Analysis of Synaptic Connections at the Electron Microscopic Level Using Viral Vectors
Anterograde tracing using Sindbis virus vector resolves several problems of conventional tracers. Usage of the Sindbis virus as a neural tracer prevents the axons passing through the injection site from being labeled because the virus does not infect from axons. Combining the methodological merit of the virus tracer with electron microscopic technique, we analyze morphological characteristics of ultrastructure of axon terminals which are provided specifically from the region of interest, and thus we can systematically extract information of synaptic organization from complicated neural networks. Here, we introduce a method where Sindbis virus labeling is combined with an immunoelectron microscopic technique. Using this technique, we observe synaptic contacts and GABA immunoreactivity in ultrastructure of axon terminals which are revealed to be projected from the virus injection site.
KeywordsSindbis virus Anterograde tracing Electron microscopy GFP Post-embedding immunostaining
This work was supported by the Ministry of Education, Science, Sports, and Culture of Japan Grants (15H04266, 15H01430 and 15K14333). We thank Dr. H. Bokor for the helpful advice on post-embedding immunostaining methods.