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Open Access Research article

Midline crossing is not required for subsequent pathfinding decisions in commissural neurons

Jennifer Bonner1*, Michael Letko1, Oliver Brant Nikolaus2, Lisa Krug1, Alexandria Cooper1, Benjamin Chadwick1, Phoebe Conklin1, Amy Lim2, Chi-Bin Chien2 and Richard I Dorsky2

Author Affiliations

1 Biology Department, Skidmore College, Saratoga Springs, NY, 12866, USA

2 Department of Neurobiology and Anatomy, University of Utah School of Medicine, Salt Lake City, UT, 84132, USA

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Neural Development 2012, 7:18  doi:10.1186/1749-8104-7-18

Published: 6 June 2012

Abstract

Background

Growth cone navigation across the vertebrate midline is critical in the establishment of nervous system connectivity. While midline crossing is achieved through coordinated signaling of attractive and repulsive cues, this has never been demonstrated at the single cell level. Further, though growth cone responsiveness to guidance cues changes after crossing the midline, it is unclear whether midline crossing itself is required for subsequent guidance decisions in vivo. In the zebrafish, spinal commissures are initially formed by a pioneer neuron called CoPA (

    Co
mmissural
    P
rimary
    A
scending). Unlike in other vertebrate models, CoPA navigates the midline alone, allowing for single-cell analysis of axon guidance mechanisms.

Results

We provide evidence that CoPA expresses the known axon guidance receptors dcc, robo3 and robo2. Using loss of function mutants and gene knockdown, we show that the functions of these genes are evolutionarily conserved in teleosts and that they are used consecutively by CoPA neurons. We also reveal novel roles for robo2 and robo3 in maintaining commissure structure. When midline crossing is prevented in robo3 mutants and dcc gene knockdown, ipsilaterally projecting neurons respond to postcrossing guidance cues. Furthermore, DCC inhibits Robo2 function before midline crossing to allow a midline approach and crossing.

Conclusions

Our results demonstrate that midline crossing is not required for subsequent guidance decisions by pioneer axons and that this is due, in part, to DCC inhibition of Robo2 function prior to midline crossing.

Keywords:
Robo; DCC; Zebrafish; Midline crossing; Axon guidance