We examined the effect of nicotine on cellular proliferation, as measured by [3H]thymidine (TdR) incorporation and cell count, and on alkaline phosphatase activity in UMR 106-01 rat osteoblastic osteosarcoma cells. The cells were cultured with varying concentrations of nicotine in serum-free medium for 2 to 72 hours. Nicotine produced a dose-dependent suppression of TdR incorporation, with maximum suppression seen at 10 mM (7% of control); the EC50 for suppression of TdR incorporation was 10 microM. 1 microM nicotine decreased cell number by 20% to 30%. The time course of the effect of 100 microM nicotine on DNA synthesis was measured by TdR incorporation. TdR uptake was measured at 2, 4, 6, 24, 48, and 72 hours. After the addition of nicotine, the following biphasic response in TdR incorporation was observed: a 15% decrease at 2 hours, recovery to near control value at 6 hours, a 27% decrease by 24 hours, and a maximum decrease of 88% by 48 hours. Over a dose range of 1 nM to 10 mM, nicotine produced a dose-dependent increase in alkaline phosphatase activity with maximum stimulation seen at 1 microM (189% of control). We conclude that nicotine suppresses cellular proliferation and stimulates alkaline phosphatase activity in UMR 106-01 osteoblast-like cells. These results may be of significance in the development of osteoporosis and alveolar bone loss associated with the use of tobacco.