|
|
|
|
|
|
|
||||
| Home Help Feedback Subscriptions Archive Search Table of Contents | ||||
|
Fig. 2. Neutralizing endogenous BDNF influences presynaptic sites in individual RGC
arbors. Time-lapse confocal imaging of individual DsRed2-labeled RGC axons
expressing GFP-synaptobrevin illustrates the effects of neutralizing
endogenous tectal BDNF on synapse number and axon arbor morphology. (A) Image
reconstructions of a RGC axon in a vehicle-treated (control) tadpole show the
localization of GFP-synaptobrevin clusters (yellow) within specific regions of
the arborizing, DsRed2-labeled axons (red). (B) The number and distribution of
GFP-synaptobrevin clusters was significantly altered in RGC arbors in tadpoles
that received a single injection of anti-BDNF following the first imaging
session (0h). Anti-BDNF not only influences axon arbor complexity but also
decreases the number and density of GFP-synaptobrevin clusters per axon arbor.
(C) Magnified region of the arbor shown in B illustrates the localization of
GFP-synaptobrevin clusters to branch points and branch termini, and their
disappearance after anti-BDNF treatment. By separating the green component
(middle panel, GFP fluorescence) from the red component (overlay DsRed2 and
GFP fluorescence; top panel) one can clearly distinguish specific
GFP-synaptobrevin puncta from the background fluorescence signal. The line
scans (bottom panels) obtained from raw confocal data show the intensity of
the DsRed2 and GFP-synaptobrevin signals at the level of the axon branch
demarcated by the light-blue hairlines (top panels). The green arrowheads
(middle panels) indicate sites containing GFP-synaptobrevin clusters that are
crossed by the line scan. In the 0 and 4 hour images, the proximal part of the
line scan (1 pixel width) travels near GFP-synaptobrevin puncta but only
crosses the arbor area where background fluorescent signal is observed.
Background fluorescence intensity values remain similar after repeated imaging
and that fluorescence intensity values of specific GFP-synaptobrevin puncta
are at least twice as great as those of background signals. Scale bar: 20
µm in A,B; 10 µm in C. Posterior is upwards, anterior is downwards.
|
