PARATHYROID-HORMONE TRANSPORT EFFECTS AND HORMONAL PROCESSING IN PRIMARY CULTURED RAT PROXIMAL TUBULAR CELLS

The development of satisfactory cell culture models for the study of p arathyroid hormone (PTH)-induced inhibition of P(i) transport has prov en difficult. Using subcellular fractionation techniques we investigat ed the response of primary cultures of rat proximal tubular cells to P TH-(1-34). Specific binding of I-125-bPTH-(1-34) occurred at 2-degrees -C. After 5 min of rewarming, trypsin-releasable radioactivity decreas ed from 90 to 50% indicating internalization of the ligand. Cell disru ption, followed by density centrifugation with 17%. Percoll either dir ectly after binding at 2-degrees-C or post-rewarming for 20 min, showe d a shift of I-125 label from the plasma membrane (5'-nucleotidase) to lysosomal fractions (beta-D-glucosaminidase), confirming the sequenti al occurrence of cell surface binding, internalization and transport t o lysosomes of I-125-bPTH-(1-34). Reculture at 37-degrees-C revealed s teady accumulation of trichloroacetic acid-soluble radioactivity in th e medium, indicating degradation of I-125-bPTH-(1-34). Phosphate trans port in the absence of sodium was minimal. Incubation of the cells wit h bPTH-(1-34) resulted in up to 50% inhibition of sodium-dependent pho sphate transport. Prior phosphate depletion abrogated the response to PTH.