Skip to main content

Neural Patterning and CNS Functions of Wnt in Zebrafish

  • Protocol
Book cover Wnt Signaling

Part of the book series: Methods in Molecular Biology ((MIMB,volume 469))

Abstract

Wnt signaling plays several important roles in the development of the zebrafish central nervous system (CNS). This chapter outlines both the known and postulated roles of Wnts from the earliest step of neural plate induction to relatively late events such as axon pathfinding and synaptogenesis. The common tools useful for examining Wnt function and nervous system development in zebrafish are first reviewed. Examples are then provided for specific phenotypes resulting from gain and loss of Wnt activity at multiple developmental stages. Finally, specific assays and reagents that can be used to investigate the function of novel Wnt pathway components in CNS development are listed.

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

Access this chapter

Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 84.99
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 109.99
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

References

  1. Wilkinson, D. G., Bailes, J. A., McMa-hon, A. P. (1987) Expression of the proto-oncogene int-1 is restricted to specific neural cells in the developing mouse embryo. Cell 50, 79–88.

    Article  PubMed  CAS  Google Scholar 

  2. McMahon, A. P., Bradley, A. (1990) The Wnt-1 (int-1) proto-oncogene is required for development of a large region of the mouse brain. Cell 62, 1073–1085.

    Article  PubMed  CAS  Google Scholar 

  3. Amsterdam, A., Hopkins, N. (2006) Muta-genesis strategies in zebrafish for identifying genes involved in development and disease. Trends Genet 22, 473–478.

    Article  PubMed  CAS  Google Scholar 

  4. Fritz, A., Rozowski, M., Walker, C., et al. (1996) Identification of selected gamma-ray induced deficiencies in zebrafish using multiplex polymerase chain reaction. Genetics 144, 1735–1745.

    PubMed  CAS  Google Scholar 

  5. Amsterdam, A. (2003) Insertional mutagen-esis in zebrafish. Dev Dyn 228, 523–534.

    Article  PubMed  CAS  Google Scholar 

  6. Clark, K. J., Geurts, A. M., Bell, J. B., et al. (2004) Transposon vectors for gene-trap insertional mutagenesis in vertebrates. Genesis 39, 225–233.

    Article  PubMed  CAS  Google Scholar 

  7. Kawakami, K. (2005) Transposon tools and methods in zebrafish. Dev Dyn 234, 244–254.

    Article  PubMed  CAS  Google Scholar 

  8. Nasevicius, A., Ekker, S. C. (2000) Effective targeted gene ‘ knockdown ’ in zebrafish. Nat Genet 26, 216–220.

    Article  PubMed  CAS  Google Scholar 

  9. Helde, K. A., Wilson, E. T., Cretekos, C. J., et al. (1994) Contribution of early cells to the fate map of the zebrafish gastrula. Science 265, 517–520.

    Article  PubMed  CAS  Google Scholar 

  10. Woo, K., Fraser, S. E. (1995) Order and coherence in the fate map of the zebrafish nervous system. Development 121, 2595–2609.

    PubMed  CAS  Google Scholar 

  11. Amsterdam, A., Becker, T. S. (2005) Trans-genes as screening tools to probe and manipulate the zebrafish genome. Dev Dyn 234, 255–268.

    Article  PubMed  CAS  Google Scholar 

  12. Jowett, T. (2001) Double in situ hybridization techniques in zebrafish. Methods 23, 345–358.

    Article  PubMed  CAS  Google Scholar 

  13. Oxtoby, E., Jowett, T. (1993) Cloning of the zebrafish krox-20 gene (krx-20) and its expression during hindbrain development. Nucleic Acids Res 21, 1087–1095.

    Article  PubMed  CAS  Google Scholar 

  14. Lee, J. E., Wu, S. F., Goering, L. M., et al. (2006) Canonical Wnt signaling through Lef1 is required for hypothalamic neurogen-esis. Development 133, 4451–4461.

    Article  PubMed  CAS  Google Scholar 

  15. Thorpe, C. J., Weidinger, G., Moon, R. T. (2005) Wnt/beta-catenin regulation of the Sp1-related transcription factor sp5l promotes tail development in zebrafish. Development 132, 1763–1772.

    Article  PubMed  CAS  Google Scholar 

  16. Schneider, S., Steinbeisser, H., Warga, R. M., et al. (1996) Beta-catenin translocation into nuclei demarcates the dorsalizing centers in frog and fish embryos. Mech Dev 57, 191–198.

    Article  PubMed  CAS  Google Scholar 

  17. Bellipanni, G., Varga, M., Maegawa, S., et al. (2006) Essential and opposing roles of zebrafish beta-catenins in the formation of dorsal axial structures and neurectoderm. Development 133, 1299–1309.

