Fig. 2: Synapse formation in OPN by regenerating RGC axons 6 months after pre-OPN OTI.
A Schematic diagram showing the labeling of synapses formed by RGC axons (anterogradely traced with CTB, blue) with the presynaptic marker (Bassoon, green) and postsynaptic marker (Homer1, red). B Fluorescence images showing the colocalization of CTB (blue) and Bassoon (green), with Homer1 (red) being adjacent to Bassoon, indicating synapses formed by regenerated RGC axons (marked by white arrows) in the OPN of 6 m Regen. mice. Scale bar: 2 µm. C Quantification of the percentage of synapses in CTB-labeled boutons in the Sham and 6 m Regen. groups. Data presents mean ± SEM. n = 3 mice. Statistical significance: p = 0.6362 student’s t-test, two-sided. D Schematic illustration of identifying dAPEX2-tagged dark mitochondria from presynaptic RGC axons in OPN with TEM. The mitochondrial matrix of the RGC axon terminal was tagged with COX4-dAPEX2 via AAV-COX4-dAPEX2 intravitreal injection, where dAPEX2 labeled mitochondria show high contrast after peroxidase reaction and OsO4 treatment. E TEM images of OPN slices from different groups showing dAPEX2-labeled mitochondria in presynaptic RGC axons (indicated by red arrows), unlabeled mitochondria from other regions (indicated by green arrows), synapses (indicated by purple triangles), and the edge of PSD (indicated by black arrows). The pseudo-light pink color represents the presynaptic area, while the light blue color represents the postsynaptic area. Scale bar: 200 nm. F Quantification of the number of synaptic vehicles (SV) per bouton (left panel) and PSD lengths (right panel). Data presents mean ± SEM. n = 4 (Sham) and n = 3 (6 m Regen.). Statistical significance: p = 0.7936 (left panel), p = 0.4446 (right panel) Student’s t-test, two-sided. ns, not significant, p > 0.05 Source data are provided as a Source Data file.
