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Figure 8 A confirms the specificity of bouton labeling using GFP-tagged synaptophysin.
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Retrograde labelling occurred in the postsynaptic profile of 15 18% type A boutons labelled from any injection site.
In resting neurons, APP surface fluorescence was co-localized to synaptic boutons labeled with αSyt1-cypHer in only a limited number of regions (Figure 4A).
To ask whether EE induced modifications of mossy fibers and olivocerebellar axons, we examined boutons labeled by anti-v-glut2 antibodies.
We found that the changes in GLR-1 puncta observed in stimulated tm446 mutants were not due to an overall decrease in the number or size of synapses, as the number of presynaptic boutons labeled with SNB-1 GFP SNB-1 GFPange (Fig. 4A,B).
Interestingly, terminations in all superficial layers (I-IIIa) were similar, as a vast majority of thalamo-cortical boutons labeled with neural tracers were also CB+, and therefore originated from the CB+ population of projection neurons in the ventral anterior nucleus (Fig. 9A C).
These terminals included some boutons which co-expressed VGLUT2 and GAD65 (Fig. 4b g, arrows) and boutons labeled for VGLUT2 only (Fig. 4b g, arrowheads).
In control animals, the distributions of neurons and boutons labeled have a small bias toward other areas preferentially responding to the same eye.
Boutons labeled with a terminal type-specific marker were selected in a single-channel image obtained from the center of the optical stack (approximately 5 μm from the surface of the section), and analysed by Adobe Photoshop CS5.
DOI: http://dx.doi.org/10.7554/eLife.03558.010 To assess the presence of mitochondria at NMJs, we counted MitoGFP positive puncta in boutons labeled by anti-Discs Large 1 (Dlg1 [ Parnas et al., 2001]).
(a) Images of synaptic bouton triple-labelled for Endo, Mnb and BRP at rest and after stimulation with high K+ for 30 s.
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