Tohoku J. Exp. Med., 2006, 210(4)

Chloride-Dependent Intracellular pH Regulation via Extracellular Calcium-Sensing Receptor in the Medullary Thick Ascending Limb of the Mouse Kidney

ULVIYYA FIZULI ASLANOVA,¹ TETSUJI MORIMOTO,¹ ELNUR ILHAM FARAJOV,² NAONORI KUMAGAI,¹ MINAKO NISHINO,¹ NORIKO SUGAWARA,¹ ATSUSHI OHSAGA,³ YOSHIO MARUYAMA,³ SHIGERU TSUCHIYA,¹ SHORI TAKAHASHI⁴ and YOSHIAKI KONDO²

¹Department of Pediatrics, Tohoku University Graduate School of Medicine, Sendai, Japan, ²Department of Medical Informatics, Tohoku University Graduate School of Medicine, Sendai, Japan, ³Department of Physiology I, Tohoku University Graduate School of Medicine, Sendai, Japan, ⁴Department of Pediatrics, Nihon University School of Medicine, Tokyo, Japan

The extracellular calcium-sensing receptor (CaSR) located in either luminal or basolateral cell membranes of various types of renal tubules including proximal tubules, Henle's loop and collecting ducts has been thought to play a fundamental role in electrolyte metabolism. To further identify the physiological roles of the CaSR, we examined the effects of Ca²⁺ and calcimimetics neomycin (Neo), gentamicin and gadolinium chloride (Gd³⁺) on the intracellular pH (pHi) of in vitro microperfused mouse medullary thick ascending limb (mTAL) cells of Henle's loop, by loading the cells with fluorescent pH indicator 2',7'-bis-(2-carboxyethyl)-5-(and-6)-carboxyfluorescein and measuring the ratio of fluorescence emission at 530 nm after exciting the dye at 490 and 440 nm. In a steady-state condition in Hepes-buffered solution, the pHi in the mTALs was 7.29 ± 0.04 (n = 9). A concentration of 200 μmol/l Neo in the basolateral side decreased the pHi after 1 min by −0.13 ± 0.02 (n = 34, p < 0.0001). The other calcimimetics showed similar effects on pHi, whereas none of these calcimimetics in the lumen affected pHi. Na⁺ removal or the inhibition of Na⁺ and proton transport with amiloride, bumetanide, or bafilomycin did not eliminate the effect of Neo on pHi. On the other hand, Cl⁻ removal clearly eliminated the Neo-induced pHi decrease (−0.06 ± 0.01 vs −0.00 ± 0.05 in Cl⁻ removal, n = 4, p < 0.003). Thus, we have demonstrated for the first time that the CaSR is involved in the regulation of the pHi in the mTAL and requires Cl⁻ to exert its effect.

keywords —— intracellular pH; renal medulla; chloride; amiloride; BCECF

© 2006 Tohoku University Medical Press

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Tohoku J. Exp. Med., 2006, 210, 291-300

Correspondence: Tetsuji Morimoto, M.D., Ph.D., Department of Pediatrics, Tohoku University Graduate School of Medicine, 1-1 Seiryo-machi, Aoba-ku, Sendai 980-8574, Japan.

e-mail: morimoto-thk@umin.ac.jp