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Isolation of multipotent neural stem or progenitor cells from both the dentate gyrus and subventricular zone of a single adult mouse

Abstract

In adult mammals, the subventricular zone (SVZ) of the lateral ventricles and the subgranular zone of the dentate gyrus (DG) show ongoing neurogenesis, and multipotent neural stem or progenitor cells (NSCs) in these two regions exhibit different intrinsic properties. However, investigation of the mechanisms underlying such differences has been limited by a lack of efficient methods for isolating NSCs, particularly from the adult DG. Here we describe a protocol that enables us to isolate self-renewing and multipotent NSCs from the SVZ and the DG of the same adult mouse. The protocol involves the microdissection of the SVZ and DG from one adult mouse brain, isolation of NSCs from specific regions and cultivation of NSCs in vitro. The entire procedure takes 2–3 h. As only one mouse is needed for each cell isolation procedure, this protocol will be particularly useful for studies with limited availability of mice, such as mice that contain multiple genetic modifications.

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Figure 1: A schematic overview of the major steps in this protocol.
Figure 2: Microdissection of the SVZ and the DG of a single adult mouse.
Figure 3: Microdissection of the SVZ and the DG from the brain of an adult nestin-GFP transgenic mouse.
Figure 4: Representative images of neurospheres.

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Acknowledgements

We thank C.T. Strauss for editing the manuscript, and Y. Xing, E. Polich, W. Gilbert and I.J. Bhuiyan for technical support. This work was supported by grants from the US National Institutes of Health (NIH) to X.Z. (RO1MH080434 and RO1MH078972), from the International Rett Syndrome Foundation to X.Z. and a center grant from the NIH to the Waisman Center (P30HD03352). W.G. is funded by a postdoctoral fellowship from University of Wisconsin Center for Stem Cells and Regenerative medicine. E.M.J. is funded by a NIH Molecular Biosciences Training grant (MBTG: T32 GM07215), and N.E.P. is funded by a NIH Molecular and Cellular Pharmacology Training grant (T32 GM008688).

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Authors and Affiliations

Authors

Contributions

W.G. and X.Z. developed the methodology. W.G., N.E.P. and E.M.J. followed the protocol to isolate cells from a nestin-GFP mouse. W.G. and X.Z. wrote the protocol.

Corresponding author

Correspondence to Xinyu Zhao.

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The authors declare no competing financial interests.

Supplementary information

Supplementary Video 1

Dissection of the SVZ (AVI 2801 kb)

Supplementary Video 2

Dissection of the DG (AVI 2675 kb)

Supplementary Figure 1

DG NSCs isolated using this protocol exhibit basic neural stem cell properties in vitro. (PDF 166 kb)

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Guo, W., Patzlaff, N., Jobe, E. et al. Isolation of multipotent neural stem or progenitor cells from both the dentate gyrus and subventricular zone of a single adult mouse. Nat Protoc 7, 2005–2012 (2012). https://doi.org/10.1038/nprot.2012.123

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