With climate warming and frequent extreme weather events, the structure and function of grassland ecosystems have been significantly altered. The Qilian Mountains are a wet island, serving as an important ecological barrier in China and central Asia. It is highly sensitive and prone to climate change, yet the vulnerability of grasslands to climate variability in this region remains unclear. In this study, based a NDVI dataset and meteorological data, we evaluated the relative vulnerability of grassland ecosystems to short-term climate variability by combining three indices: exposure, sensitivity, and resilience, and explored its relationship with water use efficiency (WUE). The results show that the spatial distribution patterns of grassland vulnerability in the Qilian Mountains from 2000 to 2018 were largely determined by exposure, and sensitivity is positively correlated with exposure and negatively correlated with resilience. The vulnerability of alpine meadow and alpine steppe is higher than that of temperate typical steppe and temperate desert steppe. Climate and topography gradients together affect the degree of grassland vulnerability. Grasslands with higher vulnerability are mainly found at medium elevation (3200–4000 m) area, where the topography is flat. The northern grasslands are primarily influenced by temperature, while the western and southern parts are mainly affected by precipitation. Alpine meadow and alpine steppe are more vulnerable to temperature and temperate desert steppe is more vulnerable to changes in precipitation. The higher the vulnerability, the lower the WUE of the grasslands. This study provides a reference for assessing the impact of climate change on grassland ecosystems and developing grassland vegetation management strategies.