Different calcium sensitivity in osteoclasts on glass and on bone and maintenance of cytoskeletal structures on bone in the presence of high extracellular calcium

The sensitivity of rat osteoclasts to increased extracellular calcium concentrations (Ca²⁺]_e) was investigated by single cell measurements of free cytosolic calcium concentrations (Ca²⁺]_i), by changes in microfilament organization of resorbing osteoclasts, and by in vitro bone resorption assays. Osteoclasts cultured on glass and on bone showed clear differences in their responses, as in 44% and 52% of osteoclasts on glass but in only 21% and 25% of osteoclasts on bone Ca²⁺]_i increased when Ca²⁺]_e was increased from 2 mM to 6 or 10 mM via perfusion, respectively. Bone resorption was inhibited without changes in the osteoclast numbers only by 10 mM Ca²⁺]_e in 2 day cultures. Furthermore, there were no changes in the organization of microfilament structures in resorbing osteoclasts after increased Ca²⁺]_e (up to 20 mM Ca²⁺]_e, 30 min incubation). These results suggest that the sensitivity of osteoclasts to increased Ca²⁺]_e is dependent on their activation phase (resting/migrating vs. resorbing) and that resorbing osteoclasts are not sensitive to increased Ca²⁺]_e or that the sensing system cannot be reached in polarized resorbing osteoclasts. In contrast, increasing Ca²⁺]_i through the use of calcium ionophores dispersed specific microfilament structures at the sealing zone transiently in a few minutes. This shows that Ca²⁺]_i is used as a signaling mechanism to inactivate osteoclasts, with a similar end result on microfilament structures at the sealing zone as caused by increased concentration of cAMP and activation of protein kinase C. © 1996 Wiley-Liss, Inc.