Link to the University of Pittsburgh Homepage
Link to the University Library System Homepage Link to the Contact Us Form

A dual infection pseudorabies virus conditional reporter approach to identify projections to collateralized neurons in complex neural circuits

Card, JP and Kobiler, O and Ludmir, EB and Desai, V and Sved, AF and Enquist, LW (2011) A dual infection pseudorabies virus conditional reporter approach to identify projections to collateralized neurons in complex neural circuits. PLoS ONE, 6 (6).

Published Version
Available under License : See the attached license file.

Download (2MB) | Preview
[img] Plain Text (licence)
Available under License : See the attached license file.

Download (1kB)


Replication and transneuronal transport of pseudorabies virus (PRV) are widely used to define the organization of neural circuits in rodent brain. Here we report a dual infection approach that highlights connections to neurons that collateralize within complex networks. The method combines Cre recombinase (Cre) expression from a PRV recombinant (PRV-267) and Cre-dependent reporter gene expression from a second infecting strain of PRV (PRV-263). PRV-267 expresses both Cre and a monomeric red fluorescent protein (mRFP) fused to viral capsid protein VP26 (VP26-mRFP) that accumulates in infected cell nuclei. PRV-263 carries a Brainbow cassette and expresses a red (dTomato) reporter that fills the cytoplasm. However, in the presence of Cre, the dTomato gene is recombined from the cassette, eliminating expression of the red reporter and liberating expression of either yellow (EYFP) or cyan (mCerulean) cytoplasmic reporters. We conducted proof-of-principle experiments using a well-characterized model in which separate injection of recombinant viruses into the left and right kidneys produces infection of neurons in the renal preautonomic network. Neurons dedicated to one kidney expressed the unique reporters characteristic of PRV-263 (cytoplasmic dTomato) or PRV-267 (nuclear VP26-mRFP). Dual infected neurons expressed VP26-mRFP and the cyan or yellow cytoplasmic reporters activated by Cre-mediated recombination of the Brainbow cassette. Differential expression of cyan or yellow reporters in neurons lacking VP26-mRFP provided a unique marker of neurons synaptically connected to dual infected neurons, a synaptic relationship that cannot be distinguished using other dual infection tracing approaches. These data demonstrate Cre-enabled conditional reporter expression in polysynaptic circuits that permits the identification of collateralized neurons and their presynaptic partners. © 2011 Card et al.


Social Networking:
Share |


Item Type: Article
Status: Published
CreatorsEmailPitt UsernameORCID
Card, JPcard@pitt.eduCARD
Kobiler, O
Ludmir, EB
Desai, V
Sved, AFsved@pitt.eduSVED0000-0002-4631-7842
Enquist, LW
ContributionContributors NameEmailPitt UsernameORCID
Date: 22 June 2011
Date Type: Publication
Journal or Publication Title: PLoS ONE
Volume: 6
Number: 6
DOI or Unique Handle: 10.1371/journal.pone.0021141
Schools and Programs: Dietrich School of Arts and Sciences > Neuroscience
Refereed: Yes
MeSH Headings: Animals; Base Sequence; Genes, Reporter; Genome, Viral; Herpesvirus 1, Suid--genetics; Herpesvirus 1, Suid--physiology; Male; Microscopy, Fluorescence; Neurons--cytology; Neurons--virology; Open Reading Frames; Pseudorabies--physiopathology; Rats; Rats, Sprague-Dawley
Other ID: NLM PMC3116869
PubMed Central ID: PMC3116869
PubMed ID: 21698154
Date Deposited: 05 Sep 2012 17:25
Last Modified: 02 Feb 2019 15:56


Monthly Views for the past 3 years

Plum Analytics

Actions (login required)

View Item View Item