Transneuronal circuit tracing with neurotropic viruses

Abstract

Because neurotropic viruses naturally traverse neural pathways, they are extremely valuable for elucidating neural circuits. Naturally occurring herpes and rabies viruses have been used for transneuronal circuit tracing for decades. Depending on the type of virus and strain, virus can travel preferentially in the anterograde or the retrograde direction. More recently, genetic modifications have allowed for many improvements. These include: reduced pathogenicity; addition of marker genes; control of synaptic spread; pseudotyping for infection of selected cells; addition of ancillary genetic elements for combining circuit tracing with manipulation of activity or functional assays. These modifications, along with the likelihood of future developments, suggest that neurotropic viruses will be increasingly important and effective tools for future studies of neural circuits.

Pseudoptyped Rabies Virus for Tracing Connections of Targeted Neurons

A. Normal and modified rabies virions. The normal rabies virion (left) includes an RNA core within the viral envelope. The envelope is coated with the rabies glycoprotein (RG). It is possible to produce modified rabies virions in which the RG gene has been deleted from the genome and replaced with coding sequence for EGFP (middle). When these modified virions are grown in culture they can be coated with RG, conferring the normal infectious properties of rabies virus (right, top), or pseudotyped with an envelope protein from another virus (e.g. EnvA), conferring the infectious properties of that virus (right, bottom). Although these modified virions are capable of infecting cells and replicating to produce large quantities of EGFP, they are not able to spread out of those cells without the help of another DNA expression vector that provides RG. This is because RG is absolutely essential for viral spread. B. Selective Infection. EnvA Pseudotyped rabies virus can be used to selectively infect neurons which have been targeted for expression of the EnvA receptor, TVA. Because there are not endogenous receptors for EnvA in the mammalian brain, other cells are not infected. Because, the RG gene has been deleted from the rabies genome, complementation is required to allow spread of the virus from infected cells. This can be accomplished by targeted expression of RG in the same cells which express TVA. C. Monosynaptically-restricted spread. Following infection and RG complementation in the initially infected neurons, the rabies virus is able to spread retrogradely to directly presynaptic neurons. However, because these presynaptic neurons lack RG expression, the virus cannot spread beyond these cells. After Wickersham et al. [35].