Extracellular Cl(-) modulates shrinkage-induced activation of Na(+)/H(+) exchanger in rat mesangial cells.
To examine the effect of hyperosmolality on Na(+)/H(+) exchanger (NHE) activity in mesangial cells (MCs), we used a pH-sensitive dye, 2',7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein-AM, to measure intracellular pH (pH(i)) in a single MC from rat glomeruli. All the experiments were performed in CO(2)/HCO(-)(3)-free HEPES solutions. Exposure of MCs to hyperosmotic HEPES solutions (500 mosmol/kgH(2)O) treated with mannitol caused cell alkalinization. The hyperosmolality-induced cell alkalinization was inhibited by 100 microM ethylisopropylamiloride, a specific NHE inhibitor, and was dependent on extracellular Na(+). The hyperosmolality shifted the Na(+)-dependent acid extrusion rate vs. pH(i) by 0.15-0.3 pH units in the alkaline direction. Removal of extracellular Cl(-) by replacement with gluconate completely abolished the rate of cell alkalinization induced by hyperosmolality and inhibited the Na(+)-dependent acid extrusion rate, whereas, under isosmotic conditions, it caused no effect on Na(+)-dependent pH(i) recovery rate or Na(+)-dependent acid extrusion rate. The Cl(-)-dependent cell alkalinization rate under hyperosmotic conditions was partially inhibited by pretreatment with 5-nitro-2-(3-phenylpropylamino)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 pH(i), 3) the NHE activation under hyperosmotic conditions, but not under isosmotic conditions, requires extracellular Cl(-), and 4) the Cl(-)-dependent NHE activation under hyperosmotic conditions partly occurs via Cl(-) channel and microtubule-dependent processes.
Miyata
Y
Y
Department of Nephrology, Jichi Medical School, Minamikawachi, Kawachi, Tochigi 329-0498, Japan.
Muto
S
S
Yanagiba
S
S
Asano
Y
Y
D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid / Q000494:pharmacology; D000584:Amiloride / Q000031:analogs & derivatives / Q000494:pharmacology; D000818:Animals; D048429:Cell Size; D002478:Cells, Cultured; D002712:Chlorides / Q000494:pharmacology*; D003078:Colchicine / Q000494:pharmacology; D005452:Fluoresceins / Q000494:pharmacology; D005920:Glomerular Mesangium / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D006863:Hydrogen-Ion Concentration; D006982:Hypertonic Solutions; D008297:Male; D009579:Nitrobenzoates / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley; D017923:Sodium-Hydrogen Exchangers / Q000187:drug effects / Q000502:physiology*
D002712:Chlorides; D005452:Fluoresceins; D006982:Hypertonic Solutions; D009579:Nitrobenzoates; D017923:Sodium-Hydrogen Exchangers; C057433:2',7'-bis-(2-carboxyethyl)-5(6)-carboxyfluorescein acetoxymethyl ester; C058176:5-nitro-2-(3-phenylpropylamino)benzoic acid; D000584:Amiloride; D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid; D003078:Colchicine; C039614:ethylisopropylamiloride