Tunneling Nanotube (TNT) , a dynamic cell-cell contact, is dependent on actin polimerization. TNTs are efficient in transporting ions, proteins and organelles intercellularly, which are important mechanisms in physiological and phathological precesses. Reported studies on the existence and function of TNTs among neural cells focus on cultured cell for the convenience in detecting TNTs’ ultrastructure. In this study, the adeno-associated virus (AAV-GFAP-EGFP-p2A-cre) was injected in the cerebral cortex of knock-in mice ROSA26 GNZ. GFAP promoter initiated the expression of Enhanced Green Fluorescent Protein (EGFP) in infected astrocytes. At 10 days post injection (10 DPI), we found that EGFP could transfer from astrocytes in layerⅠ-Ⅲ to neurons in layer Ⅴ. The dissemination of EGFP was not through endocytosis or exosome. And the intercellular transportation of EGFP was F-actin dependent. Therefore, we concluded EGFP transported from astrocytes to neurons in coetex via F-actin dependent TNTs. Although it is hardly to detect the ultrastrucute of TNTs in brain for its transiency and the noisy background, we established an animal model and indirect experimental methods to explore TNTs in vivo.