

Organotypic slice culture based on in ovo electroporation for chicken embryonic central nervous system

A method to investigate radial glia cell behavior using two-photon time-lapse microscopy in an ex vivo model of spinal cord development

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