The alpha-herpesviruses: molecular pathfinders in nervous system circuits

Abstract

Several neuroinvasive viruses can be used to study the mammalian nervous system. In particular, infection by pseudorabies virus (PRV), an alpha-herpesvirus with broad host range, reveals chains of functionally connected neurons in the nervous systems of a variety of mammals. The specificity of PRV trans-neuronal spread has been established in several systems. One attenuated strain, PRV-Bartha, causes a reduced inflammatory response and also spreads only from infected post- to pre-synaptic neurons. We review the basics of PRV tracing and then discuss new developments and novel approaches that have enabled a more detailed understanding of the architecture of the nervous system. As questions and techniques evolve in the field of neuroscience, advances in PRV tracing will certainly follow.

Figure 1

Trans-synaptic spread of PRV infection in a neural circuit. (a) After infecting primary neurons (shown in green), wild-type strains of PRV, such as PRV-Becker or PRV-Kaplan, promote infection that spreads from the cell body to axon terminals of pre-synaptic neurons (retrograde spread). Virions of wild-type strains might also be transported down the axon to infect the cell bodies of post-synaptic neurons (anterograde spread). (b) PRV-Bartha replicates well in neurons, but infected animals have reduced symptoms compared to wild-type virus infections. In addition, PRV-Bartha infection spreads only from post- to pre-synaptic neurons in a circuit. Infection by PRV-Bartha cannot spread from pre- to post-synaptic neurons because structural components of virions cannot be sorted into axons, a process that requires the actions of viral proteins gE, gI and US9. The genes encoding these proteins are deleted in the PRV-Bartha genome.

Figure 2

Retrograde tracing from molecularly defined neurons. (a) The PRV genome includes a unique long (UL) and unique short (US) segment and internal (IR) and terminal (TR) repeat regions. (b) Ba2001 has a deletion in the gene encoding the viral thymidine kinase (Δtk). Without TK, PRV cannot replicate in non-dividing cells, such as neurons. Glycoprotein G (gG) is a non-essential protein and the gG locus is a common location for the insertion of reporter genes. (c) The gG locus in Ba2001 has a lox–stop–lox cassette between the CMV immediate-early promoter (CMV) and the genes for the tau–EGFP marker and TK gene from HSV. Tau–EGFP and HSV–tk are separated by an internal ribosome entry signal (IRES). (d) Recombination between the lox sites occurs only in cells expressing Cre. This enables the CMV promoter to drive expression of the tau–EGFP and HSV–tk bicistronic message. The tau–EGFP acts as a fluorescent marker in infected cells. The HSV–tk enables Ba2001 to replicate in neurons. (e) In a transgenic mouse engineered to express Cre from a specific promoter, PRV-Bartha will infect all neurons within range of the injection site (pink circle). Ba2001 will initiate replication and trace retrograde circuitry only from neurons that express Cre-recombinase