Objective An orthogonal design was used to optimize the experimental parameters for theestablishment of a rat model of conditioned fear memory and to identify the optimal experimental conditions. Methods Themodel of conditioned fear memory in rats was established based on pavlovian conditioned fear theory. Three factors, namelysound intensity, cycle times and electric shock intensity, were adopted, and each factor was set at 3 levels. Orthogonal testwas carried out, and the freezing time ratio of rat was taken as the index to observe the changes in different parameters ofexpression of fear memory and determine the best experimental conditions. The conditioned fear memory model wasestablished again under the optimal conditions of the previous experiment, and the fear memory retention test was conducted24 h, 1 week, 2 weeks, 4 weeks and 8 weeks later. The rats without fear training were used as the control group. ResultsThe intuitive analysis results showed that the effects of various factors on the duration of fear memory of rats was soundintensity=cycle times > electric shock intensity in turn. The ANOVA test showed that the cycle times had a significantimpact on freezing time ratio ( P < 0. 05), but the sound intensity and electric shock intensity had no significant impact ( P > 0. 05). The optimal experimental conditions were as follows: 75 dB sound, 0. 8 mA electric shock, and 15 cycles. Fearmemory was maintained at 24 h and 1 week after the establishment of the model, which was significantly different from thatof the control group ( P < 0. 001 and P < 0. 05). Fear memory had faded 2 weeks later, at which point there was nosignificant difference in freezing time ratio between the model and control groups ( P > 0. 05). Conclusions Thisexperiment clarifies the primary and secondary factors that influence the establishment of a rat model of conditioned fearmemory The optimal experimental method for model establishment are identified. This study lays a foundation for the standardization and normalization of fear memory model.