The rapid development of urbanization brings challenges to the improvement of ecological quality. Solving the conflict between urbanization development and ecological quality improvement is the key to realize regional sustainable development. By constructing the evaluation index system of urbanization and ecological quality, this paper uses the coupling coordination degree model (CCDM) and geographically and temporally weighted regression model (GTWR) to reveal the degree of conflict and measure the spatio-temporal heterogeneity of the interaction between urbanization and ecological quality in Changsha-Zhuzhou-Xiangtan urban agglomeration from 2000 to 2020. The results showed as follows:(1) During the study period, the urbanization level of Changsha-Zhuzhou-Xiangtan urban agglomeration presented an upward trend, showing the characteristics that the central region was higher than the surrounding areas. The ecological quality indicated a trend of decreasing first and then increasing, and showed the characteristics of high in the surrounding area and low in the center. (2) The conflict degree between urbanization level and ecological quality of Changsha-Zhuzhou-Xiangtan urban agglomeration was gradually reduced, which had experienced the transformation from urbanization lag to ecological quality lag, and showed the characteristics of transition from high in the center and low in the surrounding area to low in the north and high in the south. (3) The interaction between urbanization level and ecological quality showed obvious stage characteristics. The negative effect of urbanization level and ecological quality was mainly distributed in the central developed areas, and the positive effect was mainly distributed in the surrounding areas of underdeveloped economy. Among them, the urbanization of population and space aggravated the deterioration of the natural ecosystem, while the reduction of ecological function and the fragmentation of structure also restricted the development of urbanization.