Transcellular chloride pathways in ambystoma proximal tubule

The transport mechanisms of Ambystoma proximal tubule that mediate transcellular Cl- absorption linked to Na+ were investigated in isolated perfused tubules using Cl--selective and voltage-recording microelectrodes. In control solutions intracellular activity of Cl- (aiCl) is 11.3 +/- 0.5 mm, the basolateral (V1), apical (V2), and transepithelial (V3) potential differences are -68 +/- 1.2 mV, +62 +/- 1.2 mV and -6.4 +/- 0.3 mV, respectively. When Na+ absorption is decreased by removal of organic substrates from the lumen, aiCl falls by 1.3 +/- 0.3 mm and V2 hyperpolarizes by +11.4 +/- 1.7 mV. Subsequent removal of Na+ from the lumen causes aiCl to fall further by 2.3 +/- 0.4 mm and V2 to hyperpolarize further by +15.3 +/- 2.4 mV. The contribution of transporters and channels to the observed changes of aiCl was examined using ion substitutions and inhibitors. Apical Na/Cl or Na/K/2Cl symport is excluded because bumetanide, furosemide or hydrochlorothiazide have no effect on aiCl. The effects of luminal HCO-3 removal and/or of disulfonic stilbenes argue against the presence of apical Cl-base exchange such as Cl-HCO3 or Cl-OH. The effects of basolateral HCO-3 removal, of basolateral Na+ removal and/or of disulfonic stilbenes are compatible with presence of basolateral Na-independent Cl-base exchange and Na-driven Cl-HCO3 exchange. Several lines of evidence favor conductive Cl- transport across both the apical and basolateral membrane. Addition of the chloride-channel blocker diphenylamine-2-carboxylate to the lumen or bath, increases the aiCl by 2.4 +/- 0.6 mm or 2.9 +/- 1.0 mm respectively. Moreover, following inhibition by DIDS of all anion exchangers in HCO-3-free Ringer, the equilibrium potential for Cl- does not differ from the membrane potential V2. Finally, the logarithmic changes in aiCl in various experimental conditions correlate well with the simultaneous changes in either basolateral or apical membrane potential. These findings strongly support the presence of Cl- channels at the apical and basolateral cell membranes of the proximal tubule.