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In vivo time-lapse imaging shows dynamic oligodendrocyte progenitor behavior during zebrafish development

Abstract

Myelinating oligodendrocytes arise from migratory and proliferative oligodendrocyte progenitor cells (OPCs). Complete myelination requires that oligodendrocytes be uniformly distributed and form numerous, periodically spaced membrane sheaths along the entire length of target axons. Mechanisms that determine spacing of oligodendrocytes and their myelinating processes are not known. Using in vivo time-lapse confocal microscopy, we show that zebrafish OPCs continuously extend and retract numerous filopodium-like processes as they migrate and settle into their final positions. Process remodeling and migration paths are highly variable and seem to be influenced by contact with neighboring OPCs. After laser ablation of oligodendrocyte-lineage cells, nearby OPCs divide more frequently, orient processes toward the ablated cells and migrate to fill the unoccupied space. Thus, process activity before axon wrapping might serve as a surveillance mechanism by which OPCs determine the presence or absence of nearby oligodendrocyte-lineage cells, facilitating uniform spacing of oligodendrocytes and complete myelination.

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Figure 1: OPC migration in zebrafish spinal cord.
Figure 2: OPC processes are rapidly and extensively remodeled.
Figure 3: OPC migration is variable in direction and distance.
Figure 4: OPC processes retract after contact.
Figure 5: OPCs divide and migrate to replace ablated cells.

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Acknowledgements

Thanks to B. Carter for comments on the manuscript. This work was supported by US National Institutes of Health grant NS046668, National Multiple Sclerosis Foundation grant RG 3420 and a zebrafish initiative funded by the Vanderbilt University Academic Venture Capital Fund.

Author information

Affiliations

Authors

Contributions

B.B.K. produced the migration data shown in Figure 1 and Figure 3 and the movie from which Figure 4a was obtained. N.T. produced the process activity data shown in Figure 2 and Figure 4b and the Tg(nkx2.2a:megfp) ablation data shown in Figure 5f. A.J.L. performed the Tg(olig2:egfp) ablations shown in Figure 5a–e. J.S. created the Tg(olig2:egfp) and Tg(nkx2.2:megfp) transgenic lines. T.J.C. and R.N.K. cloned and characterized the sox10 promoter fragment. B.A. supervised the experiments and wrote the manuscript.

Corresponding author

Correspondence to Bruce Appel.

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The authors declare no competing financial interests.

Supplementary information

Supplementary Video 1

Excerpt from a 36-h time-lapse of a Tg(nkx2.2a:megfp) embryo showing migratory behavior of OPCs and extension and retraction of filopodium-like processes. The images are from the side, focused on a portion of the trunk spinal cord. Dorsal is up and anterior left. EGFP expression also marks axons that descend from the hindbrain. Sequence shown begins at 46 hpf and ends at 68 hpf. Images were collected every 3 min, and the movie runs at 10 frames per second. (MOV 50688 kb)

Supplementary Video 2

Time-lapse sequence taken from the spinal cord of a Tg(nkx2.2a:megfp) embryo injected with p7.2sox10:mrfp plasmid. OPCs are green only (mEGFP+), red only (mRFP+) or yellow (mEGFP+ + mRFP+) because nkx2.2a:mEGFP expression marks a subset of OPCs, and injected DNA is distributed mosaically. Differentially labeled OPC processes interdigitate and withdraw. Images were collected every 1.5 min, and the movie runs at 5 frames per second. Dorsal is up and anterior is to the left. (MOV 24782 kb)

Supplementary Video 3

Time-lapse sequence following laser ablation of migrated EGFP+ oligodendrocyte lineage cells in hemisegments 6–10 of a 4-dpf Tg(olig2:egfp) larva. EGFP+ OPCs in adjacent hemisegments divide and migrate into region where dorsal oligodendrocyte lineage cells were ablated. Dorsal is up and anterior left. Time-lapse begins approximately 1.5 h following ablation and ends 14 h later. Images were acquired every 5 min, and the movie runs at 5 frames per second. Scale bar equals 48 μm. (MOV 11036 kb)

Supplementary Video 4

Time-lapse sequence of a 2-dpf Tg(nkx2.2a:megfp) embryo following ablation of three dorsally migrated EGFP+ OPCs. Nearby OPCs in dorsal and ventral spinal cord extend multiple processes into the ablated region and migrate into the area. Images were collected every 2 min, and the movie runs at 5 frames per second. Dorsal is up and anterior left. (MOV 22861 kb)

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Kirby, B., Takada, N., Latimer, A. et al. In vivo time-lapse imaging shows dynamic oligodendrocyte progenitor behavior during zebrafish development. Nat Neurosci 9, 1506–1511 (2006). https://doi.org/10.1038/nn1803

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