Efficient in utero gene transfer system to the developing mouse brain using electroporation: Visualization of neuronal migration in the developing cortex

We report a novel gene transfer system using electroporation. We used this technique to introduce a marker gene plasmid containing enhanced green fluo rescent protein into mouse brains at embryonic day 12-17 without removing t he embryos from the uterus. The embryos were allowed to continue to develop in utero, and more than 80% were born normally expressing the exogenous ge ne. Enhanced green fluorescent protein driven by the cytomegalovirus promot er was strongly expressed in the ventricular zone, radial fibers and migrat ing neuroblasts, but not in mature neurons, suggesting that the cytomegalov irus promoter is silenced after the cells differentiate into mature neurons . Since there is still no convenient way of visualizing the migrating neuro blasts, especially of distinguishing them from the surrounding mature neuro ns in the cortical plate, this system should provide a good tool for analys ing neuronal migration. In the postnatal lateral cortex, neuroblasts migrat ed almost "tangentially" along the obliquely running "radial" fibers beneat h the cortical plate, and after entering the cortical plate, turned towards the marginal zone and migrated radially. Neurons with primitive dendrites were observed only along the border between the marginal zone and the corti cal plate, and never at other sites, such as in the middle of the cortical plate. These results imply that the neuroblasts do terminate migration and start differentiation to mature neurons when they encounter the marginal zo ne, as has long been suggested. By contrast, when elongation factor 1 alpha promoter was used, prominent fluorescence allowed visualization of the ent ire mature neurons as well. The labeled neurons were observed to send axons to the contralateral cortex where they arborized extensively. Thus, this system is much easier and more efficient than virus-mediated gen e transfer, and is useful for gain-of-function analysis of neural cell fate determination, migration, positioning and axon path-finding in mouse embry os. (C) 2001 IBRO. Published by Elsevier Science Ltd. All rights reserved.