Adeno-associated virus (AAV) vectors represent promising tools for central nervous system (CNS) gene therapy; however, achieving efficient widespread brain transduction remains challenging. This study compares the transduction of three AAV serotypes, AAV2, AAV8, and AAV-B10, following intracerebroventricular (ICV) administration in adult mice. Using advanced tissue clearing methods with high-resolution light-sheet fluorescence microscopy, we visualized whole-brain AAV distribution at single-cell resolution and employed semi-automated spatial quantification to analyze regional transduction. Each vector carried identical enhanced green fluorescent protein (eGFP) expression cassettes under the ubiquitous CAG promoter. AAV-B10 demonstrated superior parenchymal transduction across multiple brain regions, with significantly higher expression in the cortex, midbrain, striatum, and thalamus compared to other serotypes. In contrast, AAV2 exhibited distinctive tropism for ventricular structures, with robust transduction of ependymal cells lining the ventricular walls and choroid plexus. AAV8 showed intermediate distribution targeting the choroid plexus epithelium. Following ICV administration, peripheral leakage of AAV genomes was observed, with AAV8 showing strong liver enrichment of eGFP transgene and AAV-B10 displaying broader yet weaker distribution across peripheral tissues. These serotype-specific distribution profiles provide critical insights for optimizing vector selection based on therapeutic requirements and target regions, potentially enhancing the efficacy of AAV-based gene therapies for neurological disorders.
