Skip to main content
SpringerLink
Log in

Analysis of Axonal Regrowth and Dendritic Remodeling After Optic Nerve Crush in Adult Zebrafish

  • Protocol
  • First Online:
Axon Regeneration

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2636))

Abstract

Neurodegenerative diseases and central nervous system (CNS) injuries are frequently characterized by axonal damage, as well as dendritic pathology. In contrast to mammals, adult zebrafish show a robust regeneration capacity after CNS injury and form the ideal model organism to further unravel the underlying mechanisms for both axonal and dendritic regrowth upon CNS damage. Here, we first describe an optic nerve crush injury model in adult zebrafish, an injury paradigm that inflicts de- and regeneration of the axons of retinal ganglion cells (RGCs), but also triggers RGC dendrite disintegration and subsequent recovery in a stereotyped and timed process. Next, we outline protocols for quantifying axonal regeneration and synaptic recovery in the brain, using retro- and anterograde tracing experiments and an immunofluorescent staining for presynaptic compartments, respectively. Finally, methods to analyze RGC dendrite retraction and subsequent regrowth in the retina are delineated, using morphological measurements and immunofluorescent staining for dendritic and synaptic markers.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Log in via an institution

Subscribe and save

Springer+ Basic
$34.99 /Month
  • Get 10 units per month
  • Download Article/Chapter or eBook
  • 1 Unit = 1 Article or 1 Chapter
  • Cancel anytime
Subscribe now

Buy Now

Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 189.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 169.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 249.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

Similar content being viewed by others

References

  1. Benowitz LI, He Z, Goldberg JL (2017) Reaching the brain: advances in optic nerve regeneration. Exp Neurol 287:365–373

    Article  PubMed  Google Scholar 

  2. Berry M, Ahmed Z, Logan A (2019) Return of function after CNS axon regeneration: lessons from injury-responsive intrinsically photosensitive and alpha retinal ganglion cells. Prog Retin Eye Res 71:57–67. https://doi.org/10.1016/j.preteyeres.2018.11.006

    Article  CAS  PubMed  Google Scholar 

  3. Di Polo A (2015) Dendrite pathology and neurodegeneration: focus on mTOR. Neural Regen Res 10:559–561

    Article  PubMed  PubMed Central  Google Scholar 

  4. Johnston D, Frick A, Poolos NP (2016) Dendrites and disease. In: Stuart G, Spruston N, Häusser M (eds) Dendrites, 3rd edn. Oxford Academic, Oxford, p 677–702. https://doi.org/10.1093/acprof:oso/9780198745273.001.0001

  5. Emoto K (2011) Dendrite remodeling in development and disease. Develop Growth Differ 53:277–286

    Article  CAS  Google Scholar 

  6. Della SL, Inman DM, Lupien CB et al (2013) Differential progression of structural and functional alterations in distinct retinal ganglion cell types in a mouse model of glaucoma. J Neurosci 33:17444–17457. https://doi.org/10.1523/JNEUROSCI.5461-12.2013

    Article  CAS  Google Scholar 

  7. Frankfort BJ, Kareem Khan A, Tse DY et al (2013) Elevated intraocular pressure causes inner retinal dysfunction before cell loss in a mouse model of experimental glaucoma. Investig Ophthalmol Vis Sci 54:762–770. https://doi.org/10.1167/iovs.12-10581

    Article  Google Scholar 

  8. Morquette B, Morquette P, Agostinone J et al (2015) REDD2-mediated inhibition of mTOR promotes dendrite retraction induced by axonal injury. Cell Death Differ 22:612–625. https://doi.org/10.1038/cdd.2014.149

    Article  CAS  PubMed  Google Scholar 

  9. Peterson SL, Benowitz LI (2018) Mammalian dendritic regrowth: a new perspective on neural repair. Brain 141:1891–1894

    Article  PubMed  PubMed Central  Google Scholar 

  10. Beckers A, Moons L (2019) Dendritic shrinkage after injury: a cellular killer or a necessity for axonal regeneration? Neural Regen Res 14:1313–1316

