An enhancer-AAV toolbox to target and manipulate distinct interneuron subtypes

Establishing tools that can target and manipulate specific neuronal populations is critical to understanding neural circuits and developing targeted therapies. Recombinant adeno-associated virus (rAAV) is a popular tool for gene delivery and is widely used in basic research as well as gene therapy. Significant effort has been made to develop rAAV tools that can drive cell-type or tissue specific transgene expression. One of the strategies to develop such tools is to incorporate cell-type specific enhancer elements into the rAAV genome. In recent years, we and others have identified a number of enhancers that, when incorporated into rAAV vectors, can restrict the transgene expression to particular neuronal populations. Yet, viral tools to access and manipulate fine neuronal subtypes are still limited. Here, we performed systematic analysis of single cell ATAC-seq and RNA-seq data to identify enhancer candidates for each of the cortical interneuron subtypes.We established a set of enhancer-AAV tools that are highly specific for distinct cortical interneuron populations and striatal cholinergic neurons. These enhancers, when used in the context of different effectors, can target (fluorescent proteins), observe activity (gCaMP) and manipulate (opto- or chemo-genetics) specific neuronal subtypes. We also validated our enhancer-AAV tools across species. Thus, we provide the field with a powerful set of tools to study neural circuits and functions and to develop precise and targeted therapy. To identify potential cell type specific enhancers, we took advantage of the single-cell assay for transposase accessible chromatin using sequencing (scATAC-seq) data we have previously generated in the lab. To enrich for GABAergic cells, Dlx6a-Cre::INTACT mice were used. GFP+ nuclei from cortex (anterior lateral motor cortex, ALM) and striatum were isolated by FACS and scATAC-seq data were collected