Down-regulation of parathyroid hormone (PTH) receptors in cultured bone cells is associated with agonist-specific intracellular processing of PTH-receptor complexes.
Exposure of cultured embryonic chicken bone cells to the PTH agonists bovine (b) PTH-(1-34) and [8Nle, 18Nle, 34Tyr]bPTH-(1-34)amide [bPTH-(1-34)A] reduces the subsequent cAMP response to the hormone and decreases the specific binding of 125I-labeled PTH to these cultures. To determine whether PTH receptor down-regulation in cultured bone cells is mediated by cellular internalization of PTH-receptor complexes, we measured the uptake of [125I]bPTH-(1-34) into an acid-resistant compartment. Uptake of radioactivity into this compartment was inhibited by incubating cells at 4 C with phenylarsineoxide and unlabeled bPTH-(1-34). Tracer uptake into the acid-resistant compartment at any time was directly proportional to total cell binding at 22 C. Thus, it is likely that PTH-receptor complexes are internalized by bone cells. This mechanism may explain the loss of cell surface receptors after PTH pretreatment. To determine whether internalized PTH-receptor complexes are reinserted into the plasma membrane, we measured PTH binding and PTH stimulation of cAMP production after cells were exposed to monensin, a known inhibitor of receptor recycling. Monensin (25 microM) had no effect on PTH receptor number or affinity and did not alter PTH-stimulated cAMP accumulation. However, monensin (25 microM) incubated with cells pretreated with various concentrations of bPTH-(1-34) for 1 h potentiated the effect of the hormone to reduce subsequent [125I]bPTH-(1-34) binding and PTH-stimulated cAMP accumulation by more than 2 orders of magnitude. Chloroquine also potentiated PTH-induced down-regulation of PTH receptors. By contrast, neither agent influenced PTH binding or PTH-stimulated cAMP production in cells pretreated with the antagonist bPTH-(3-34)A. Thus, monensin potentiated PTH receptor loss only in cells pretreated with PTH agonists, indicating that antagonist-occupied receptors may be processed differently from agonist-occupied receptors in bone cells. The data further suggest that the attenuation of PTH stimulation of cAMP production in treated bone cells may be, at least in part, due to receptor-mediated endocytosis of the hormone.
D000818:Animals; D001152:Arsenicals / Q000494:pharmacology; D001842:Bone and Bones / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D002642:Chick Embryo; D002738:Chloroquine / Q000494:pharmacology; D000242:Cyclic AMP / Q000096:biosynthesis; D004357:Drug Synergism; D008985:Monensin / Q000494:pharmacology; D010281:Parathyroid Hormone / Q000378:metabolism / Q000494:pharmacology; D010446:Peptide Fragments / Q000378:metabolism / Q000494:pharmacology; D011956:Receptors, Cell Surface / Q000187:drug effects / Q000378:metabolism*; D018016:Receptors, Parathyroid Hormone
D001152:Arsenicals; D010281:Parathyroid Hormone; D010446:Peptide Fragments; D011956:Receptors, Cell Surface; D018016:Receptors, Parathyroid Hormone; C029341:oxophenylarsine; C025076:parathyroid hormone (1-34), bovine; D002738:Chloroquine; D008985:Monensin; D000242:Cyclic AMP