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Bollaerts I, Veys L, Geeraerts E et al (2018) Complementary research models and methods to study axonal regeneration in the vertebrate retinofugal system. Brain Struct Funct 223:545–567

    Article  PubMed  Google Scholar 

  12. Becker T, Becker CG (2014) Axonal regeneration in zebrafish. Curr Opin Neurobiol 27:186–191

    Article  CAS  PubMed  Google Scholar 

  13. Van Houcke J, Bollaerts I, Geeraerts E et al (2017) Successful optic nerve regeneration in the senescent zebrafish despite age-related decline of cell intrinsic and extrinsic response processes. Neurobiol Aging 60:1–10. https://doi.org/10.1016/j.neurobiolaging.2017.08.013

    Article  CAS  PubMed  Google Scholar 

  14. Udvadia AJ (2008) 3.6 kb Genomic sequence from Takifugu capable of promoting axon growth-associated gene expression in developing and regenerating zebrafish neurons. Gene Expr Patterns 8:382–388. https://doi.org/10.1016/j.gep.2008.05.002

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Dhara SP, Rau A, Flister MJ et al (2019) Cellular reprogramming for successful CNS axon regeneration is driven by a temporally changing cast of transcription factors. Sci Rep 9:1–12. https://doi.org/10.1038/s41598-019-50485-6

    Article  CAS  Google Scholar 

  16. Diekmann H, Kalbhen P, Fischer D (2015) Characterization of optic nerve regeneration using transgenic zebrafish. Front Cell Neurosci 9. https://doi.org/10.3389/fncel.2015.00118

  17. Van Dyck A, Bollaerts I, Beckers A et al (2021) Müller glia–myeloid cell crosstalk accelerates optic nerve regeneration in the adult zebrafish. Glia. https://doi.org/10.1002/glia.23972

  18. Beckers A, Van Dyck A, Bollaerts I et al (2019) An antagonistic axon-dendrite interplay enables efficient neuronal repair in the adult zebrafish central nervous system. Mol Neurobiol 56:3175–3192. https://doi.org/10.1007/s12035-018-1292-5

    Article  CAS  PubMed  Google Scholar 

  19. Bollaerts I, Van Houcke J, Andries L et al (2017) Neuroinflammation as fuel for axonal regeneration in the injured vertebrate central nervous system. Mediat Inflamm 2017:9478542

    Article  Google Scholar 

  20. Lemmens K, Bollaerts I, Bhumika S et al (2016) Matrix metalloproteinases as promising regulators of axonal regrowth in the injured adult zebrafish retinotectal system. J Comp Neurol 524:1472–1493. https://doi.org/10.1002/cne.23920

    Article  CAS  PubMed  Google Scholar 

  21. Liu Q, Londraville R, Marrs JA et al (2008) Cadherin-6 function in zebrafish retinal development. Dev Neurobiol 68:1107–1122. https://doi.org/10.1002/dneu.20646

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Fox MA, Sanes JR (2007) Synaptotagmin I and II are present in distinct subsets of central synapses. J Comp Neurol 503:280–296. https://doi.org/10.1002/cne.21381

    Article  CAS  PubMed  Google Scholar 

  23. Nevin LM, Robles E, Baier H, Scott EK (2010) Focusing on optic tectum circuitry through the lens of genetics. BMC Biol 8:126

    Article  PubMed  PubMed Central  Google Scholar 

  24. Aref AA, Sayyad FE, Mwanza JC et al (2014) Diagnostic specificities of retinal nerve fiber layer, optic nerve head, and macular ganglion cell-inner plexiform layer measurements in myopic eyes. J Glaucoma. https://doi.org/10.1097/IJG.0b013e31827b155b

  25. Greenberg BM, Frohman E (2010) Optical coherence tomography as a potential readout in clinical trials. Ther Adv Neurol Disord 3:153–160

