Previous in vitro studies have shown that salmon calcitonin had direct effects to increase parameters associated with embryonic chicken bone formation and to increase mouse and chicken osteoblast-line cell proliferation. The current studies demonstrate increased cell proliferation (i.e., [3H]-thymidine incorporation into DNA and tetrazolium salt reduction/deposition) in the osteoblastic murine cell line MC-3T3-E1 in response to salmon calcitonin (P less than 0.005) and to human calcitonin (P less than 0.005), but not to human calcitonin gene-related peptide. The current studies also show that salmon calcitonin increased several indices of murine bone formation. We found that 72 hours of exposure to salmon calcitonin [at 5 mU/ml-about 0.37 nM; mU/ml = milliunits of calcitonin activity/ml incubation medium (at 4,000 U/mg protein)] increased net 45Ca deposition (121% of control, P less than 0.05), net [3H]-proline incorporation 149% of control, P less than 0.001), and alkaline phosphatase activity (146% of control, P less than 0.01), in neonatal mouse half-calvaria. The calcitonin-dependent increase in alkaline phosphatase activity was not affected by co-incubation with 1 nM parathyroid hormone. Co-incubation with fluoride (which also increased net [3H]-proline incorporation and alkaline phosphatase activity in neonatal mouse half-calvaria, P less than 0.05, for each) enhanced the osteogenic response to low-dose calcitonin, (i.e., co-incubation with fluoride shifted the biphasic calcitonin dose-response curve to a range of lower calcitonin concentrations). The calcitonin-fluoride combinations had proportional effects on net [3H]-proline incorporation and alkaline phosphatase in the treated mouse calvaria (r = 0.78, P less than 0.005).