Abstract: Li₄Ti₅O₁₂ spinel has been regarded as one of the ideal anode materials for Li ion batteries and is a good candidate for developing high performance electric automobiles and smart grids for its characteristics of the long electrochemical cycle-life, high-structural stability and power density. Atomic-scale visualization of Li₄Ti₅O₁₂ spinel upon lithiation is realized using spherical aberration corrected transmission electron microscope (STEM), electron energy loss spectroscopy (EELS) and first-principles calculations. Upon lithiation, the Li₄Ti₅O₁₂ spinel transforms into the rock-salt Li₇Ti₅O₁₂ phase by developing an almost ideal hetero-interface and the Ti ions demonstrate different chemical states on the different sides of the interface. Further observation indicates that the elongation and shrinkage of Ti—O bonds in TiO₆ octahedra (breathing model) results in the fundamental changes of electronic conductivity in lithiated Li₄Ti₅O₁₂, where the insulating Li₄Ti₅O₁₂ is transformed into a quasi-conducting Li₇Ti₅O₁₂ phase. But these changes show limited influence on ionic conductivity. Using atomic-scale EELS analysis, a spontaneous oxidization process of Ti³⁺ to Ti⁴⁺ is revealed on the surface of Li₇Ti₅O₁₂ phase. This spontaneously surficial charge transfer reaction is revealed to be strongly related with the package swelling issue of Li₄Ti₅O₁₂ battery during cycling. Moreover, a three-phase separation, including the Li₄Ti₅O₁₂, Li₇Ti₅O₁₂ and Na₆LiTi₅O₁₂ phases, is found during Na insertion when Li₄Ti₅O₁₂ is employed as anode for sodium ion battery. All these important findings provide a rewarding avenue for the structural design and optimization of battery materials and lay a solid foundation for industrial application of Li₄Ti₅O₁₂ anode.