Osteoporosis is a well-recognized complication of long-term heparin use. However, the mechanisms by which heparin can influence bone metabolism are unclear. We report here that unfractionated heparin stimulates the process of bone resorption and that the low molecular weight heparins (LMWHs), enoxaparin, fragmin, logiparin, and ardeparin produce significantly less calcium loss than unfractionated heparin. To assess calcium loss from bone, we quantified the release of 45Ca into the culture medium of fetal rat calvaria. 45Ca release was increased in a dose-dependent manner by the addition of either unfractionated heparin or the LMWHs; but more than 50-fold higher LMWH concentrations were required to obtain an equivalent effect to unfractionated heparin. Thus, at concentration > or = 2 micrograms/mL (0.35 anti-Xa units/mL), unfractionated heparin stimulated 45Ca release 1.53 +/- 0.06 fold. 45Ca release was increased to a similar extent by the addition of either 10(- 7) mol/L parathyroid hormone (PTH) or 10(-6) mol/L 1,25 dihydroxyvitamin D3 (1,25 Vit D3). In contrast to unfractionated heparin, LMWH concentrations > or = 100 micrograms/mL (> or = 14.0 anti- Xa units/mL) were required before maximum isotope release was observed. At concentrations well above therapeutic levels, the LMWHs stimulated 45Ca release by only 1.25 /+- 0.01-fold. Heparins with high and low antithrombin III affinities stimulated 45Ca release equally well. Both size and sulfation were found to be major determinants of heparin's ability to promote isotope release. Thus, the ability of defined heparin fragments to stimulate 45Ca release correlated with their molecular weight, and after N-desulfation the ability of heparin to induce isotope release was greatly diminished. Dermatan sulfate had no effect on 45Ca release. We conclude that size and sulfation are major determinants of heparin's ability to promote bone resorption and that the risk of heparin-induced osteoporosis may be reduced by the use of LMWH preparations.