    Article  PubMed  CAS  Google Scholar 

  18. Kelly, C., Chin, A. J., Leatherman, J. L., et al. (2000) Maternally controlled (beta)-catenin-mediated signaling is required for organizer formation in the zebrafish. Development 127, 3899–3911.

    Google Scholar 

  19. Westfall, T. A., Brimeyer, R., Twedt, J., et al. (2003) Wnt-5/pipetail functions in vertebrate axis formation as a negative regulator of Wnt/beta-catenin activity. J Cell Biol 162, 889–898.

    Article  PubMed  CAS  Google Scholar 

  20. Leung, T., Soll, I., Arnold, S. J., et al. (2003) Direct binding of Lef1 to sites in the boz promoter may mediate pre-midblastula-transition activation of boz expression. Dev Dyn 228, 424–432.

    Article  PubMed  CAS  Google Scholar 

  21. Ryu, S. L., Fujii, R., Yamanaka, Y., et al. (2001) Regulation of dharma/bozozok by the Wnt pathway. Dev Biol 231, 397–409.

    Article  PubMed  CAS  Google Scholar 

  22. Dorsky, R. I., Sheldahl, L. C., Moon, R. T. (2002) A transgenic Lef1/beta-catenin-dependent reporter is expressed in spatially restricted domains throughout zebrafish development. Dev Biol 241, 229–237.

    Article  PubMed  CAS  Google Scholar 

  23. Shimizu, T., Yamanaka, Y., Ryu, S. L., et al. (2000) Cooperative roles of Bozo-zok/Dharma and Nodal-related proteins in the formation of the dorsal organizer in zebrafish. Mech Dev 91, 293–303.

    Article  PubMed  CAS  Google Scholar 

  24. Okuda, Y., Yoda, H., Uchikawa, M., et al. (2006) Comparative genomic and expression analysis of group B1 sox genes in zebrafish indicates their diversification during vertebrate evolution. Dev Dyn 235, 811–825.

    Article  PubMed  CAS  Google Scholar 

  25. Lekven, A. C., Thorpe, C. J., Waxman, J. S., et al. (2001) Zebrafish wnt8 encodes two wnt8 proteins on a bicistronic transcript and is required for mesoderm and neurectoderm patterning. Dev Cell 1, 103–114.

    Article  PubMed  CAS  Google Scholar 

  26. Erter, C. E., Wilm, T. P., Basler, N., et al. (2001) Wnt8 is required in lateral mesen-dodermal precursors for neural posterioriza-tion in vivo. Development 128, 3571–3583.

    PubMed  CAS  Google Scholar 

  27. Dorsky, R. I., Itoh, M., Moon, R. T., et al. (2003) Two tcf3 genes cooperate to pattern the zebrafish brain. Development 130, 1937–1947.

    Article  PubMed  CAS  Google Scholar 

  28. Kapsimali, M., Caneparo, L., Houart, C., et al. (2004) Inhibition of Wnt/Axin/beta-catenin pathway activity promotes ventral CNS midline tissue to adopt hypothalamic rather than floorplate identity. Development 131, 5923–5933.

    Article  PubMed  CAS  Google Scholar 

  29. Kim, C. H., Oda, T., Itoh, M., et al. (2000) Repressor activity of Headless/Tcf3 is essential for vertebrate head formation. Nature 407, 913–916.

    Article  PubMed  CAS  Google Scholar 

  30. van de Water, S., van de Wetering, M., Joore, J., et al. (2001) Ectopic Wnt signal determines the eyeless phenotype of zebrafish masterblind mutant. Development 128, 3877–3888.

    PubMed  Google Scholar 

  31. Heisenberg, C. P., Houart, C., Take-Uchi, M., et al. (2001) A mutation in the Gsk3-bind-ing domain of zebrafish Masterblind/Axin1 leads to a fate transformation of telen-cephalon and eyes to diencephalon. Genes Dev 15, 1427–1434.

    Article  PubMed  CAS  Google Scholar 

  32. Kilian, B., Mansukoski, H., Barbosa, F. C., et al. (2003) The role of Ppt/Wnt5 in regulating cell shape and movement during zebrafish gastrulation. Mech Dev 120, 467–476.

    Article  PubMed  CAS  Google Scholar 

  33. Heisenberg, C. P., Tada, M., Rauch, G. J., et al. (2000) Silberblick/Wnt11 mediates convergent extension movements during zebrafish gastrulation. Nature 405, 76–81.

    Article  PubMed  CAS  Google Scholar 

  34. Jessen, J. R., Topczewski, J., Bingham, S., et al. (2002) Zebrafish trilobite identifies new roles for Strabismus in gastrulation and neuronal movements. Nat Cell Biol 4, 610–615.

