Parathyroid hormone (PTH)-stimulated cyclic adenosine monophosphate (cAMP) in rat osteoblastlike (OB) cells has been shown to be modulated by steroid hormones; glucocorticoids are known to increase the level, while the effects of 1,25(OH)2D3 are inhibitory. In the present study, we found that the PTH-stimulated cAMP responses are similar in neonatal mouse and fetal rat OB cells. Dexamethasone (0.13-13 nM) augmented PTH-stimulated cAMP in both species. Mouse cells showed a higher maximal response to dexamethasone (100% increment) than rat cells (60-70% increment) with similar sensitivity to dexamethasone (ED50 approximately 1.0 nm). On the other hand, 1,25(OH)2D3 decreased PTH-stimulated cAMP, but the effect required pharmacological levels of hormone; mouse cells responded at a lower dose (1.3 nM) and were more sensitive than rat cells (responded at 13 nM) to 1,25(OH)2D3 treatment. Introduction of physiological concentrations of 1,25(OH)2D3 (0.013-1.3 nm) in addition to dexamethasone (13 nM) resulted in a synergistic enhancement of PTH-stimulated cAMP in rat cells. In contrast, a dose-dependent antagonistic effect was observed in mouse cells. In summary, our findings demonstrate species and concentration-dependent differences in hormonal responses to 1,25(OH)2D3 and a complex interplay among PTH, dexamethasone, and 1,25(OH)2D3.