Tunnel excavation has significant disturbance on groundwater system and related geo-environment, especially in karst regions like southwestern China. The present research was conducted to quantitatively understand the negative impacts posed by tunnel construction on the karst groundwater system and reveal the behavior of karst groundwater system under the tunnel disturbance, with the aid of field survey, hydrogeological analysis, and numerical simulation. The results suggested that negative impacts such as loss of surface and underground water, ground collapse, and house deformation would be posed directly and indirectly to the karst groundwater system and its dependent geo-environment as the result of groundwater level drawdown by tunnel excavation. The degree and range of groundwater drainage impact were determined by the lithological and hydrogeological characteristics of strata. These negative impacts were dominantly distributed in the karst depressions valleys, and the direct ones occurred at first and followed by the indirect ones. Simulation results showed groundwater level drawdown would not occur synchronously in spatial, but always occurred around the tunnel axis at first and gradually expanded towards far away over time. The maximum disturbance on groundwater system can reach to approximately 25 m vertically and 3000 m horizontally for present modeling tunnel. With the aid of numeral simulation, three response stages were identified for the karst groundwater system behavior to the tunnel disturbance. The impacts of tunnel practice on groundwater and surface water bodies can be gradually eliminated since the second stage, but would continue if existing failure of tunnel waterproof until a new balance state achieved. The present research can improve the understanding of the impacts of tunnel excavation on karst groundwater system and dependent geo-environment, and provide reference to the protection of water resources and geo-environment in karst regions like Chongqing worldwide.