The effect of directional grain structure and strain rate on the impact properties and dislocation substructure of 6061-T6 aluminum alloy is studied. Impact tests are performed at strain rates ranging from 1x103 to 5x103s-1 using a split Hopkinson pressure bar system. Cylindrical specimens are prepared from the rolled plates in longitudinal direction, transverse direction and through-thickness direction, respectively. The results show that the flow stress is strongly dependent on the strain rate and displays complex variations with grain structure direction. The flow stress increases with increasing strain rate. For all tested strain rates, the flow stress is the highest in the transverse specimen, followed by the through-thickness specimen and longitudinal specimen. However, at the strain rate of 5×102s-1, the flow stress in longitudinal specimen is higher than that in through-thickness specimen due to the change of dislocation multiplication rate. The plastic flow occurs within the deformation regions, and becomes more pronounced at high strain rates, especially for the longitudinal specimen. Dislocation density increases markedly with increasing strain rate. Strengthening effect is the highest in the transverse specimen, followed by the longitudinal specimen and through-thickness specimen.