Heterogeneity of H-K-ATPase-mediated acid secretion along the mouse collecting duct

1 Research Service, North Florida/South Georgia Veterans Health System, and ; 2 Division of Nephrology, Hypertension, and Transplantation, Department of Medicine, University of Florida, Gainesville, Florida Submitted August 11, 2009 ; accepted in final form November 17, 2009 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 HK 1 and HK 2 H-K-ATPase activity, and 3 ) the OMCD 1 cell line best exhibits H-K-ATPase. microperfusion; intracellular pH; acid-base balance; intercalated cell; 2',7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein Address for reprint requests and other correspondence: C. S. Wingo, Research Service, North Florida/South Georgia Veterans Health System, 1601 SW Archer Rd., Gainesville, FL 32608 (e-mail: cswingo{at}ufl.edu ).