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Am J Physiol Renal Physiol 283: F399-F406, 2002. First published March 12, 2002; doi:10.1152/ajprenal.00346.2001
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Vol. 283, Issue 3, F399-F406, September 2002

Calcium-sensing receptor regulation of PTH-dependent calcium absorption by mouse cortical ascending limbs

Hiroki I. Motoyama1 and Peter A. Friedman1,2

Departments of 1 Pharmacology and 2 Medicine, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania 15261

Resting Ca2+ absorption by cortical thick ascending limbs (CALs) is passive and proceeds through the paracellular pathway. In contrast, parathyroid hormone (PTH) stimulates active, transcellular Ca2+ absorption (JCa). The Ca2+-sensing receptor (CaSR) is expressed on serosal membranes of CALs. In the present study, we tested the hypothesis that activation of the CAL CaSR indirectly inhibits passive Ca2+ transport and directly suppresses PTH-induced cellular JCa. To test this theory, we measured JCa and Na absorption (JNa) by single perfused mouse CALs. Net absorption was measured microfluorimetrically in samples collected from tubules perfused and bathed in symmetrical HEPES-buffered solutions or those in which luminal Na+ was reduced from 150 to 50 mM. We first confirmed that Gd3+ activated the CaSR by measuring intracellular Ca2+ concentration ([Ca2+]i) in CALs loaded with fura 2. On stepwise addition of Gd3+ to the bath, [Ca2+]i increased, with a half-maximal rise at 30 µM Gd3+. JCa and transepithelial voltage (Ve,) were measured in symmetrical Na+-containing solutions. PTH increased JCa by 100%, and 30 µM Gd3+ inhibited this effect. Ve was unchanged by either PTH or Gd3+. Similarly, NPS R-467, an organic CaSR agonist, inhibited PTH-stimulated JCa without altering Ve. Neither PTH nor Gd3+ affected JNa. Addition of bumetanide to the luminal perfusate abolished JNa and Ve. These results show that CaSR activation directly inhibited PTH-induced transcellular JCa and that cellular Ca2+ and Na+ transport can be dissociated. To test the effect of CaSR activation on passive paracellular Ca2+ transport, JCa was measured under asymmetrical Na conditions, in which passive Ca2+ transport dominates transepithelial absorption. PTH stimulated JCa by 24% and was suppressed by Gd3+. In this setting, Gd3+ reduced Ve by 32%, indicating that CaSR activation inhibited both transcellular and paracellular Ca2+ transport. We conclude that the CaSR regulates both active transcellular and passive paracellular Ca2+ reabsorption but has no effect on JNa by CALs.

calcium transport; thick ascending limbs; kidney; parathyroid hormone; transepithelial voltage


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