Efficient infection of organotypic hippocampal slice cultures with adenovirus carrying the transgene REST/NRSF

Organotypic hippocampal slice cultures provide a useful platform maintaining hippocampal structure and synaptic connections of the brain over weeks in culture with ease of in vitro manipulations. Gene transfer is a particularly desirable tool for using with them but current difficulties with transformation of transgenes into these cultures is a barrier to their use in research. Previous quantifications of viral infections have shown low transformation rates and have relied upon invasive microinjections. In this paper we present an efficient way of infecting organotypic cultures with adenovirus at the acute slice stage that does not require injection. We use the adenoviral delivery system to introduce the transcription factor REST and a GFP marker, providing around 41 % cellular infection spread throughout the entire slice culture and promoting transgene expression for weeks in vitro. GFP expression was observed most intensely in the slices when they were infected just a few hours after plating and was shown to infect neurons and microglia. We decided to use the transcription factor REST/NRSF as an example transgene which was delivered into cells via the adenoviral construct, conferring overexpression of REST in addition to the GFP marker. This outlines a technique whereby adenoviral infection of organotypic cultures can infect neurons with good efficiency and confer successful manipulation of genetic factors within the cell.

Automated summary

The study presents an efficient method to infect organotypic cultures with adenovirus at the acute slice stage without injection. By introducing the transcription factor REST and a GFP marker using the adenoviral delivery system, a high cellular infection spread of around 41% was achieved, promoting transgene expression for weeks in vitro. This technique allows for successful manipulation of genetic factors within neurons and microglia in organotypic cultures.