Calcium-sensing receptor regulation of PTH-inhibitable proximal tubule phosphate transport

Inorganic phosphate (P-i) is absorbed by proximal tubules through a cellular pathway that is inhibited by parathyroid hormone (PTH). The calcium-sensing receptor (CaSR) is expressed on apical membranes of proximal tubules. In the present studies, we determined the effect of luminal and/or basolateral PTH on phosphate absorption and tested the hypothesis that CaSR activation blocks PTH-inhibitable phosphate absorption. Single proximal S3 tubules were dissected from the kidneys of mice and studied by the Burg technique. Tubules were bathed with DMEM culture media supplemented with 6% BSA and perfused with an ultrafiltrate prepared from the bathing solution. P-33 and FITC-inulin were added to the luminal perfusate to measure phosphate absorption (J(Pi)) and fluid absorption (J(v)), respectively. J(Pi) averaged 2.9 pmol . min(-1) . mm(-1) under control conditions and decreased by 20% upon addition of serosal PTH. PTH had no effect on Jv. Inclusion of PTH in the luminal perfusate reduced J(Pi) to 2.1 pmol . min(-1) . mm(-1). Combined addition of PTH to perfusate and bathing solutions reduced JPi to 1.5 pmol . min(-1) . mm(-1) without affecting J(v). Indirect immunofluorescence studies revealed abundant PTH receptor (PTH1R) expression on brush-border membranes, with lower amounts on basolateral membranes. CaSRs were localized primarily, but not exclusively, to brush-border membranes. CaSR activation with luminal Gd3+ abolished the inhibitory action of PTH on J(Pi). Addition of Gd3+ to the serosal bathing solution had no effect on PTH-sensitive J(Pi). Gd3+ did not affect basal, i.e., PTH-independent J(Pi). Gd3+ had no effect on J(v) when added to lumen or bath. Dopamine-inhibitable J(Pi) was not affected by Gd3+. Experiments with proximal-like opossum kidney cells showed that elevated extracellular Ca2+ or NPS R467, a type II calcimimetic, inhibited PTH action on P-i uptake. In conclusion, PTH1Rs are expressed on apical and basolateral membranes of mouse proximal tubules. Stimulating apical or basolateral PTH1R inhibits phosphate absorption. CaSR activation specifically regulates PTH-suppressible phosphate absorption.