A Modified Sindbis Vector for Prolonged Gene Expression in Neurons

Division of Neuroscience, Baylor College of Medicine, Houston, Texas 77030 Submitted 14 May 2003; accepted in final form 25 June 2003 Sindbis viruses have been widely used in neurobiology to express a variety of genes in cultured neurons, in cultured slices, and in vivo. They provide fast onset and high levels of expression of foreign genes, but the expression is limited to a short time window due to a shut-off of host protein synthesis. We have used a mutation in an essential gene (nsP2) of the life cycle of Sindbis, which allows the functional analysis of changes in protein expression for 6 days after infection. This Sindbis mutant (nsP2) was used to express enhanced green fluorescent protein (EGFP) in hippocampal neurons in culture and in vivo without any sign of toxicity, based on two-photon imaging and electrophysiology. In addition, the EGFP mutant virus can be injected in vivo to visualize spines and other details of neuronal structure. The Sindbis mutant described here provides an improved tool in neurobiology with reduced cytotoxicity and a prolonged time window of expression for novel applications in imaging and behavior. In addition, the use of this vector for the functional expression of mammalian voltage-gated ion channels in organotypic slices is demonstrated. Address for reprint requests and other correspondence: A. Jeromin, Div. of Neuroscience, Baylor College of Medicine, Houston, TX 77030 (E-mail: jeromin{at}ltp.neusc.bcm.tmc.edu ).