Characterization of Na+-H+ antiport in type II alveolar epithelial cells.
The presence of a Na+-H+ exchange pathway in the plasma membrane of type II alveolar epithelial cells was explored using the pH-sensitive fluorescent probe 2,7-biscarboxyethyl-5,6-carboxyfluorescein (BCECF) to monitor changes in cytosolic pH. Freshly prepared pneumocytes suspended in medium at pH 7.4 had an intracellular pH of 7.07 +/- 0.07. Acid-loaded cells equilibrated in sodium-free buffer showed rapid cytoplasmic alkalinization when exposed to sodium. This response to sodium was inhibited greater than 90% by 10(-4) M amiloride. The presence of the K+ ionophore, valinomycin, had no effect on the rate of Na+-dependent alkalinization, indicating the electroneutrality of the system. Li+ partially supported the alkalinization process, but other monovalent cations, notably K+, Rb+, and Cs+, were without effect. Kinetic analysis for Na+ at the external binding site yielded KNat (dissociation constant) = 62 +/- 3 mM. Hill equation analysis of the data derived a Hill coefficient (n) = 1.2 +/- 0.1 for Na+, consistent with a 1:1 stoichiometry for Na+ and H+ for the transporter. The Ki for amiloride inhibition of proton efflux at the external locus was 0.45 microM. These findings define the transport pathway as Na+-H+ antiport, with kinetic parameters somewhat similar to those described for other cell types. Antiport activity was detected at intracellular pH (pHi) values of 6.8 or below, with no activity observed at pHi 7.0-7.2. It is suggested that Na+-H+ exchange is a major mechanism whereby pneumocytes recover from an acid load and that this transport pathway may play an important role in vectorial reabsorption of Na+ from the alveolar air spaces.
D000584:Amiloride / Q000037:antagonists & inhibitors; D000818:Animals; D002138:Calibration; D002352:Carrier Proteins / Q000378:metabolism*; D004847:Epithelial Cells; D004848:Epithelium / Q000378:metabolism; D005452:Fluoresceins; D006863:Hydrogen-Ion Concentration; D007425:Intracellular Membranes / Q000378:metabolism; D007700:Kinetics; D008297:Male; D011522:Protons; D011650:Pulmonary Alveoli / Q000166:cytology / Q000378:metabolism*; D051381:Rats; D011919:Rats, Inbred Strains; D017923:Sodium-Hydrogen Exchangers
D002352:Carrier Proteins; D005452:Fluoresceins; D011522:Protons; D017923:Sodium-Hydrogen Exchangers; D000584:Amiloride; C043829:2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein