The dose-response effects of exercise in reduced gravity on musculoskeletal health have not been well documented. It is not known whether or not individualized exercise prescriptions can be effective in preventing the substantial loss in bone mineral density and muscle function that have been observed in space flight and in bed rest. In this study, typical daily loads to the lower extremities were quantified in free-living subjects who were then randomly assigned to control or exercise groups. Subjects were confined to 6-degree head-down bed rest for 84 days. The exercise group performed individually prescribed 1 g loaded locomotor exercise to replace their free-living daily load. Eleven subjects (5 exercise, 6 control) completed the protocol. Volumetric bone mineral density results from quantitative computed tomography demonstrated that control subjects lost significant amounts of bone in the intertrochanteric and total hip regions (p < 0.0125), whereas the exercise group showed no significant change from baseline in any region (p > 0.0125). Pre-and post-bed rest muscle volumes were calculated from analysis of magnetic resonance imaging data. The exercise group retained a larger percentage of their total quadriceps and gastrocnemius muscle volume (- 7.2% ± 5.9, - 13.8% ± 6.1, respectively) than their control counterparts (- 23.3% ± 5.9, - 33.0 ± 8.2, respectively; p < 0.01). Both groups significantly lost strength in several measured activities (p < 0.05). The declines in peak torque during repeated exertions of knee flexion and knee extension were significantly less in the exercise group than in the control group (p < 0.05) but work done was not significantly different between groups (p > 0.05). The decline in VO2max was 17% ± 18 in exercising subjects (p < 0.05) and 31% ± 13 in control subjects (p = 0.003; difference between groups was not significant p = 0.26). Changes in blood and urine measures showed trends but no significant differences between groups (p > 0.05). In summary, the decline in a number of important measures of musculoskeletal and cardiovascular health was attenuated but not eliminated by a subject-specific program of locomotor exercise designed to replace daily load accumulated during free living. We conclude that single daily bouts of exposure to locomotor exercise can play a role in a countermeasures program during bed rest, and perhaps space flight, but are not sufficient in their own right to ensure musculoskeletal or cardiovascular health.