Image the visual cortex with V4

[JIAXIN FU]

Hi all, 

We're trying to visualise individual neurons with the UCLA miniscope V4 in the visual cortices (the postrhinal cortex and V1) under a 3mm cranial window. 

After a few trials, we could only see general fluorescence in the FOV (please see: https://drive.google.com/file/d/1WB_TVA84JZrkxN5NVZTXakdF8RWiJne1/view?usp=drive_link). Could anyone share some experience for your successful attempts? I'm wondering the combination of parameters you use and how you troubleshoot to arrive at it.

We've tried with: 

1. pGP-AAV9-hsyn-jfCaMP8m; titre in the range of 10^12; inject in at least 3 sites (non-overlapping) of 30nl/site; injection speed of 1nl/s; with/without dura removal

2. AAV1-HSyn1-GCAMP6f-P2A-nls-dTomato with the same parameters as above

Apart from the difficulty of visualising individual neurons, we saw frequent bone regrowth and tissue damage even with dura removal. 

On a different note, if anyone has tried retrograde tracing from another brain region, specifically the ACC/amygdala, to the visual cortices, could you share which virus you use for that? 

Thank you very much for your suggestions in advance!!