Ouabain-induced cell swelling in rabbit cortical collecting tubule: NaCl transport by principal cells

Abstract

Ouabain had no effect on the volume of intercalated cells of DOCA-stimulated rabbit cortical collecting tubules, but caused principal cells to swell rapidly at an initial rate of 67%/min. Principal cells swelled 133% then activated regulatory volume decrease mechanisms and shrank at an initial rate of -3%/min to a new volume 13% above control. The initial rate of ouabain swelling was completely inhibited by perfusate Na+ removal or reduced 95% by luminal addition of 10(-5) M amiloride. Luminal, peritubular, or bilateral Cl- removal each caused cell shrinkages of 10% and reduced the rate of ouabain swelling by 70, 85, and 99%, respectively. The presence of an apical Cl- transport step in principal cells was confirmed by increasing luminal K+ from 5 to 53 mM, which caused cell swelling of 22%. This volume increase was completely blocked by luminal Cl- removal, but was unaffected by peritubular Cl- substitution. Perfusion of CCT with 0.1 mM acetazolomide, 0.1 mM DPC or 0.5 mM SITS caused principal cell shrinkages of 7-9% and reduced the rate of ouabain swelling by 60, 70, and 40%, respectively. The initial rate of ouabain swelling was inhibited 70% by bilateral CO2/HCO3 removal and 50% by whole animal acid loading. Taken together these results demonstrate that ouabain swelling is due to cellular NaCl accumulation and that Na+ enters the cell primarily through apical Na+ channels. Cellular Cl- entry occurs at least partially through the apical membrane and may be mediated by a Cl-/HCO3- exchanger. Brief (45-90 sec) exposure of principal cells to ouabain is associated with a rapid inhibition of Na+ and/or Cl- entry steps, whereas long-term (greater than 5 min) ouabain exposure completely blocks one or both of these transport pathways.