Abstract
Visualizing markers for neural stem cells (NSCs) and morphological analysis are frequently used for identification of NSCs in tissues. However, NSCs are defined as cells with the ability to both self-renew and produce descendants that can differentiate into neurons, astrocytes and oligodendrocytes. The neural colony forming cell (NCFC) assay is a single-step semisolid based assay for the identification of NSCs. In this assay, NSCs generate clonally derived colonies due to their high proliferative potential. The relative comparison of NSC populations between tissues is possible by counting the colonies obtained from the NCSC assay. Furthermore, the colonies can be isolated to establish monolayer cultures of clonal NSCs. Using clonal cultures of NSCs, it is possible to assess differentiation stage and differentiation potential of each NSC. Here, we describe a semi quantitative method for the enumeration of NSCs using the NCFC assay, with slight modification from the original protocol (Louis et al., Stem Cells 26:988–996, 2008). A method to establish monolayer culture of NSCs from a colony derived from NCFC assay is also described.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
Similar content being viewed by others
References
Altman J, Das GD (1965) Autoradiographic and histological evidence of postnatal hippocampal neurogenesis in rats. J Comp Neurol 124:319–335
Kaplan MS, Hinds JW (1977) Neurogenesis in the adult rat: electron microscopic analysis of light radioautographs. Science 197:1092–1094
Bayer SA, Yackel JW, Puri PS (1982) Neurons in the rat dentate gyrus granular layer substantially increase during juvenile and adult life. Science 216:890–892
Reynolds BA, Weiss S (1992) Generation of neurons and astrocytes from isolated cells of the adult mammalian central nervous system. Science 255:1707–1710
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 90:2074–2077
Lois C, Alvarez-Buylla A (1994) Long-distance neuronal migration in the adult mammalian brain. Science 264:1145–1148
Suh H, Consiglio A, Ray J et al (2007) In vivo fate analysis reveals the multipotent and self-renewal capacities of Sox2+ neural stem cells in the adult hippocampus. Cell Stem Cell 1:515–528
Ehninger D, Kempermann G (2008) Neurogenesis in the adult hippocampus. Cell Tissue Res 331:243–250
Lugert S, Basak O, Knuckles P et al (2010) Quiescent and active hippocampal neural stem cells with distinct morphologies respond selectively to physiological and pathological stimuli and aging. Cell Stem Cell 6:445–456
Kempermann G, Jessberger S, Steiner B et al (2004) Milestones of neuronal development in the adult hippocampus. Trends Neurosci 27:447–452
Encinas JM, Vaahtokari A, Enikolopov G (2006) Fluoxetine targets early progenitor cells in the adult brain. Proc Natl Acad Sci U S A 103:8233–8238
Takei Y (2019) Age-dependent decline in neurogenesis of the hippocampus and extracellular nucleotides. Hum Cell 32:88–94
Uda Y, Xu S, Matsumura T et al (2016) P2Y4 nucleotide receptor in neuronal precursors induces glutamatergic subtype markers in their descendant neurons. Stem Cell Rep 6:474–482
Louis SA, Rietze RL, Deleyrolle L et al (2008) Enumeration of neural stem and progenitor cells in the neural colony-forming cell assay. Stem Cells 26:988–996
Acknowledgments
This work was supported by JSPS Grant-in-Aid for Exploratory Research (Exploratory), Grant Number 18K18454.
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2022 Springer Science+Business Media, LLC, part of Springer Nature
About this protocol
Cite this protocol
Amagase, Y., Izumi-Nakaseko, H., Sugiyama, A., Takei, Y. (2022). Quantification and Clonal Culture of Neural Stem Cells from the Hippocampus of Adult Mouse. In: Kannan, N., Beer, P. (eds) Stem Cell Assays. Methods in Molecular Biology, vol 2429. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-1979-7_23
Download citation
DOI: https://doi.org/10.1007/978-1-0716-1979-7_23
Published:
Publisher Name: Humana, New York, NY
Print ISBN: 978-1-0716-1978-0
Online ISBN: 978-1-0716-1979-7
eBook Packages: Springer Protocols