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Isolation of Neural Stem and Progenitor Cells from the Adult Brain and Live Imaging of Their Cell Cycle with the FUCCI System

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Cellular Quiescence

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1686))

Abstract

Neural stem cells (NSCs) enter quiescence in early embryonic stages to create a reservoir of dormant NSCs able to enter proliferation and produce neuronal precursors in the adult mammalian brain. Various approaches of fluorescent-activated cell sorting (FACS) have emerged to allow the distinction between quiescent NSCs (qNSCs), their activated counterpart (aNSCs), and the resulting progeny. In this article, we review two FACS techniques that can be used alternatively. We also show that their association with transgenic Fluorescence Ubiquitination Cell Cycle Indicator (FUCCI) mice allows an unprecedented overlook on the cell cycle dynamics of adult NSCs.

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References

  1. Luskin MB (1993) Restricted proliferation and migration of postnatally generated neurons derived from the forebrain subventricular zone. Neuron 11(1):173–189

    Article  CAS  PubMed  Google Scholar 

  2. Lois C, Alvarez-Buylla A (1993) Proliferating subventricular zone cells in the adult mammalian forebrain can differentiate into neurons and glia. Proc Natl Acad Sci U S A 90(5):2074–2077

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  3. Doetsch F, Caille I, Lim DA, Garcia-Verdugo JM, Alvarez-Buylla A (1999) Subventricular zone astrocytes are neural stem cells in the adult mammalian brain. Cell 97(6):703–716

    Article  CAS  PubMed  Google Scholar 

  4. Kaplan MS, Hinds JW (1977) Neurogenesis in the adult rat: electron microscopic analysis of light radioautographs. Science 197(4308):1092–1094

    Article  CAS  PubMed  Google Scholar 

  5. Gage FH, Kempermann G, Palmer TD, Peterson DA, Ray J (1998) Multipotent progenitor cells in the adult dentate gyrus. J Neurobiol 36(2):249–266

    Article  CAS  PubMed  Google Scholar 

  6. Ponti G, Obernier K, Guinto C, Jose L, Bonfanti L, Alvarez-Buylla A (2013) Cell cycle and lineage progression of neural progenitors in the ventricular-subventricular zones of adult mice. Proc Natl Acad Sci U S A 110(11):E1045–E1054. doi:10.1073/pnas.1219563110

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Furutachi S, Matsumoto A, Nakayama KI, Gotoh Y (2013) p57 controls adult neural stem cell quiescence and modulates the pace of lifelong neurogenesis. EMBO J 32(7):970–981. doi:10.1038/emboj.2013.50

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Fuentealba LC, Rompani SB, Parraguez JI, Obernier K, Romero R, Cepko CL, Alvarez-Buylla A (2015) Embryonic origin of postnatal neural stem cells. Cell 161(7):1644–1655. doi:10.1016/j.cell.2015.05.041

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Furutachi S, Miya H, Watanabe T, Kawai H, Yamasaki N, Harada Y, Imayoshi I, Nelson M, Nakayama KI, Hirabayashi Y, Gotoh Y (2015) Slowly dividing neural progenitors are an embryonic origin of adult neural stem cells. Nat Neurosci 18(5):657–665. doi:10.1038/nn.3989

    Article  CAS  PubMed  Google Scholar 

  10. Li G, Fang L, Fernandez G, Pleasure SJ (2013) The ventral hippocampus is the embryonic origin for adult neural stem cells in the dentate gyrus. Neuron 78(4):658–672. doi:10.1016/j.neuron.2013.03.019

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Daynac M, Chicheportiche A, Pineda JR, Gauthier LR, Boussin FD, Mouthon MA (2013) Quiescent neural stem cells exit dormancy upon alteration of GABAAR signaling following radiation damage. Stem Cell Res 11(1):516–528. doi:10.1016/j.scr.2013.02.008

    Article  CAS  PubMed  Google Scholar 

  12. Mich JK, Signer RA, Nakada D, Pineda A, Burgess RJ, Vue TY, Johnson JE, Morrison SJ (2014) Prospective identification of functionally distinct stem cells and neurosphere-initiating cells in adult mouse forebrain. Elife 3:e02669. doi:10.7554/eLife.02669

    Article  PubMed  PubMed Central  Google Scholar 

  13. Daynac M, Tirou L, Faure H, Mouthon MA, Gauthier LR, Hahn H, Boussin FD, Ruat M (2016) Hedgehog controls quiescence and activation of neural stem cells in the adult ventricular-subventricular zone. Stem Cell Reports 7:735–748. doi:10.1016/j.stemcr.2016.08.016

