Calcitonin inhibits proton extrusion in resorbing rat osteoclasts via protein kinase A

Although calcitonin is well known to be a potent inhibitor of bone resorption, it remains unknown how it regulates osteoclastic H⁺ transport. In this study, we examined the effects of calcitonin on H⁺ extrusion in cultured rat resorbing osteoclasts using an intracellular pH (pHi) indicator, BCECF [2′7′-bis-(2-carboxyethyl)- 5-carboxyfluorescein]. Resorbing osteoclasts were identified by their formation of resorbing pits on calcium phosphate-coated quartz coverslips. Both basal pHi and H⁺ extrusion activity were significantly higher compared to non-resorbing osteoclasts. Two types of H⁺-extruding systems were identified by pharmacological and immunocytochemical means: a bafilomycin-A₁- sensitive and an amiloride-sensitive system [H⁺ extrusion mediated by a vacuolar type proton pump (V-ATPase) and by a Na⁺/H⁺ exchanger (NHE), respectively]. Calcitonin inhibited both H⁺ extrusion activities in a dose-dependent manner and this action was mimicked by protein kinase A (PKA) activators, but not by protein kinase C (PKC) activators. Pretreatment with PKA inhibitors completely suppressed calcitonin-induced inhibition, whereas neither PKC inhibitors nor calcium chelators suppressed it. These results indicate that calcitonin inhibits H⁺ extrusion generated by V-ATPase and NHE via PKA activation. These inhibitory mechanisms of H⁺ transport by calcitonin are important for the regulation of bone resorption.