Immunocytochemical localization of vacuolar H+-ATPase and Cl--HCO3- anion exchanger (erythrocyte band-3 protein) in avian osteoclasts: effect of calcium-deficient diet on polar expression of the H+-ATPase pump.
Osteoclasts attach to the bone surface and resorb bone by secreting protons into an isolated subosteoclastic compartment. Previous studies have shown the presence of a vacuolar type H+-ATPase, and a functional Cl--HCO3- anion exchanger in the osteoclast. In the present studies, using a monoclonal antibody to the 31-kDa subunit of H+-ATPase and a rabbit antiserum to the erythrocyte band-3 protein (Cl--HCO3- anion exchanger) we have immunocytochemically localized the respective pumps in bone sections obtained from chickens fed a normal or a calcium-deficient diet for 4 weeks. Our results indicate that although H+-ATPase is either evenly distributed throughout the osteoclast or is more polarized at its ruffled membrane juxtaposed to the bone surface, the band-3 protein immunoreactivity is always localized to the plasma membrane which is not attached to the bone surface (basolateral membrane). Four weeks of a calcium-deficient diet resulted in a significant increase in the percentage of osteoclasts that were polarized for the H+-ATPase pump at their ruffled membrane, and a trend toward increased total number of osteoclasts, although the latter did not reach statistical significance (P = 0.09). These changes were not accompanied by a significant increase in the intensity of staining for H+-ATPase. Band-3 protein immunoreactivity was always prominent, limited to the basolateral membrane, and did not alter with calcium-deficient diet or with changes in the degree of H+-ATPase polarization.
Bastani
B
B
Division of Nephrology, St. Louis University Health Sciences Center, 3635 Vista Ave., 9N-FDT, St. Louis, Missouri 63110-0250, USA.
Ross
F P
FP
Kopito
R R
RR
Gluck
S L
SL
D000251:Adenosine Triphosphatases / Q000378:metabolism*; D000818:Animals; D001457:Anion Exchange Protein 1, Erythrocyte / Q000378:metabolism*; D001717:Birds; D001862:Bone Resorption; D002118:Calcium / Q000172:deficiency*; D002645:Chickens; D005260:Female; D005270:Femur Head / Q000166:cytology; D019084:Fluorescent Antibody Technique, Indirect; D010010:Osteoclasts / Q000166:cytology / Q000378:metabolism*; D017494:Proton Pumps; D025262:Vacuolar Proton-Translocating ATPases
D001457:Anion Exchange Protein 1, Erythrocyte; D017494:Proton Pumps; D000251:Adenosine Triphosphatases; D025262:Vacuolar Proton-Translocating ATPases; D002118:Calcium