Karst groundwater of Jinci spring is one of the significant water supply sources for Taiyuan, Shanxi, China. However, groundwater overexploitation had affected karst groundwater balance over the years. Based on the available data from 1961 to 2008, and using a combination of the traditional water balance method, geographic information system (GIS) method and other hydrogeological researches, this study revealed the relationship between the overexploitation of karst groundwater and the cessation of Jinci spring, analyzed the variations of karst groundwater level after the depletion of the spring, identified the interaction between the karst aquifer in the mountainous area and the alluvial aquifer in the plain area, and delineated the critical zones of the impact of human activity on karst groundwater. The main findings include: (1) due to the consumption of 29 Mm³/a groundwater from the karst aquifer in 1961–1994, the reduction in storage led to the cessation of Jinci spring; (2) the decline rates of karst groundwater level varied from 1.27 to 3.99 m/a in 1994–2008; (3) groundwater overexploitation in the alluvial aquifer can negatively affect the karst aquifer; and (4) delineation of the critical zones are as follows: the coalmine area of Xishan Mountain, the peripheral fault zones, the plain area and the leakage section of the Fenhe River. The results are beneficial for alleviating the pressure of karst aquifer carrying water supply, landscape and aquatic ecosystem in the downstream of the spring, and provide scientific basis for the ecological protection of the spring.