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Optimized culture of retinal ganglion cells and amacrine cells from adult mice

Yong H Park, Joshua D Snook, Iris Zhuang, Guofu Shen, View ORCID ProfileBenjamin J Frankfort
doi: https://doi.org/10.1101/2020.06.16.155069
Yong H Park
1Department of Ophthalmology, Baylor College of Medicine, Houston, Texas, United States
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Joshua D Snook
1Department of Ophthalmology, Baylor College of Medicine, Houston, Texas, United States
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Iris Zhuang
1Department of Ophthalmology, Baylor College of Medicine, Houston, Texas, United States
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Guofu Shen
1Department of Ophthalmology, Baylor College of Medicine, Houston, Texas, United States
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Benjamin J Frankfort
1Department of Ophthalmology, Baylor College of Medicine, Houston, Texas, United States
2Department of Neuroscience, Baylor College of Medicine, Houston, Texas, United States
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  • ORCID record for Benjamin J Frankfort
  • For correspondence: benjamin.frankfort@bcm.edu
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Abstract

Cell culture is widely utilized to study the cellular and molecular biology of different neuronal cell populations. Current techniques to study enriched neurons in vitro are primarily limited to embryonic/neonatal animals and induced pluripotent stem cells (iPSC). Although the use of these cultures is valuable, the accessibility of purified primary adult neuronal cultures would allow for improved assessment of certain neurological diseases and pathways at the cellular level. Using a modified 7-step immunopanning technique to isolate for retinal ganglion cells (RGCs) and amacrine cells (ACs) from adult mouse retinas, we have successfully developed a model of neuronal culture that maintains for at least one week. Isolations of Thy1.2+ cells are enriched for RGCs, with the isolation cell yield being congruent to the theoretical yield of RGCs in a mouse retina. ACs of two different populations (CD15+ and CD57+) can also be isolated. The populations of these three adult neurons in culture are healthy, with neurite outgrowths in some cases greater than 500µm in length. Optimization of culture conditions for RGCs and CD15+ cells revealed that neuronal survival and the likelihood of neurite outgrowth respond inversely to different culture media. Serially diluted concentrations of puromycin decreased cultured adult RGCs in a dose-dependent manner, demonstrating the potential usefulness of these adult neuronal cultures in screening assays. This novel culture system can be used to model adult neurons in vivo. Studies can now be expanded in conjunction with other methodologies to study the neurobiology of function, aging, and diseases.

Competing Interest Statement

The authors have declared no competing interest.

Copyright 
The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under a CC-BY 4.0 International license.
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Posted June 18, 2020.
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Optimized culture of retinal ganglion cells and amacrine cells from adult mice
Yong H Park, Joshua D Snook, Iris Zhuang, Guofu Shen, Benjamin J Frankfort
bioRxiv 2020.06.16.155069; doi: https://doi.org/10.1101/2020.06.16.155069
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Optimized culture of retinal ganglion cells and amacrine cells from adult mice
Yong H Park, Joshua D Snook, Iris Zhuang, Guofu Shen, Benjamin J Frankfort
bioRxiv 2020.06.16.155069; doi: https://doi.org/10.1101/2020.06.16.155069

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