The aim of this study is to investigate impact response and dislocation substructure of Ti-6Al-4V alloy using a compressive split-Hopkinson pressure bar and transmission electron microscopy technique. Cylindrical specimens are deformed at strain rates ranging from 8×10 2 s -1 to 4.0×10 3 s -1 and temperatures of 0°C and -200°C respectively. It is shown that the impact properties of Ti-6Al-4V alloy depend strongly on strain rate and temperature. For a constant temperature, the flow stress and strain rate sensitivity increase with increasing strain rate, while the activation volume decreases. Meanwhile, for a constant strain rate, the activation volume increases with increasing temperature, while the flow stress and strain rate sensitivity decrease. Transmission electron microscopy (TEM) observations show that the dislocation density increases with an increasing strain rate, but decreases with an increasing temperature. The mechanical properties of the impacted specimens are related to the microstructural evolution.