Journal: The Journal of Neuroscience
Article Title: Identification of Two Classes of Somatosensory Neurons That Display Resistance to Retrograde Infection by Rabies Virus
Figure Lengend Snippet: Monosynaptic tracing initiated from spinal Grp::Cre neurons. A , A' , Grp::Cre neurons are located in lamina II of the spinal cord. The neuropil and cell bodies of GRP::Cre neurons overlap with central terminals of IB4+ and TH+ primary afferent fibers. B , High-resolution imaging of a tdTomato+ Grp neuron (red) indicates the presence of Homer1+ (green) postsynapses on Grp neurons, which are near IB4+ (blue) terminals. C , High-resolution imaging of tdTomato+ Grp neuropil (red) indicates the presence of Homer1+ (blue) postsynapses on Grp neurons near vGlut3+ (green) primary afferent terminals. D , Rabies-mediated monosynaptic retrograde tracing reveals many primary (mCherry+ and eGFP+) and secondary (only eGFP+) infected spinal neurons. E , Secondary infected DRG neurons are mostly NF200+ (arrows). F , Quantification of eGFP-labeled DRG neuron subtypes after retrograde transduction mediated by either SAD-G or oG rabies glycoprotein. SAD.RabiesΔG-eGFP (Enva) was always injected at 3.5 × 10 8 focus forming units per milliliter. AAV1.EF1a.flex.mCherry-2A-SADG and pAAV1.Ef1a.DIO.oG.WPRE.hGH were injected at 9.5 × 10 12 GC/ml. Scale bars: A , D , E , 100 μm; B , C , 5 μm. Error bars represent the SD.
Article Snippet: AAV.flex.mCherry-2A-RabG vector was cloned by excising the ChR2-mCherry fusion protein from pAAV-Ef1a-DIO-hChR2(H134R)-mCherryWPRE-pA (kindly provided by Dr. Karl Deisseroth, Stanford University) with AscI and NheI and replacing it with PCR amplified mCherry-2A-RabG cDNA. pAAV-Ef1a-DIO-oG-WPRE-hGH was obtained from Addgene (74290; ) and packaged by the Viral Vector Facility (Zurich).
Techniques: Imaging, Retrograde Tracing, Infection, Labeling, Transduction, Injection