Sensing of extracellular cations in CasR-deficient osteoblasts - Evidence for a novel cation-sensing mechanism

We isolated osteoblastic cell lines from wild-type (CasR(+/+)) and receptor null (CasR(-/-)) mice to investigate whether CasR is present in osteoblast s and accounts for their responses to extracellular cations, Osteoblasts fr om both CasR(+/+) and CasR(-/-) mice displayed an initial period of cell re plication followed by a culture duration-dependent increase in alkaline pho sphatase activity, expression of osteocalcin, and mineralization of extrace llular matrix. In addition, a panel of extracellular cations, including alu minum and the CasR agonists gadolinium and calcium, stimulated DNA synthesi s, activated a transfected serum response element-luciferase reporter const ruct, and inhibited agonist-induced cAMP in CasR(-/-) osteoblasts. The func tional responses to these cations were identical in CasR(+/+) and CasR(-/-) osteoblasts, Thus, the absence of CasR alters neither the maturational pro file of isolated osteoblast cultures nor their in vitro responses to extrac ellular cations. In addition, CasR transcripts could not be detected by rev erse transcription-polymerase chain reaction with mouse specific primers in either CasR(+/+) or CasR(-/-) osteoblasts, and immunoblot analysis with a CasR-specific antibody was negative for CasR protein expression in osteobla sts, The presence of a cation-sensing response in osteoblasts from CasR(-/- ) mice indicates the existence of a novel osteoblastic extracellular cation -sensing mechanism.