Surface binding and clearance of calcitonin by avian osteoclasts.

Osteoclasts, isolated from chick tibias and maintained in culture for 4-6 days, were treated with biotinylated calcitonin followed by fluorescent avidin. In 2 min the entire cell surface fluoresced, by 7.5 min fluorescence was centrally clustered and by 10 min fluorescence had diminished to background levels. Specific binding was blocked by excess unlabeled calcitonin and not influenced by unrelated peptide. The results show that avian osteoclasts bind calcitonin specifically and that the bound calcitonin is rapidly cleared from the cell surface, which indicates involvement of receptor-mediated endocytosis.

Effect of estrogen on acidification in osteoclasts.

Isolated chicken osteoclasts were neutralized with NH4Cl, then recovery of acid formation was monitored by measuring acridine orange fluorescence over a period of 4 hrs. Estradiol-17 beta and diethylstilbestrol (DES), in nanomolar amounts, blocked acidification 20% to 60%. Effects were detectable in 30 min and maximal by one hr. Estradiol-17 alpha was ineffective. This action of estradiol is believed to be mediated at the plasma membrane since the response was rapid and was not affected by cycloheximide inhibition of protein synthesis. Stimulation of acidification by PTH and cyclic AMP was blocked by estradiol. Estradiol was inhibitory to the same extent as calcitonin; these effects were not additive. Estradiol-17 beta and DES, in micromolar but not in nanomolar amounts, blocked proton pumping in isolated plasma membrane vesicles.