The relationship between microstructure and diffusion of oxygen in YBCO films are studied in this work. The dislocation structure in magnetron sputtered c-axis oriented YBCO films deposited on (100)SrTiO3 substrates consists of vertical (//c-axis) screw dislocation forests together with layered horizontal (⊥c-axis) edge dislocation net-works. It is found that in the oxygenation process of YBCO films, oxygen diffusion is greatly enhanced by short circuit paths which we suggest to be the horizontally (⊥c-axis) layered edge dislocation net-works. Base on the structure of c-axis oriented YBCO films, the oxygen diffusion process was examined by applying the cylinder diffusion model. The diffusion coefficient of oxygen and the corresponding activation energy were measured.