ACELL June 47/6

Miyata, Yukio, Shigeaki Muto, Satoru Yanagiba, and Yasushi Asano. Extracellular Cl2 modulates shrinkageinduced activation of Na1/H1 exchanger in rat mesangial cells. Am J Physiol Cell Physiol 278: C1218–C1229, 2000.—To examine the effect of hyperosmolality on Na1/H1 exchanger (NHE) activity in mesangial cells (MCs), we used a pHsensitive dye, 28,78-bis(2-carboxyethyl)-5(6)-carboxyfluorescein-AM, to measure intracellular pH (pHi) in a single MC from rat glomeruli. All the experiments were performed in CO2/HCO3 -free HEPES solutions. Exposure of MCs to hyperosmotic HEPES solutions (500 mosmol/kgH2O) treated with mannitol caused cell alkalinization. The hyperosmolalityinduced cell alkalinization was inhibited by 100 μM ethylisopropylamiloride, a specific NHE inhibitor, and was dependent on extracellular Na1. The hyperosmolality shifted the Na1dependent acid extrusion rate vs. pHi by 0.15–0.3 pH units in the alkaline direction. Removal of extracellular Cl2 by replacement with gluconate completely abolished the rate of cell alkalinization induced by hyperosmolality and inhibited the Na1-dependent acid extrusion rate, whereas, under isosmotic conditions, it caused no effect on Na1-dependent pHi recovery rate or Na1-dependent acid extrusion rate. The Cl2-dependent cell alkalinization rate under hyperosmotic conditions was partially inhibited by pretreatment with 5-nitro-2-(3phenylpropylamino)benzoic acid, DIDS, and colchicine. We conclude: 1) in MCs, hyperosmolality activates NHE to cause cell alkalinization, 2) the acid extrusion rate via NHE is greater under hyperosmotic conditions than under isosmotic conditions at a wide range of pHi, 3) the NHE activation under hyperosmotic conditions, but not under isosmotic conditions, requires extracellular Cl2, and 4) the Cl2-dependent NHE activation under hyperosmotic conditions partly occurs via Cl2 channel and microtubule-dependent processes.