Many retrograde dyes work well for retrogradely labeling the somata of sensory neurons (DRG), such as Fast Blue, DiI, Fluoro-Gold, etc.
Does anything also fill the axons and show the afferent terminations in the spinal cord?
Does Cholera Toxin B (CTB) do this?
I don’t think CTB would work well for this. We haven’t used it in the way that you propose, but when I inject into the brain for the purpose of labeling spinal projection neurons, only the cell bodies and proximal dendrites are filled with the CTB.
You might be better off with a viral approach, as some vectors will go retrograde and then generate bright reporter expression in the transduced cell. The reporter will then fill all parts of the cell, depending on how much is produced and how the reporter is trafficked. If you have a nice strong promoter for your reporter then you should get some decent axonal labeling.
Thanks @Liz. Good to know.
Yes, virus works very nicely. But thus far, I haven’t found a virus that with high efficiency and indiscriminately (not preferring certain neurons) retrogradely infects DRG neurons.
The best solution I’ve found so far is using AAV1-Cre from the paw into a LSL reporter mouse. This seems to work fairly well.
I was thinking about this more, and you may consider a CAV vector. I have not tried CAV in anything other than my PB --> spinal projection neuron circuit so I have no idea how it might compare to AAV in your model. However, within my circuit, I would rank CAV as the second best virus I’ve tried in terms of retrograde labeling (first place is still AAV1 from UPenn, 3rd place is AAVretro from Addgene. I really ought to write all this down somewhere.) If you are having tropism issues with AAV, it may be better or different in CAV.
Lastly on a selfish note, did you inject into the skin of the paw or into the underlying muscle? What volume of virus and age of animal did you use to obtain this picture? A long time ago we were contemplating comparison of different afferent populations using viral tracing but we did not have the transgenic mouse tools at the time so it dropped off our radar. Maybe it’s time to revisit…
Hi @liz
Thanks again for your insights. I have still never seen a paper that uses CAV from the paw in the DRG. I thought about it, but given how CAV is not super accessible, I didn’t pursue more. Maybe it will work. Anyone know? @esypek?
Retrograde labeling with virus definitely depends on the cell types and regions. I think a lot of people are satisfied with retro-AAV in many parts of the CNS, but not all. For the DRG, there still isn’t a good solution. I’ve tried HSV from MIT and I wasn’t super impressed.
I think the Janelia folks may do a second version of retro-AAV, and they will consider peripheral neurons. AAV is definitely the friendliest and most widely used viral vector now, and having a good one for the periphery would be huge.
Lastly on a selfish note, did you inject into the skin of the paw or into the underlying muscle? What volume of virus and age of animal did you use to obtain this picture? A long time ago we were contemplating comparison of different afferent populations using viral tracing but we did not have the transgenic mouse tools at the time so it dropped off our radar. Maybe it’s time to revisit…
I injected into the skin. I’ve thought about labeling different afferents based on location, which is why I’ve been messing around with different viruses. I suspect muscle afferents would be labeled too. You would also get motor neurons though, but that likely won’t matter if you’re just trying to label.