In order to improve the ecological operation effect and increase the success rate of ecological operation practice, a set of ecological operation target quantification methods based on the bio-hydrological response model for stimulating reproduction of four major Chinese carps is proposed. The method includes the definition of upwelling event, the selection of eco-hydrological indicators, the definition of natural fish reproduction event and effective reproduction response, the construction of bio-hydrological response model and multi-indicator discriminative preference. The actual observed data of 47 upwelling events and 24 effective reproduction responses of four fish in the Yichang River section for 6 years after the operation of the Three Gorges Reservoir from 2013 to 2019 (except 2016), were used to construct the model and quantify the ecological planning goals of the reservoir. The results showed that (1) the single-indicator bio-hydrological response model based on the four indicators of initial flow (Q₀), peak flood flow (Q_peak), total flow growth (Q_sum) and daily flow growth rate (dQ) had the ability to discriminate whether the natural reproduction of the four major carp was responsive to hydrological conditions; (2) the optimal thresholds for the above evaluation indicators were 14960 m³/s, 19610 m³/s, 4050 m³/s and 1405 m³/(s·d), respectively; the performance of the single indicator model was ranked as dQ>Q_peak>Q_sum>Q₀; (3) the comprehensive performance of the multi-indicator bio-hydrological response model based on the combination of dQ or Q_peak was the best. The model results are simple and predictable, and can be used to quantify the ecological operating targets, which can provide a reference for discriminating fish reproduction response and formulating reservoir ecological planning in different watersheds and reservoirs.