    PubMed  CAS  Google Scholar 

  35. Park, M., Moon, R. T. (2002) The planar cell-polarity gene stbm regulates cell behaviour and cell fate in vertebrate embryos. Nat Cell Biol 4, 20–25.

    Article  PubMed  CAS  Google Scholar 

  36. Topczewski, J., Sepich, D. S., Myers, D. C., et al. (2001) The zebrafish glypican knypek controls cell polarity during gastrulation movements of convergent extension. Dev Cell 1, 251–264.

    Article  PubMed  CAS  Google Scholar 

  37. Veeman, M. T., Slusarski, D. C., Kaykas, A., et al. (2003) Zebrafish prickle, a modulator of noncanonical Wnt/Fz signaling, regulates gastrulation movements. Curr Biol 13, 680–685.

    Article  PubMed  CAS  Google Scholar 

  38. Ciruna, B., Jenny, A., Lee, D., et al. (2006) Planar cell polarity signalling couples cell division and morphogenesis during neurula-tion. Nature 439, 220–224.

    Article  PubMed  CAS  Google Scholar 

  39. Bingham, S., Higashijima, S., Okamoto, H., et al. (2002) The Zebrafish trilobite gene is essential for tangential migration of branchi-omotor neurons. Dev Biol 242, 149–160.

    Article  PubMed  CAS  Google Scholar 

  40. Lin, F., Sepich, D. S., Chen, S., et al. (2005) Essential roles of G{alpha}12/13 signaling in distinct cell behaviors driving zebrafish convergence and extension gastrulation movements. J Cell Biol 169, 777–787.

    Article  PubMed  CAS  Google Scholar 

  41. Sepich, D. S., Solnica-Krezel, L. (2005) Analysis of cell movements in zebrafish embryos: morphometrics and measuring movement of labeled cell populations in vivo. Methods Mol Biol 294, 211–233.

    PubMed  Google Scholar 

  42. Higashijima, S., Hotta, Y., Okamoto, H. (2000) Visualization of cranial motor neurons in live transgenic zebrafish expressing green fluorescent protein under the control of the islet-1 promoter/enhancer. J Neuro-sci20, 206–218.

    CAS  Google Scholar 

  43. Lekven, A. C., Buckles, G. R., Kostakis, N., et al. (2003) Wnt1 and wnt10b function redundantly at the zebrafish midbrain-hindbrain boundary. Dev Biol254, 172–187.

    Article  PubMed  CAS  Google Scholar 

  44. Buckles, G. R., Thorpe, C. J., Ramel, M. C., et al. (2004) Combinatorial Wnt control of zebrafish midbrain-hindbrain boundary formation. Mech Dev 121, 437–447.

    Article  PubMed  CAS  Google Scholar 

  45. Amoyel, M., Cheng, Y. C., Jiang, Y. J., et al. (2005) Wnt1 regulates neurogenesis and mediates lateral inhibition of boundary cell specification in the zebrafish hindbrain. Development 132, 775–785.

    Article  PubMed  CAS  Google Scholar 

  46. Riley, B. B., Chiang, M. Y., Storch, E. M., et al. (2004) Rhombomere boundaries are Wnt signaling centers that regulate meta-meric patterning in the zebrafish hindbrain. Dev Dyn 231, 278–291.

    Article  PubMed  CAS  Google Scholar 

  47. Prince, V. E., Holley, S. A., Bally-Cuif, L., et al. (2001) Zebrafish lunatic fringe demarcates segmental boundaries. Mech Dev 105, 175–180.

    Article  PubMed  CAS  Google Scholar 

  48. Chizhikov, V. V., Millen, K. J. (2005) Roof plate-dependent patterning of the vertebrate dorsal central nervous system. Dev Biol 277, 287–295.

    Article  PubMed  CAS  Google Scholar 

  49. Krauss, S., Korzh, V., Fjose, A., et al. (1992) Expression of four zebrafish wnt-related genes during embryogenesis. Development 116, 249–259.

    PubMed  CAS  Google Scholar 

  50. Cheesman, S. E., Layden, M. J., Von Ohlen, T., et al. (2004) Zebrafish and fly Nkx6 proteins have similar CNS expression patterns and regulate motoneuron formation. Development 131, 5221–5232.

    Article  PubMed  CAS  Google Scholar 

  51. Lewis, K. E., Bates, J., Eisen, J. S. (2005) Regulation of iro3 expression in the zebrafish spinal cord. Dev Dyn 232, 140–148.

    Article  PubMed  CAS  Google Scholar 

  52. Seo, H. C., Saetre, B. O., Havik, B., et al. (1998) The zebrafish Pax3 and Pax7 homo-logues are highly conserved, encode multiple isoforms and show dynamic segment-like expression in the developing brain. Mech Dev 70, 49–63.

