title: A bicistronic lentiviral vector-based method for differential transsynaptic tracing of neural circuits.
abstract: We developed a bicistronic HIV1-derived lentiviral vector system co-expressing green fluorescent protein (AcGFP1) and wheat germ agglutinin (WGA) mediated by picornaviral 2A peptide. This system was first applied to the analysis of the rat cerebellar efferent pathways. When the lentiviral vector was injected into a specific lobule, the local Purkinje cell population (first-order neurons) was efficiently infected and co-expressed both AcGFP1 and WGA protein. In the second-order neurons in the cerebellar and vestibular nuclei, WGA but not AcGFP1 protein was differentially detected, demonstrating that the presence of AcGFP1 protein enables discrimination of first-order neurons from second-order neurons. Furthermore, WGA protein was detected in the contralateral ventrolateral thalamic nucleus (third-order nucleus). This system also successfully labeled rat cortical pathways from the primary somatosensory cortex and monkey cerebellar efferent pathways. Thus, this bicistronic lentiviral vector system is a useful tool for differential transsynaptic tracing of neural pathways originating from local brain regions.
authors.affiliation: Department of Physiology, The University of Tokyo School of Medicine, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan.
authors.forename: Yohei
authors.initials: Y
authors.lastname: Ohashi
authors.forename: Tadashi
authors.initials: T
authors.lastname: Tsubota
authors.forename: Ayana
authors.initials: A
authors.lastname: Sato
authors.forename: Kenji W
authors.initials: KW
authors.lastname: Koyano
authors.forename: Keita
authors.initials: K
authors.lastname: Tamura
authors.forename: Yasushi
authors.initials: Y
authors.lastname: Miyashita
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D002531:Cerebellum / Q000033:anatomy & histology; D018014:Gene Transfer Techniques; D005822:Genetic Vectors / Q000235:genetics / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D016086:Lentivirus / Q000235:genetics / Q000378:metabolism; D008251:Macaca; D008297:Male; D009434:Neural Pathways / Q000033:anatomy & histology* / Q000378:metabolism; D009474:Neurons / Q000378:metabolism / Q000648:ultrastructure*; D051381:Rats; D017208:Rats, Wistar; D013194:Staining and Labeling / Q000379:methods*; D013569:Synapses / Q000502:physiology / Q000648:ultrastructure*; D014909:Wheat Germ Agglutinins / Q000235:genetics / Q000378:metabolism
chemical_list: D014909:Wheat Germ Agglutinins; D049452:Green Fluorescent Proteins