Detecting axonal proteins or mRNA from sensory or motor neurons in target tissues (skin, viscera, muscle)


#1

I’m looking for papers that successfully detect the axonal proteins or mRNAs in lysates of innervated tissues. For example, do people detect channel proteins (Nav1.7, Nav1.8, TRPV1) or structural proteins (NF200) in skin lysates? We know those proteins are there. We can see them on IHC in the axons themselves. But these are probably very small quantities. Can you detect the proteins by Western or an ELISA or high-sensitivity proteomics? Similarly for muscle, where there is both motor axon and sensory axon innervation? For mRNAs, you’d only be able to find the axonally-transported ones. Can you do RNA-seq and detect these or is it too little? How about smFISH?

@CandlerPaige @liz @tberta @thicunha @sshiers @smithtp @vanja


#2

This paper looks promising although it looks like they pool whole nerves rather than looking at the target tissue.


#3

Lots of people have done this with cultures but not in vivo. We are trying it with trap. Its tough but we r getting there. For axonal rna of course.


#4

Hey @Theodore.price
Thanks for chiming in. I figured you’d know. I’ve seen those in vitro papers but nothing for in vivo or whole tissue. It must be possible. Just a matter of the sensitivity of our methods. You ever seen in situ hybridization of local mRNA sensory axons in skin? Good luck with the TRAP.


#5

@achamess. It is a interesting question. I never saw. Since the levels of these proteins must be very low at the peripheral sites to be detected by WB, maybe you can try immunoprecipitation before WB to concentrate the samples.


#6

I’m reminded of this paper https://www.ncbi.nlm.nih.gov/pubmed/24904046 which used TRAP to compare axonal vs somatic mRNA in Purkinje neurons.

I would imagine you might be able to see some axonally localized mRNAs in innervated tissue with in situ hybridization, but I haven’t ever tried it for any of our projects.