MEC Synapse Annotation Protocol v1.0

General task

Go through each synapse indicated by either a painted cleft or a pre-existing line annotation and check it for validity, then scan the volume for any missed synapses.

Step-by-step

1. The volume starts with an annotation layer marking the points where AI-generated synapses occur. Copy this layer (click-and-drag the layer, then shift+click) and rename it "to_check". Also make a second, empty annotation layer named "checked".
2. Scan through the volume and add annotation points anywhere you see existing tracer-made line annotations that don't coincide with an AI-made synapse.
3. Work through the "to_check" list, deleting each point as you check it. Each time you delete a point, add a corresponding point to the "checked" layer — even when you're erasing a synapse, so you know the area has been covered.
4. Edit the cleft painting to add or remove as needed:
   • Actual gaps between the pre- and post-partners' membranes usually aren't visible in mouse tissue, so "cleft" here means all the space from the inside of the presynaptic partner's membrane to the inside of the postsynaptic partner's membrane, plus any PSD you notice.
   • If the clefts of two separate synapses touch, paint one with a different segment by changing the "target value" number in the brush menu (this creates a new segment, same as in VAST).
5. Create a line annotation for each synapse in the "New Tracer Synapses" annotation layer that came with the volume (it has Nico's shader script preloaded). Place the first point inside the presynaptic partner and the second point inside the postsynaptic partner.
6. Any axo-axonic synapses go in their own annotation layer (requested by Nico for labeling purposes).
7. Once all existing synapse sites have been checked, scan the volume for any missed by both the previous tracers and the AI, and add them as well.

Axo-axonic synapse criteria

• Bouton–bouton synapses can only be marked if the boutons aren't synapsing with anything else.
• Bouton–shaft synapses must show clear PSD to qualify.
• Axo-somatic synapses likely won't have PSD but still need to be marked. (Criteria pending — awaiting the outcome of Ben and Zhihao's conversation.)

Examples