A detailed understanding of the hip loading environment is needed to help prevent hip fractures, minimize hip pain, rehabilitate hip injuries, and design osteogenic exercises for the hip. The purpose of this study was to compare femoral neck stress during stair ascent and descent and to identify the contribution of muscles and reaction forces to the stress environment in mature adult subjects (n = 17; age: 50–65 y). Motion analysis and inverse dynamics were combined with musculoskeletal modeling and optimization, then used as input to an elliptical femoral neck cross-sectional model to estimate femoral neck stress. Peak stress values at the 2 peaks of the bimodal stress curves (stress vs time plot) were compared between stair ascent and descent. Stair ascent had greater compressive stress than descent during the first peak at the anterior (ascent: −18.0 [7.9] MPa, descent: −12.9 [5.4] MPa, P < .001) and posterior (ascent: −34.4 [10.9] MPa, descent: −27.8 [10.1] MPa, P < .001) aspects of the femoral neck cross section. Stair descent had greater tensile stress during both peaks at the superior aspect (ascent: 1.3 [7.0] MPa, descent: 24.8 [9.7] MPa, peak 1: P < .001; ascent: 15.7 [6.1] MPa, descent: 18.0 [8.4] MPa, peak 2: P = .03) and greater compressive stress during the second peak at the inferior aspect (ascent: −43.8 [9.7] MPa, descent: −51.1 [14.3] MPa, P = .004). Understanding this information can provide a more comprehensive view of bone loading at the femoral neck for older population.