Patterns of gene expression define the status of a cell in physiological as well as pathological conditions. The interplay of gene promoters and enhancers has been identified as a major mechanism to control specific expression patterns. Furthermore, combinations of enhancer elements with a preferable activity profile together with a small core promoter are starting to be used to drive transcription in gene delivery vectors.However, the identification of enhancers often relies on indirect measures, such as DNA conservation or epigenetic signatures. In addition, functional screens for enhancer activities are typically performed in immortalized cell lines in culture. This significantly hampers the identification of enhancers to be used in gene therapy vectors such as AAV, as the delivered DNA remains episomal and free from chromatin and the transcriptional profile of immortalized cell lines substantially differs from fully differentiated cells embedded in their natural tissue.Here, we transposed the Self-Transcribing Active Regulatory Region Sequencing (STARR-seq) technique to the mouse brain using adeno-associated-viruses (AAV) for the delivery of a highly complex screening library tiling entire genomic regions and revealing the enhancer activities in a total of 3Mb of the mouse genome.We identified 483 sequences with enhancer activity, including sequences that were not predicted by DNA accessibility or histone marks. Characterizing the expression patterns of fluorescent reporters controlled by nine candidate sequences, we observed differential expression patterns also in sparse cell types. Together, our study provides an entry point for the unbiased study of enhancer activities in organisms during health and disease.