    Article  PubMed  CAS  Google Scholar 

  53. Appel, B., Korzh, V., Glasgow, E., et al. (1995) Motoneuron fate specification revealed by patterned LIM homeobox gene expression in embryonic zebrafish. Development 121, 4117–4125.

    PubMed  CAS  Google Scholar 

  54. Higashijima, S., Masino, M. A., Mandel, G., et al. (2004) Engrailed-1 expression marks a primitive class of inhibitory spinal interneu-ron. J Neurosci 24, 5827–5839.

    Article  PubMed  CAS  Google Scholar 

  55. Kimura, Y., Okamura, Y., Higashijima, S. (2006) alx, a zebrafish homolog of Chx10, marks ipsilateral descending excitatory interneurons that participate in the regulation of spinal locomotor circuits. J Neurosci 26, 5684–5697.

    Article  PubMed  CAS  Google Scholar 

  56. Megason, S. G., McMahon, A. P. (2002) A mitogen gradient of dorsal midline Wnts organizes growth in the CNS. Development 129, 2087–2098.

    PubMed  CAS  Google Scholar 

  57. Nyholm, M. K., Wu, S. F., Dorsky, R. I., et al. (2007) The zebrafish zic2a-zic5 gene pair acts downstream of canonical Wnt signaling to control cell proliferation in the developing tectum. Development 134, 735–746.

    Article  PubMed  CAS  Google Scholar 

  58. Park, H. C., Boyce, J., Shin, J., et al. (2005) Oligodendrocyte specification in zebrafish requires notch-regulated cyclin-dependent kinase inhibitor function. J Neurosci 25, 6836–6844.

    Article  PubMed  CAS  Google Scholar 

  59. Yarden, A., Salomon, D., Geiger, B. (1995) Zebrafish cyclin D1 is differentially expressed during early embryogenesis. Biochim Biophys Acta 1264, 257–260.

    Article  PubMed  Google Scholar 

  60. Liu, Y., Shi, J., Lu, C. C., et al. (2005) Ryk-mediated Wnt repulsion regulates posterior-directed growth of corticospinal tract. Nat Neurosci 8, 1151–9.

    Article  PubMed  CAS  Google Scholar 

  61. Lu, W., Yamamoto, V., Ortega, B., et al. (2004) Mammalian Ryk is a Wnt corecep-tor required for stimulation of neurite outgrowth. Cell 119, 97–108.

    Article  PubMed  CAS  Google Scholar 

  62. Lyuksyutova, A. I., Lu, C. C., Milanesio, N., et al. (2003) Anterior-posterior guidance of commissural axons by Wnt-frizzled signaling. Science 302, 1984–1988.

    Article  PubMed  CAS  Google Scholar 

  63. Krylova, O., Herreros, J., Cleverley, K. E., et al. (2002) WNT-3, expressed by motone-urons, regulates terminal arborization of neurotrophin-3-responsive spinal sensory neurons. Neuron 35, 1043–1056.

    Article  PubMed  CAS  Google Scholar 

  64. Lucas, F. R., Salinas, P. C. (1997) WNT-7a induces axonal remodeling and increases synapsin I levels in cerebellar neurons. Dev Biol 192, 31–44.

    Article  PubMed  CAS  Google Scholar 

  65. Mueller, T., Wullimann, M. F. (2003) Anatomy of neurogenesis in the early zebrafish brain. Brain Res Dev Brain Res 140, 137–155.

    Article  PubMed  CAS  Google Scholar 

  66. Dawid, I. B., Chitnis, A. B. (2001) Lim homeobox genes and the CNS: a close relationship. Neuron 30, 301–303.

    Article  PubMed  CAS  Google Scholar 

  67. Higashijima, S., Schaefer, M., Fetcho, J. R. (2004) Neurotransmitter properties of spinal interneurons in embryonic and larval zebrafish. J Comp Neurol 480, 19–37.

    Article  PubMed  CAS  Google Scholar 

  68. Wilson, S. W., Ross, L. S., Parrett, T., et al. (1990) The development of a simple scaffold of axon tracts in the brain of the embryonic zebrafish, Brachydanio rerio. Development 108, 121–145.

    PubMed  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Editor information

Editors and Affiliations

Rights and permissions

Reprints and permissions

Copyright information

© 2008 Springer Science+Business Media, LLC

About this protocol

Cite this protocol

Dorsky, R.I. (2008). Neural Patterning and CNS Functions of Wnt in Zebrafish. In: Vincan, E. (eds) Wnt Signaling. Methods in Molecular Biology, vol 469. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-60327-469-2_20

Download citation

  • DOI: https://doi.org/10.1007/978-1-60327-469-2_20

  • Publisher Name: Humana Press, Totowa, NJ

  • Print ISBN: 978-1-60327-468-5

  • Online ISBN: 978-1-60327-469-2

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics