Neuronal conversion of pre-labeled astrocyte using adeno-asociated viruses in vivo

Neuronal reprogramming using adeno-associated viruses (AAVs) carrying a GFAP mini-promoter to drive neurogenic transcription factor expression in astrocytes in vivo were refuted due to off-target neuronal expression. Here, we show successful neuronal conversion of pre-labeled astrocytes using AAV-GFAP to express phospho-deficient Ascl1SA6, and demonstrate that miR124 target sites reduce off-target neuronal expression. Finally, single cell transcriptomic and pseudotime trajectory analyses confirmed Ascl1SA6-driven astrocyte-to-neuron conversion, and more limited oligodendrocyte conversion events Overall design: Following AAV injection into the motor cortex of C57/BL6 male mice, brains were harvested 7 and 50 days post injection. Cortex was microdissected under the microscope and single cell suspension was prepared for single cell analysis using scRNA-seq.