Hi,
Our lab did imaging of rab-3(ox699); shyEx246(Pglr-4>Flippase) in head of dauer worms. However, we could not identify what neuron was expressing the GFP. Can someone advice me on this? The imaging result can be seen here (sorry, i could not put the link so i add a space between https and drive as well as between google and com; https:// drive.google. com/drive/folders/1Jw5_43ujNfxOdu9u29t-b_MitxPejsu5?usp=sharing).
Thank you!
I tried to get to your image with the posted link (with the two spaces removed) and was unable to do so. Janet
Hi! I’m sorry, I forgot to change the settings. You should be able to access it now. Thank you!
I am no expert in identifying neurons, but it looks like you have 4 fold-symmetry in the head, which is consistent with CEP labeling. A first try for cell identity might be to do DiI labeling of your worms (probably before induction of dauer, since I believe that dauer formation blocks acess of the chemosensory neurons to DII). This is super easy - see WormBook for protocols. You would see processes in the head that would be distinct from the CEP’s (which do not label with DiI). If you see overlap of staining with the GFP, then you instead have some subset of the DiI/FITC labeling chemisensory cells labelled. Hope this is useful - perhaps someone who is more familiar with head neurons may have another suggestion. Good luck, Janet
I agree with Janet, it looks like CEP neurons, but there might even be some expression in ADE as well. You could order a strain from the CGC that carries otIs181 (dat-1p::mCherry expression) and cross it in together with your stuff. If you want further confirmation that it’s in the dopaminergic neurons, I actually have rab-3(ox699) with wyIs834 (integrated dat-1p::FLP) growing right now if you want some.