Inhibition of NA+/K+ ATPase under hypertonic conditions in rat mast cells

Ionic fluxes that contribute to changes in membrane potential and variations of pHi (intracellular pH) are not well known in mast cells, although they can be important in the stimulus-secretion coupling. Cellular volume regulation implies changes in the concentration of intracellular ions, such as sodium and potassium and volume changes can be imposed varying the tonicity of the medium. We studied the physiology of sodium and examined the effect of ouabain on [Na-22] entry in mast cells in isotonic and hypertonic media. We also recorded changes in membrane potential and pHi using the fluorescent dyes bis-oxonol (Bis-(1,3-diethylthiobarbituric acid) trimethine oxonol) and BCECF (2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein acetoxymethyl ester) in hypertonic conditions. The results show that [Na-22] influx increases four fold in hypertonic solutions and it is mediated mainly by an amiloride-sensitive Na+/H+ exchanger. This transporter is involved in the shrinkage-activated cellular alkalinization and the pHi recovery is accelerated by inhibition of the Na+/K+ ATPase with ouabain in the absence of extracellular calcium. Under hypertonic conditions Na-22 influx is apparently not increased by ouabain, while the Na+/K+ ATPase inhibitor clearly increases [Na-22] uptake and also induces membrane depolarization in isotonic conditions. All together, these findings suggest that Na+/K+ ATPase is partially inhibited in hypertonic conditions.