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 53mm, 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 0.1mm acetazolomide, 0.1mm DPC or 0.5mm 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 demounstrate 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⁻/HCO ₃ ⁻ 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 (>5min) ouabvain exposure completely blocks one or both of these transport pathways.