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Regulation of intracellular sodium and ph by the electrogenic proton pump in frog rana esculenta skin


, : Regulation of intracellular sodium and ph by the electrogenic proton pump in frog rana esculenta skin. Pfluegers Archiv European Journal of Physiology 406(4): 362-366

We have investigated the role of the electrogenic hydrogen ion pump in the regulation of intracellular sodium ion activity .**GRAPHIC**. and intracellular pH (pHi) in frog skin epithelial cells using double-barreled ion sensitive microelectrodes. When Rana esculenta skin is mounted in an Ussing chamber and bathed in 1 mM Na2SO4 buffered to pH 7.34 with imidazole on the apical side and in normal Ringer on the serosal side, the apical addition of the carbonic anhydrase inhibitor ethoxzolamide (10-4 M) blocks net H+ ion excretion and Na absorption, producing a depolarization of 25-30 mV of the apical membrane potential (.psi.mc). We demonstrate that these changes are accompanied by a fall in .**GRAPHIC**. from 6.2 .+-. 0.5 mmol/l to 3.4 .+-. 0.6 mmol/l and an increase in pHi from 7.20 .+-. 0.03 to 7.38 .+-. 0.08 (n = 12 skins). Voltage clamping .psi.mc to its control value in the presence of ethoxzolamide restored .**GRAPHIC**. but the pHi remained alkaline. Furthermore, the fall in .**GRAPHIC**. produced by ethoxzolamide could be mimicked by voltage clamping .psi.mc towards the value of the Nernst potential for Na at the apical membrane. These results indicate that the maintenance of the cellular Na+ transport pool is dependent on a favourable electrical driving force and counter-current generated by an electrogenic H+ pump at the apical membrane. Addition of amiloride (10-5 mol/l) or substition of external Na+ by Mg2+ or K+ caused a hyperpolarization of .psi.mc and a fall in .**GRAPHIC**. These effects were accompanied by an inhibition of H+ excretion and a fall in pHi of 0.14 .+-. 0.08 units (n = 6 skins). However, when the effect of Na+ transport inhibition on .psi.mc was prevented by imposing a voltage clamp no effects of amiloride on H+ excretion or pHi could be reproduced in control skins by voltage clamping .psi.mc to -100 mV. The metabolic inhibitors vanadate (1 mmol/l) and di-cyclo hexyl carbodiimide (5 .times. 10-5 mol/l) inhibited H+ excretion and decreased pHi from 7.28 .+-. 0.08 to 7.02 .+-. 0.06 and from 7.30 .+-. 0.06 to 7.12 .+-. 0.05 (n = 6 skins), respectively. These results indicate that an apical membrane H+ ATPase plays a role in regulating pHi and the mechanism is sensitive to membrane potential.

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