The issue of the direct action of glucocorticoids on bone resorption was addressed in a model system consisting of a pure (99%) population of putative osteoclast precursors, namely thioglycollate-elicited rat macrophages cocultured in vitro with devitalized rat bone particles which had been prelabeled in vivo with 45Ca. After 4 days in culture, these macrophages mobilized a net 20—30% 45Ca from bone. The addition of cortisol in physiological concentrations (10-8 M) elicited significant (P < 0.001) increases in isotope mobilization; optimal enhancement occurred at 10-8 M and approximated the circulating level achieved with high dose glucocorticoid therapy. Other steroids, specifically aldosterone, 17α- and β-estradiol, 11-deoxycortisol, and progesterone, did not enhance the resorptive process. The augmentation of bone 45Ca mobilization by cortisoltreated macrophages was not due to protracted cell viability in culture or the increased avidity with which treated cells bind to bone. On the other hand, cortisol treatment was associated with stimulated protein and RNA synthesis, as evidenced by enhancement (P < 0.001) of both [3H]leucine and [3H]uridine incorporation into treated cells. This association was also underscored by the observation that increased protein and RNA synthesis and enhanced mineral mobilization were inducible phenomena, i.e. phenomena that could be observed 48-72 h after cells were pretreated with cortisol. We conclude, therefore, that glucocorticoids can directly modify both the metabolism and the mineralmobilizing activity of bone-resorbing cells.