Recombinant Semliki Forest Virus and Sindbis Virus Efficiently Infect Neurons in Hippocampal Slice Cultures

• Published in: Proceedings of the National Academy of Sciences - PNAS 1999-06, Vol.96 (12), p.7041-7046
• Main Authors: Ehrengruber, Markus U., Lundstrom, Kenneth, Schweitzer, Christophe, Heuss, Christian, Schlesinger, Sondra, Gähwiler, Beat H.
• Format: Article
• Language: English
• Subjects: Alphavirus Infections - virology; Animals; beta-Galactosidase - genetics; Biological Sciences; Culture Techniques; Cultured cells; Dendrites; DNA, Recombinant; Fluorescence; Gene Transfer Techniques; Genes; Genes, Reporter; Genetic Vectors; Green Fluorescent Proteins; Hippocampus - pathology; Hippocampus - virology; Infections; Luminescent Proteins - genetics; Nervous system; Neuroglia; Neurons; Neurons - virology; Pyramidal cells; Rats; RNA; Rodents; Semliki Forest virus; Semliki forest virus - genetics; Sindbis virus; Sindbis Virus - genetics; Viruses
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.96.12.7041

Gene transfer into nervous tissue is a powerful tool for the analysis of gene function. By using a rat hippocampal slice culture preparation, we show here that Semliki Forest virus (SFV) and Sindbis virus (SIN) vectors are useful for the effective infection of neurons. The stratum pyramidale and/or the granular cell layer were injected with recombinant virus encoding β -galactosidase (LacZ) or green fluorescent protein (GFP). By using low concentrations of injected SFV-LacZ or SIN-LacZ, we detected LacZ staining of pyramidal cells, interneurons, and granule cells. About 60% of the infected cells showed clear neuronal morphology; thus, relatively few glial cells expressed the transgene. Expression of GFP from SFV and SIN vectors gave similar results, with an even higher percentage (>90%) of the GFP-positive cells identified as neurons. Infected pyramidal cells were readily recognized in living slices, displaying GFP fluorescence in dendrites of up to fourth order and in dendritic spines. They appeared morphologically normal and viable at 1-5 days postinfection. We conclude that both SFV and SIN vectors efficiently transfer genes into neurons in hippocampal slice cultures. In combination with the GFP reporter, SFV and SIN vectors will allow the physiological examination of identified neurons that have been modified by overexpression or suppression of a specific gene product.