    Article  PubMed  PubMed Central  Google Scholar 

  26. Miki A, Medeiros FA, Weinreb RN et al (2014) Rates of retinal nerve fiber layer thinning in glaucoma suspect eyes. Ophthalmology. https://doi.org/10.1016/j.ophtha.2014.01.017

  27. Kim EK, Park HYL, Park CK (2017) Segmented inner plexiform layer thickness as a potential biomarker to evaluate open-angle glaucoma: dendritic degeneration of retinal ganglion cell. PLoS One. https://doi.org/10.1371/journal.pone.0182404

  28. Chua J, Tan B, Ke M et al (2020) Diagnostic ability of individual macular layers by spectral-domain optical coherence tomography in different stages of glaucoma. Ophthalmol Glaucoma. https://doi.org/10.1016/j.ogla.2020.04.003

  29. Riederer B, Matus A (1985) Differential expression of distinct microtubule-associated proteins during brain development. Proc Natl Acad Sci U S A 82:6006–6009. https://doi.org/10.1073/pnas.82.17.6006

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. Lieven CJ, Millet LE, Hoegger MJ, Levin LA (2007) Induction of axon and dendrite formation during early RGC-5 cell differentiation. Exp Eye Res 85:678–683. https://doi.org/10.1016/j.exer.2007.08.001

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. Kim Y, Jang YN, Kim JY et al (2020) Microtubule-associated protein 2 mediates induction of long-term potentiation in hippocampal neurons. FASEB J 34:6965–6983. https://doi.org/10.1096/fj.201902122RR

    Article  CAS  PubMed  Google Scholar 

  32. Cho T, Kashiwagi Y, Okabe S (2019) Temporal sequences of synapse disintegration triggered by afferent axon transection, time-lapse imaging study of presynaptic and postsynaptic molecules. eNeuro 6. https://doi.org/10.1523/ENEURO.0459-18.2019

  33. Meyer MP, Trimmer JS, Gilthorpe JD, Smith SJ (2005) Characterization of zebrafish PSD-95 gene family members. J Neurobiol 63:91–105. https://doi.org/10.1002/neu.20118

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgments

We thank present and former lab members of the Neural Circuit Development and Regeneration (NCDR) group, especially the ones forming the zebrafish research team, who have helped to develop these techniques over the years. This work was financially supported by fellowships and project grants from the Research Foundation-Flanders (FWO), FWO Flanders-Quebec bilateral research grant, the KU Leuven Research Council, and L’Oréal/UNESCO (For Women in Science).

Author information

Authors and Affiliations

  1. Neural Circuit Development and Regeneration Research Group, Animal Physiology and Neurobiology Section, Department of Biology, Leuven Brain Institute, KU Leuven, Leuven, Belgium

    An Beckers, Steven Bergmans, Annelies Van Dyck & Lieve Moons

Authors
  1. An Beckers
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Steven Bergmans
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Annelies Van Dyck
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Lieve Moons
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Lieve Moons .

Editor information

Editors and Affiliations

  1. Department of Biology, Appalachian State University, Boone, NC, USA

    Ava J. Udvadia

  2. Office of Technology Development, Medical College of Wisconsin, Milwaukee, WI, USA

    James B. Antczak

Rights and permissions

Reprints and permissions

Copyright information

© 2023 The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature

About this protocol

Check for updates. Verify currency and authenticity via CrossMark

Cite this protocol

Beckers, A., Bergmans, S., Van Dyck, A., Moons, L. (2023). Analysis of Axonal Regrowth and Dendritic Remodeling After Optic Nerve Crush in Adult Zebrafish. In: Udvadia, A.J., Antczak, J.B. (eds) Axon Regeneration. Methods in Molecular Biology, vol 2636. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-3012-9_9

Download citation

  • DOI: https://doi.org/10.1007/978-1-0716-3012-9_9

  • Published:

  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-3011-2

  • Online ISBN: 978-1-0716-3012-9

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics

Search

Navigation