Objective: Previous studies have indicated that the type and intensity of exercise can modulate the trabecular structure in cancellous bone; however, the effect of exercise loading direction on the femoral head structure in rats remains unclear. Therefore, this study aimed to investigate the impact of different exercise directions on the bone microstructure of the femoral head in rats using an inclined treadmill protocol.Methods:Twenty-one Sprague-Dawley (SD) rats were randomly divided into three groups (0°、10°、-10°). They underwent intermittent treadmill training at a speed of 16 m/min, five times per week, with a cumulative exercise duration of 30 minutes per session, for a total of 12 weeks. Body weight was measured every two weeks before, during, and after the intervention. Micro-CT was used to collect structural data of the right femoral head, and Hematoxylin-Eosin (HE) staining as well as Tartrate-Resistant Acid Phosphatase (TRAP) staining were performed for analysis.Results:① Body weight increased over time in the horizontal, uphill, and downhill exercise groups. At the end of the intervention, the weights were (538.30±61.53) g, (502.17±31.85) g, and (532.77±72.91) g, respectively.②At week 4, the BV/TV values of the three groups were (0.20±0.11)%, (0.14±0.12)%, and (0.32±0.19)%, respectively, showing significant differences. At week 12, Tb.BMD also differed significantly among the three groups, with values of (0.02±0.04) g/cm3, (0.05±0.04) g/cm3, and (0.07±0.03) g/cm3, respectively. At week 8, Tb.N values were (0.58±0.86) mm?1, (0.79±0.86) mm?1, and (1.59±0.45) mm?1, respectively, indicating significant differences. At week 12, Tb.Th showed significant differences among the groups, with values of (0.01±0.02) mm, (0.04±0.03) mm, and (0.07±0.03) mm, respectively.③HE staining results revealed that, at the end of the intervention, the average number of osteoblasts in the uphill and downhill exercise groups was slightly higher than that in the horizontal group, but the difference was not statistically significant.TRAP staining results indicated that the number of osteoclasts in the horizontal group was significantly higher than that in the downhill group, but not significantly different from the uphill group.④The biomechanical results of rat femurs showed no significant differences in elastic modulus among the various exercise groups.Conclusion:Compared to horizontal and uphill exercise, downhill exercise more effectively suppresses bone resorption in the femoral head of rats and significantly enhances bone mineral density, trabecular number, and thickness. This study provides experimental evidence for understanding the mechanisms by which exercise regulates femoral head growth and for developing exercise-based therapeutic and rehabilitation strategies for related conditions.