HCO(-3)-dependent pHi recovery and overacidification induced by NH+4 pulse in rat lung alveolar type II cells: HCO(-3)-dependent NH3 excretion from lungs?

Intracellular pH (pHi) after the NH+4 pulse addition and its removal were measured in isolated alveolar type II cells (ATII cells) using BCECF fluorescence. In the absence of HCO(-3), the NH+4 pulse addition increased pHi (alkali jump) and its removal decreased pH(i) (acid jump) to the control level (no overacidification). This pHi change was induced by reaction 1 (NH3 + H+ <--> NH+4). However, in the presence of HCO(-3), the NH+4 pulse removal decreased pHi (acid jump) with overacidification. The extent of overacidification was decreased by acetazolamide (a carbonic anhydrase inhibitor), bumetanide (an inhibitor of Na+/K+/2Cl(-) cotransporter [NKCC]), and NPPB (an inhibitor of Cl(-) channel). The NH+4 pulse addition led to the accumulation of NH+4 in ATII cells via reaction 1 and NKCC, and the NH+4 pulse removal induced reaction 2 (NH+4 + HCO(-3) --> NH3 + H+ HCO(-3)) in addition to the reversal of reaction 1. Thus, NH+4 that entered via NKCC reacts with HCO(-3) (reaction 2) to produce H+, which induces overacidification in the acid jump. After the overacidification, the pH(i) recovery consisted of a rapid recovery (first phase) followed by a slow recovery (second phase). The first phase was inhibited by NPPB, glybenclamide, amiloride, and an Na+-free solution, and the second phase was inhibited by DIDS, MIA, and an Na+-free solution. Both phases were accelerated by a high extracellular HCO(-3) concentration. These observations indicate that the first phase was induced by HCO(-3) entry via Cl(-) channels coupled with Na+ channels activities, and that the second phase was induced by H+ extrusion via Na+/H+ exchanger and by HCO(-3) entry via HCO(-3) cotransporter. Thus, in ATII cells, HCO(-3) entry via Cl(-) channels is essential for recovering pHi after overacidification during the acid jump and for removing NH+4 that entered via NKCC from ATII cells, suggesting HCO(-3)-dependent NH3 excretion from lungs.