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Sakaue-Sawano A, Kurokawa H, Morimura T, Hanyu A, Hama H, Osawa H, Kashiwagi S, Fukami K, Miyata T, Miyoshi H, Imamura T, Ogawa M, Masai H, Miyawaki A (2008) Visualizing spatiotemporal dynamics of multicellular cell-cycle progression. Cell 132(3):487–498. doi:10.1016/j.cell.2007.12.033

    Article  CAS  PubMed  Google Scholar 

  15. Codega P, Silva-Vargas V, Paul A, Maldonado-Soto AR, Deleo AM, Pastrana E, Doetsch F (2014) Prospective identification and purification of quiescent adult neural stem cells from their in vivo niche. Neuron 82(3):545–559. doi:10.1016/j.neuron.2014.02.039

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Azari H, Rahman M, Sharififar S, Reynolds BA (2010) Isolation and expansion of the adult mouse neural stem cells using the neurosphere assay. J Vis Exp 45:2393. doi:10.3791/2393

    Google Scholar 

  17. Daynac M, Morizur L, Kortulewski T, Gauthier LR, Ruat M, Mouthon MA, Boussin FD (2015) Cell sorting of neural stem and progenitor cells from the adult mouse subventricular zone and live-imaging of their cell cycle dynamics. J Vis Exp 103. doi:10.3791/53247

  18. Daynac M, Pineda JR, Chicheportiche A, Gauthier LR, Morizur L, Boussin FD, Mouthon MA (2014) TGFbeta lengthens the G1 phase of stem cells in aged mouse brain. Stem Cells 32(12):3257–3265. doi:10.1002/stem.1815

    Article  CAS  PubMed  Google Scholar 

  19. Daynac M, Morizur L, Chicheportiche A, Mouthon MA, Boussin FD (2016) Age-related neurogenesis decline in the subventricular zone is associated with specific cell cycle regulation changes in activated neural stem cells. Sci Rep 6:21505. doi:10.1038/srep21505

    Article  CAS  PubMed  PubMed Central  Google Scholar 

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Acknowledgments

We are indebted to members of the LRP, C. Joubert, V. Neuville, V. Barroca, J. Tilliet and all the staff of animal facilities; to J. Baijer and N. Dechamps for cell sorting ; O. Etienne for technical assistance, and A. Gouret for her administrative assistance. Flow cytometry and cell sorting were carried out at the iRCM Flow Cytometry Shared Resource, established by equipment grants from DIM-Stem-Pôle, INSERM, Fondation ARC, and CEA. This work was supported by grants of Electricité de France (EDF).

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Authors and Affiliations

  1. CEA DSV iRCM SCSR; Laboratoire de Radiopathologie, INSERM; Université Paris Diderot, Sorbonne Paris Cité, Université Paris Sud, Paris, France

    Alexandra Chicheportiche, François D. Boussin & Mathieu Daynac

  2. Molecules Circuits Department, Neuroscience Paris-Saclay Institute, CNRS, Université Paris Sud, UMR9197, Paris, France

    Martial Ruat & Mathieu Daynac

  3. Department of Neurological Surgery, Helen Diller Comprehensive Cancer Research Center, University of California San Francisco, 1450 Third Street, Box 0520, San Francisco, CA, 94158-2197, USA

    Mathieu Daynac

Authors
  1. Alexandra Chicheportiche
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  2. Martial Ruat
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  3. François D. Boussin
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  4. Mathieu Daynac
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Corresponding author

Correspondence to Mathieu Daynac .

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Editors and Affiliations

  1. Texas Children’s Hospital, Baylor College of Medicine, Houston, Texas, USA

    H. Daniel Lacorazza

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Chicheportiche, A., Ruat, M., Boussin, F.D., Daynac, M. (2018). Isolation of Neural Stem and Progenitor Cells from the Adult Brain and Live Imaging of Their Cell Cycle with the FUCCI System. In: Lacorazza, H. (eds) Cellular Quiescence. Methods in Molecular Biology, vol 1686. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-7371-2_5

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  • DOI: https://doi.org/10.1007/978-1-4939-7371-2_5

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-7370-5

  • Online ISBN: 978-1-4939-7371-2

  • eBook Packages: Springer Protocols

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