title: Parathyroid hormone transport effects and hormonal processing in primary cultured rat proximal tubular cells.
abstract: The development of satisfactory cell culture models for the study of parathyroid hormone (PTH)-induced inhibition of Pi transport has proven difficult. Using subcellular fractionation techniques we investigated the response of primary cultures of rat proximal tubular cells to PTH-(1-34). Specific binding of 125I-bPTH-(1-34) occurred at 2 degrees C. After 5 min of rewarming, trypsin-releasable radioactivity decreased from 90 to 50%, indicating internalization of the ligand. Cell disruption, followed by density centrifugation with 17% Percoll either directly after binding at 2 degrees C or post-rewarming for 20 min, showed a shift of 125I label from the plasma membrane (5'-nucleotidase) to lysosomal fractions (beta-D-glucosaminidase), confirming the sequential occurrence of cell surface binding, internalization and transport to lysosomes of 125I-bPTH-(1-34). Reculture at 37 degrees C revealed steady accumulation of trichloroacetic acid-soluble radioactivity in the medium, indicating degradation of 125I-bPTH-(1-34). Phosphate transport in the absence of sodium was minimal. Incubation of the cells with bPTH-(1-34) resulted in up to 50% inhibition of sodium-dependent phosphate transport. Prior phosphate depletion abrogated the response to PTH.
authors.affiliation: Department of Medicine, Tulane University Medical Center, New Orleans, LA 70112.
authors.forename: R M
authors.initials: RM
authors.lastname: O'Donovan
authors.forename: C C
authors.initials: CC
authors.lastname: Widnell
authors.forename: T C
authors.initials: TC
authors.lastname: Chen
authors.forename: J B
authors.initials: JB
authors.lastname: Puschett
mesh_terms: D000818:Animals; D001665:Binding Sites; D001692:Biological Transport; D002458:Cell Fractionation; D002478:Cells, Cultured; D007687:Kidney Tubules, Proximal / Q000166:cytology / Q000378:metabolism*; D010281:Parathyroid Hormone / Q000378:metabolism*; D010446:Peptide Fragments / Q000378:metabolism*; D010710:Phosphates / Q000378:metabolism; D051381:Rats
chemical_list: D010281:Parathyroid Hormone; D010446:Peptide Fragments; D010710:Phosphates; C025076:parathyroid hormone (1-34), bovine