Previous work demonstrated that parathyroid hormone (PTH) activates the Ca2+/protein kinase C (PKC) system in addition to cAMP production. Therefore, the authors explored the role of cAMP-dependent and Ca2(+)-dependent signals in the regulation of osteoblastic growth and bone resorption. In exponentially growing UMR 106-01 osteogenic sarcoma cells, PTH (10(-7) M) inhibited [3H] thymidine incorporation by 80%. This effect was reproduced by maximal doses of both dibutyryl-cAMP (dbcAMP) and forskolin. The Ca2+ ionophore ionomycin (10(-7) M) had no effect, whereas phorbol 12-myristate 13-acetate (PMA) was slightly mitogenic. The antimitogenic action of dbcAMP was dose-dependent, with ED0.5 at about 3 X 10(-5) M. Ionomycin enhanced this dbcAMP effect at submaximal doses of the cAMP analog. PMA used in combination with both dbcAMP and ionomycin induced further depression of cell proliferation, indicating synergism with cAMP. Both dbcAMP (10(-4) M) and ionomycin (10(-7) M) stimulated 45Ca release from fetal rat limb bones after five days in culture, although the Ca2+ ionophore was less potent. 1-Oleoyl 2-acetyl-glycerol (2 X 10(-6) M) was ineffective alone, and slightly inhibited the 45Ca release produced by the other second messenger analogs in all combinations. The combination of dbcAMP and ionomycin showed a synergistic effect, and fully reproduced PTH effect. In conclusion, PTH signal transduction for control of cell proliferation and bone resorption is mediated mainly by cAMP. Activation of the Ca2+/PKC message system is nevertheless necessary to express a full hormonal response in both cell and organ culture systems.