Parathyroid hormone(PTH), a major bone hormone, inhihits DNA and collagen syntheses in osteohlast-like cells in vitro, but increase the proliferation of osteoblast in vivo as secn in hyperparathyroidism. On the other hand, insulin is known to increase DNA and collagen syntheses and modify the effects of PTH in osteoblast-like cells. We have examined the effects of PTH and insulin in rat osteosarcoma UMR-l06-01 cells and whether PTH plays a role in the insulin-mediated bone formation. When 1 nM PTH and 10 nM insulin were administered to VMR-l06-01 ceils, the rates of DNA synthesis were 124% and 136% of the untreated control, respectively. When the two hormones were administered serially by exposing to 1 nM PTH for 7 days followed by 10 nM insulin lor 24h, the largest increase was observed. The protein synthesis was also increased remarkahly when the two hormones were aclministered serially: the[3H]-leucine incorporation rates, compared to the control group, were 75% and l62% with PTH ancl insulin administration, respectively, but the rate was 297% with the serial administration of the two. The collaeen synthesis, as measured by the (3H)-proline incorporation rates were 60% and l64% with PTH and insulin administration, respectively, but 351% with serial administration, again showing a dramatic effect. These results showed that 1 nM PTH decreased DNA and collagen syntheses in UMR-l06-01 cells after both a 24h and a more prolonged exposure. Similar exposures to insulin tended to increase the syntheses. The comhination of PTH and insulin tended to increase the syntheses. hut not beyond the effect of insulin alone. However, the sequential administration of PTH and insulin markedly increases ihose rales relative to the simultaneous adminstration of these two hormones. Thus, it is possihle that sequential stimulation of PTH and insulin in hone matrix exerts an synergistic effect on hone formation in vivo.