Identification and characterization of a sodium/calcium exchanger, NCX-1, in osteoclasts and its role in bone resorption

We provide the first demonstration for a Na⁺/Ca²⁺ exchanger, NCX-1, in the osteoclast. We speculate that by using Na⁺ exchange, NCX-1 couples H⁺ extrusion with Ca²⁺ fluxes during bone resorption. Microspectrofluorimetry of fura-2-loaded osteoclasts revealed a rapid and sustained, but reversible, cytosolic Ca²⁺ elevation upon Na⁺ withdrawal. This elevation was abolished by the cytosolic introduction (by gentle permeabilization) of a highly specific Na⁺/Ca²⁺ exchange inhibitor peptide, XIP, but not its inactive analogue, sXIP. Confocal microscopy revealed intense plasma membrane immunofluorescence with an isoform-specific monoclonal anti-NCX-1 antibody applied to gently permeabilized osteoclasts. Electrophysiological studies using excised outside-in membrane patches showed a low-conductance, Na⁺-selective, dichlorobenzamil-sensitive, amiloride-insensitive channel that we tentatively assigned as being an NCX. Finally, to examine for physiological relevance, an osteoclast resorption (pit) assay was performed. There was a dramatic reduction of bone resorption following NCX-1 inhibition by dichloroben- zamil and XIP (but not with S-XIP). Together, the results suggest that a functional NCX, likely NCX-1, is involved in the regulation of osteoclast cytosolic Ca²⁺ and bone resorption.