Gamble, A., Suessmilch, M., Bonestroo, A., Merits, A., Graham, G. J. , Cavanagh, J. , Edgar, J. M. and Pingen, M. (2023) Establishing mixed neuronal and glial cell cultures from embryonic mouse brains to study infection and innate immunity. Journal of Visualized Experiments(196), 65331. (doi: 10.3791/65331) (PMID:37458443)
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Abstract
Models of the central nervous system (CNS) must recapitulate the complex network of interconnected cells found in vivo. The CNS consists primarily of neurons, astrocytes, oligodendrocytes, and microglia. Due to increasing efforts to replace and reduce animal use, a variety of in vitro cell culture systems have been developed to explore innate cell properties, which allow the development of therapeutics for CNS infections and pathologies. Whilst certain research questions can be addressed by human-based cell culture systems, such as (induced) pluripotent stem cells, working with human cells has its own limitations with regard to availability, costs, and ethics. Here, we describe a unique protocol for isolating and culturing cells from embryonic mouse brains. The resulting mixed neural cell cultures mimic several cell populations and interactions found in the brain in vivo. Compared to current equivalent methods, this protocol more closely mimics the characteristics of the brain and also garners more cells, thus allowing for more experimental conditions to be investigated from one pregnant mouse. Further, the protocol is relatively easy and highly reproducible. These cultures have been optimized for use at various scales, including 96-well based high throughput screens, 24-well microscopy analysis, and 6-well cultures for flow cytometry and reverse transcription-quantitative polymerase chain reaction (RT-qPCR) analysis. This culture method is a powerful tool to investigate infection and immunity within the context of some of the complexity of the CNS with the convenience of in vitro methods.
| Item Type: | Articles |
|---|---|
| Additional Information: | This work was supported by the MS Society (grant 122) and the Yuri and Lorna Chernajovsky foundation to MP; University of Glasgow funding to JC and MP; and Wellcome Trust (217093/Z/19/Z) and Medical Research Council (MRV0109721) to GJG. |
| Keywords: | 3Rs, neuroimmunology, viral infection. |
| Status: | Published |
| Refereed: | Yes |
| Glasgow Author(s) Enlighten ID: | Cavanagh, Professor Jonathan and Edgar, Professor Julia and Suessmilch, Mrs Maria and Pingen, Dr Marieke and Bonestroo, Ms Anniek and Gamble, Mr Alistair and Graham, Professor Gerard |
| Authors: | Gamble, A., Suessmilch, M., Bonestroo, A., Merits, A., Graham, G. J., Cavanagh, J., Edgar, J. M., and Pingen, M. |
| Subjects: | Q Science > QR Microbiology > QR180 Immunology Q Science > QR Microbiology > QR355 Virology R Medicine > RC Internal medicine > RC0321 Neuroscience. Biological psychiatry. Neuropsychiatry |
| College/School: | College of Medical Veterinary and Life Sciences > School of Infection & Immunity |
| Research Centre: | College of Medical Veterinary and Life Sciences > School of Infection & Immunity > Centre for Immunobiology |
| Research Group: | Chemokine Research Group |
| Journal Name: | Journal of Visualized Experiments |
| Journal Abbr.: | JoVE |
| Publisher: | Journal of Visualized Experiments |
| ISSN: | 1940-087X |
| ISSN (Online): | 1940-087X |
| Copyright Holders: | Copyright © 2023 JOVE |
| First Published: | First published in Journal of Visualized Experiments 196:65331 |
| Publisher Policy: | Reproduced in accordance with the publisher copyright policy |
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