Pseudorabies virus expressing enhanced green fluorescent protein

A tool for in vitro electrophysiological analysis of transsynaptically labeled neurons in identified central nervous system circuits

Bret N. Smith, Bruce W. Banfield, Cynthia A. Smeraski, Christine L. Wilcox, F. Edward Dudek, Lynn W. Enquist, Gary E Pickard

Research output: Contribution to journalArticle

157 Citations (Scopus)

Abstract

Physiological properties of central nervous system neurons infected with a pseudorabies virus were examined in vitro by using whole-cell patch-clamp techniques. A strain of pseudorabies virus (PRV 152) isogenic with the Bartha strain of PRV was constructed to express an enhanced green fluorescent protein (EGFP) from the human cytomegalovirus immediate early promoter. Unilateral PRV 152 injections into the vitreous body of the hamster eye transsynaptically infected a restricted set of retinorecipient neurons including neurons in the hypothalamic suprachiasmatic nucleus (SCN) and the intergeniculate leaflet (IGL) of the thalamus. Retinorecipient SCN neurons were identified in tissue slices prepared for in vitro electrophysiological analysis by their expression of EGFP. At longer postinjection times, retinal ganglion cells in the contralateral eye also expressed EGFP, becoming infected after transsynaptic uptake and retrograde transport from infected retinorecipient neurons. Retinal ganglion cells that expressed EGFP were easily identified in retinal whole mounts viewed under epifluorescence. Whole-cell patch-clamp recordings revealed that the physiological properties of PRV 152-infected SCN neurons were within the range of properties observed in noninfected SCN neurons. Physiological properties of retinal ganglion cells also appeared normal. The results suggest that PRV 152 is a powerful tool for the transsynaptic labeling of neurons in defined central nervous system circuits that allows neurons to be identified in vitro by their expression of EGFP, analyzed electrophysiologically, and described in morphological detail.

Original languageEnglish (US)
Pages (from-to)9264-9269
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Volume97
Issue number16
DOIs
StatePublished - Aug 1 2000

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Suid Herpesvirus 1
Staphylococcal Protein A
Central Nervous System
Neurons
Suprachiasmatic Nucleus
Retinal Ganglion Cells
enhanced green fluorescent protein
In Vitro Techniques
Vitreous Body
Patch-Clamp Techniques
Cytomegalovirus
Thalamus
Cricetinae
Injections

Keywords

  • PRV 152
  • PRV-Bartha
  • Retinal ganglion cell
  • Suprachiasmatic nucleus

ASJC Scopus subject areas

  • General

Cite this

Pseudorabies virus expressing enhanced green fluorescent protein : A tool for in vitro electrophysiological analysis of transsynaptically labeled neurons in identified central nervous system circuits. / Smith, Bret N.; Banfield, Bruce W.; Smeraski, Cynthia A.; Wilcox, Christine L.; Dudek, F. Edward; Enquist, Lynn W.; Pickard, Gary E.

In: Proceedings of the National Academy of Sciences of the United States of America, Vol. 97, No. 16, 01.08.2000, p. 9264-9269.

Research output: Contribution to journalArticle

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