Abstract
In the collecting duct (CD), H-K-ATPases function in cation reabsorption and H secretion. This study evaluated H-K-ATPase-mediated H secretion along the mouse CD, measured as EIPA- and luminal bafilomycin A(1)-insensitive intracellular pH (pH(i)) recovery from acute H loading (NH(4)) using BCECF. pH(i) recovery was measured in 1) microperfused cortical, outer medullary, and inner medullary CDs (CCD, OMCD, and IMCD) from C57BL/6J mice fed a normal diet and 2) common murine CD cell lines. H-K-ATPase activity along the native, microperfused CD was greatest in the CCD, less in the OMCD, and least in the IMCD (0.10 +/- 0.02, 0.04 +/- 0.01, and 0.01 +/- 0.002 U/min, respectively). H-K-ATPase activity was 0.30 +/- 0.03 and 0.26 +/- 0.03 in A- and B-type ICs, respectively, and was sensitive to Sch-28080 or ouabain. pH(i) recovery was greatest in the OMCD(1) cell line (0.25 +/- 0.01) and less in mpkCCD(c14) (0.17 +/- 0.01), mIMCD-K2 (0.12 +/- 0.01), and mIMCD-3 (0.05 +/- 0.01) cells. EIPA inhibited the majority of pH(i) recovery in these cells (100%, 64%, 75%, and 80% in mpkCCD(c14), OMCD(1), mIMCD-K2, and mIMCD-3, respectively). In OMCD(1) cells, where EIPA-insensitive pH(i) recovery was greatest, H-K-ATPase activity was 0.10 +/- 0.01 and was significantly inhibited (80%) by Sch-28080. We conclude that 1) H-K-ATPase-mediated H secretion in the native mouse CD is greatest in the ICs of the CCD, 2) A- and B-type ICs possess HKalpha(1) and HKalpha(2) H-K-ATPase activity, and 3) the OMCD(1) cell line best exhibits H-K-ATPase.