a) Top: Image of whole Tg(olig1:memEYFP) transgenic animal at 5 days
post-fertilization (dpf). Scale bar: 1mm. Bottom: confocal image of a
Tg(olig1:memEYFP), Tg(olig1:nls-mApple) zebrafish at the level of the spinal
cord at 5 dpf. Scale bar: 50 μm. Representative images from 4 animals in
2 independent experiments.
b) Cross-sectional view of the spinal cord showing the distribution
of OPC processes in Tg(olig1:memEYFP) at 7 dpf (n=33 animals / 11 experiments).
Scale bar: 10 μm.
c) Cross-sectional view of the spinal cord showing the distribution
of axons and dendrites at 7 dpf (anti-acetylated tubulin and anti-MAP2; n=7
animals / 2 experiments). Scale bar: 10 μm.
d) Top: sparse labelling of olig1:memEYFP expressing OPCs at 4 dpf.
Bottom: Tracing of two neighbouring examplary OPCs and the spinal cord outlines
(n numbers in panel g). Dotted lines indicate the position of the y-axis
rotations shown in e.
e) 90° y-axis rotations at the level of the soma of each of
the two cells shown in panel d with a BODIPY counterstain to reveal the position
of OPC somata in axo-dendritic (cell #1) and neuron-rich (cell#2) regions (n=12
BODIPY stained animals / 4 experiments). Dotted lines indicate axo-dendritic
areas. Scale bar: 10 μm.
f) High-magnification view showing the proximity of the processes
made by the two OPCs shown in panel d within axo-dendritic areas. Quantification
shows the percentage of cell processes resident in axo-dendritic areas formed by
OPCs with their soma in neuron-rich and axo-dendritic regions at 4-5 dpf (mean
91.3% ± 2.3 SD neuron-rich vs. 98.9% ± 0.7 SD
axo-dendritic; n=5 cells/condition from 6 animals).
g) Morphology reconstructions of the two OPCs shown in panels d-f.
Quantification shows relative cell complexities of individual OPCs at 4 dpf with
their soma in different areas. Triangles indicate example cell#1 and #2. Data
are expressed as median ± 25/75% interquartile percentiles, whiskers
indicate min/max values (2.1±1.5/3.2 for OPCs in neuron-rich areas
vs. 0.6±0.4/1.5 in axo-dendritic areas,
p<0.001 (Mann-Whitney U test, U=211; n=36/38 cells from 23 animals in 11
experiments).
h) Projections of 60 minutes time-lapse imaging to show remodelled
and stable processes of OPCs with their soma in different areas. Quantification
shows stable processes over time. Dashed lines connect data points of individual
cells. Data points connected by continuous lines represent mean ± SD
within the groups. At t=60 minutes: 56.3±12.2 % stable processes for OPCs
in neuron-rich areas vs. 25.0±5.5 % in axo-dendritic
areas; p=0.001 between groups (two-way repeated-measures ANOVA of time-points
0-60 min, F(1,6)=32); n=4 cells per group from 8 animals in 8 experiments).
Scale bar: 20 μm.
i) Schematic overview depicting the positioning of OPCs in the
zebrafish spinal cord.