[Objective] Chongqing, as the economic center of the upper reaches of the Yangtze River in China and a barrier to ensure ecological security in the middle and lower reaches of the Yangtze River, is a typical ecological functional and fragile area in China. Studying the spatiotemporal evolution characteristics and ecological sensitivity of its land use is of great significance for the reconstruction of land use and ecological protection in the research area. [Methods] A comprehensive assessment of the ecological sensitivity of Chongqing City was conducted by using remote sensing data from the years 2000, 2010 and 2020. This evaluation integrated various analytical approaches with software tools such as ArcGIS and Fragstat. Specifically, the Coupled Spatial Distance Index Model, Spatial Autocorrelation Analysis, Geodetector Model, and Grid Coding techniques were applied to evaluate the ecological sensitivity comprehensively. [Results] (1) In the study area, the landscape matrix was predominantly composed of farmland, accounting for more than 45% of the total area. There had been a significant reduction in both cropland and grassland, while the areas of forest land and urban-rural construction land had increased markedly from 2000 to 2020. (2) Over the course of 20 years, the average value of the comprehensive ecological sensitivity index rose from 1.037 to 1.045, indicating an overall improvement in the ecological environment of the study area. Spatial autocorrelation analysis revealed that the Global Moran’s I index for the comprehensive ecological sensitivity in Chongqing consistently exceeded 0, indicating a clear spatial heterogeneity in the sensitivity. Specifically, sensitivity decreased in a stepwise manner from the inner to the outer regions. Areas of high and very high sensitivity were predominantly found in the western part of Chongqing, the eastern new town, and the northeastern Three Gorges Reservoir area. Regions with significant changes in sensitivity levels were located in the central urban area and the southeastern Wuling Mountainous area of Chongqing. (3) The temporal and spatial evolution of comprehensive ecological sensitivity in the study area was the result of the joint action of natural environmental factors and social economic factors. Single-factor detection results indicate that the comprehensive index of average temperature, soil type, and land use degree were the main driving factors. The results of interactive exploration showed both linear and nonlinear enhancements, and the impact of the interaction between average temperature, annual rainfall, first industry added value, and per capita GDP on comprehensive ecological sensitivity showed a significant synergistic enhancement effect. [Conclusion] Based on the characteristics of ecological sensitivity spatial and temporal evolution in the study area, Chongqing City is divided into five zones, i.e. the comprehensive development zone, ecological improvement zone, potential governance zone, ecological regression zone, and strict protection zone, and corresponding measures are proposed for each zone. The research findings can provide theoretical support and decision-making basis for the reconstruction of ecological security pattern and sustainable development of Chongqing.