title: Intracellular ClC-3 chloride channels promote bone resorption in vitro through organelle acidification in mouse osteoclasts.
abstract: ClC-7 Cl(-) channels expressed in osteoclasts are important for bone resorption since it has been shown that disruption of the ClCN7 gene in mice leads to severe osteopetrosis. We have previously reported that Cl(-) currents recorded from mouse osteoclasts resemble those of ClC-3 Cl(-) channels. The aim of the present study was to determine the expression of ClC-3 channels in mouse osteoclasts and their functional role during bone resorption. We detected transcripts for both ClC-7 and ClC-3 channels in mouse osteoclasts by RT-PCR. The expression of ClC-3 was confirmed by immunocytochemical staining. Mouse osteoclasts lacking ClC-3 Cl(-) channels (ClC-3(-/-) osteoclasts) derived from ClCN3 gene-deficient mice (ClC-3(-/-)) showed lower bone resorption activity compared with ClC-3+/+ osteoclasts derived from wild-type mice (ClC-3+/+). Treatment of ClC-3+/+ osteoclasts with small interfering RNA (siRNA) against ClC-3 also significantly reduced bone resorption activity. Electrophysiological properties of basal and hypotonicity-induced Cl(-) currents in ClC-3(-/-) osteoclasts did not differ significantly from those in ClC-3+/+ osteoclasts. Using immunocytochemistry, ClC-3 was colocalized with lysosome-associated membrane protein 2. Using pH-sensitive dyes, organelle acidification activity in ClC-3(-/-) osteoclasts was weaker than in ClC-3+/+ osteoclasts. Treatment of ClC-3+/+ osteoclasts with siRNA against ClC-3 also reduced the organelle acidification activity. In conclusion, ClC-3 Cl(-) channels are expressed in intracellular organelles of mouse osteoclasts and contribute to osteoclastic bone resorption in vitro through organelle acidification.
authors.affiliation: Dept. of Physiological Science and Molecular Biology, Fukuoka Dental College, 2-15-1 Tamura, Sawara-ku, Fukuoka 814-0193, Japan. fujipi@college.fdcnet.ac.jp
authors.forename: Fujio
authors.initials: F
authors.lastname: Okamoto
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Kajiya
authors.forename: Kazuko
authors.initials: K
authors.lastname: Toh
authors.forename: Shinichi
authors.initials: S
authors.lastname: Uchida
authors.forename: Momono
authors.initials: M
authors.lastname: Yoshikawa
authors.forename: Sei
authors.initials: S
authors.lastname: Sasaki
authors.forename: Mizuho A
authors.initials: MA
authors.lastname: Kido
authors.forename: Teruo
authors.initials: T
authors.lastname: Tanaka
authors.forename: Koji
authors.initials: K
authors.lastname: Okabe
mesh_terms: D000818:Animals; D001862:Bone Resorption / Q000378:metabolism*; D002478:Cells, Cultured; D018118:Chloride Channels / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D002712:Chlorides / Q000378:metabolism*; D003804:Dentin / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007150:Immunohistochemistry; D052119:Lysosomal-Associated Membrane Protein 2 / Q000378:metabolism; D008564:Membrane Potentials; D051379:Mice; D008813:Mice, Inbred ICR; D018345:Mice, Knockout; D015388:Organelles / Q000378:metabolism*; D010010:Osteoclasts / Q000378:metabolism*; D011485:Protein Binding; D034622:RNA Interference; D012333:RNA, Messenger / Q000378:metabolism; D034741:RNA, Small Interfering / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction
chemical_list: D018118:Chloride Channels; D002712:Chlorides; C115376:ClC-3 channel; C490803:Clcn7 protein, mouse; D052119:Lysosomal-Associated Membrane Protein 2; D012333:RNA, Messenger; D034741:RNA, Small Interfering