The distal convoluted tubule serves as a potassium switch in an intracellular chloride-dependent mechanism

Background       The kidney plays a key role in the regulation of K⁺ excretion and K⁺ homeostasis.  Previous studies have demonstrated that the thiazide-sensitive Na-Cl cotransporter (NCC) in the distal convoluted tubule (DCT) plays an important role in the regulation of renal K⁺ excretion by controlling sodium and volume delivery to the distal nephron.  It has been shown that NCC is inhibited when K⁺ intake is high and activated when dietary K⁺ intake is low. It is now generally accepted that WNKs (with-no-lysine kinases) plays a major role in NCC phosphorylation and activation. A large body of evidence has demonstrated that WNK1 and WNK4 are chloride-sensitive kinases, which are inhibited by high intracellular chloride concentration ([Cl^-]_i) and activated by low [Cl^-]_i. It has also been suggested that membrane voltage changes by extracellular K⁺ concentrations are responsible for altering the [Cl^-]_i thereby affecting WNK activity. The aim of our study is to test the hypothesis that changes in the basolateral cell voltage alter the [Cl^-]_i in the DCT.

Methods             To determine the [Cl^-]_i in the DCT, we have used isolated single DCT tubule of transgenic mice expressing Cl-sensor, a chloride-sensitive fluorescent protein modified from Chlomeleon. The Cl-sensor includes both yellow and cyan fluorescent moieties, allowing ratiometric estimation of [Cl^-]_i.  

Results                We first measured the [Cl^-]_i in the isolated DCT by establishing the calibration curve. Using this calibration curve, we estimated that basal DCT [Cl^-]_i is 7 ± 2.3 mM (n=18) . Changing the extracellular potassium concentration from 10 mM to 2 mM decreases the [Cl^-]_i. To test the role of the basolateral Kir4.1/Kir5.1 of the DCT in altering [Cl^-]_i, we examined whether inhibition of the basolateral K⁺ channels with Ba²⁺ would increase the [Cl^-]_i since Ba²⁺ has been shown to depolarize DCT membrane.  Indeed, Inhibition of Kir4.1/Kir5.1 significantly increased the [Cl^-]_i in the DCT bathed in a solution containing 2 mM K⁺.

Conclusion         Our results indicate that the intracellular Cl^- concentrations of DCT cells are low at baseline and that the depolarization increases whereas hyperpolarization decreases the intracellular Cl^- concentrations. Thus, changes in DCT voltage are associated with alteration of intracellular Cl^- concentrations.