This study is aimed at providing an effective method for determining strain-load relationship and at quantifying the strain distribution within the whole tibia under axial compressive load on rats. Rat tibial models with axial compressive load were designed. Strains in three directions (0°, 45°, and 90°) at the proximal shaft of the tibia were measured by using a strain gauge rosette, which was used to calculate the maximum and minimum principal strains. Moreover, the strain at the midshaft of the tibia was measured by a single-element strain gauge. The slopes of the strain-load curves with different peak loads were calculated to assess the stability of the strain gauge measurement. Mechanical environment in the whole tibia by the axial compressive load was quantified using finite element analysis (FEA) based on microcomputed tomography images. The von Mises elastic strain distributions of the whole tibiae were evaluated. Slopes of the strain-load curves showed no significant differences among different peak loads (ANOVA; P > 0.05), indicating that the strain-load relationship obtained from the strain gauge measurement was reasonable and stable. The FEA results corresponded to the experimental results with an error smaller than 15% (paired Student's t-test, P > 0.05), signifying that the FEA can simulate the experiment reasonably. FEA results showed that the von Mises elastic strain was the lowest in the middle and gradually increased to both sides along the lateral direction, with the maximal von Mises elastic strain being observed on the posterior side under the distal tibiofibular synostosis. The method of strain gauge measurements and FEA used in this study can provide a feasible way to obtain the mechanical environment of the tibiae under axial compressive load on the rats and serve as a reference for further exploring the mechanical response of the bone by axial compressive load.