In this study, K+ channels present in the basolateral membrane of the distal convoluted tubule (DCT) were investigated using patch‐clamp methods. In addition, Kir4.1, Kir4.2 and Kir5.1 inward rectifier channels were investigated using RT‐PCR and immunohistochemistry (Kir4.1). DCTs were microdissected from collagenase‐treated mouse kidneys. One type of K+ channel was detected in about 50 % of cell‐attached patches from the DCT basolateral membrane; this channel was inwardly rectifying and had an inward conductance (gin) of ∼40 pS at an external [K+] of 145 mm. The current‐voltage relationship was linear when inside‐out patches were exposed to a Mg2+‐free medium. Mg2+ at a concentration of 1.2 mm considerably reduced the outward conductance (gout), yielding a gin/gout ratio of ∼4.7. The polycation spermine (5 × 10−7m) reduced the open probability (Po) by 50 %. Channel activity was dependent upon the intracellular pH, with acid pH decreasing, and basic pH increasing, Po. Internal ATP (2 mm) and Ca2+ (up to 10−3m) had no effect. Channel activity declined irreversibly when the inner side of the patch was exposed to Mg2+. Kir4.1, Kir4.2 and Kir5.1 mRNAs were all detected in the DCT. The Kir4.1 protein co‐localised with the Na+‐Cl− cotransporter, which is specific to the DCT, and was located on basolateral membranes. The DCT K+ channel differs from other functionally identified renal K+ channels with regard to its inhibition by spermine and insensitivity to internal ATP and Ca2+. At the current state of knowledge, the channel is similar to Kir4.1‐Kir5.1 and Kir4.2‐Kir5.1 heteromeric channels, but not to Kir4.1 or Kir4.2 homomeric channels.

The heteromeric inwardly rectifying Kir4.1/Kir5.1 K + channel underlies the basolateral K + conductance in the distal nephron and is extremely sensitive to inhibition by intracellular pH. The functional importance of Kir4.1/Kir5.1 in renal ion transport has recently been highlighted by mutations in the human Kir4.1 gene ( KCNJ10 ) that result in seizures, sensorineural deafness, ataxia, mental retardation, and electrolyte imbalance (SeSAME)/epilepsy, ataxia, sensorineural deafness, and renal tubulopathy (EAST) syndrome, a complex disorder that includes salt wasting and hypokalemic alkalosis. Here, we investigated the role of the Kir5.1 subunit in mice with a targeted disruption of the Kir5.1 gene ( Kcnj16 ). The Kir5.1 −/− mice displayed hypokalemic, hyperchloremic metabolic acidosis with hypercalciuria. The short-term responses to hydrochlorothiazide, an inhibitor of ion transport in the distal convoluted tubule (DCT), were also exaggerated, indicating excessive renal Na + absorption in this segment. Furthermore, chronic treatment with hydrochlorothiazide normalized urinary excretion of Na + and Ca 2+ , and abolished acidosis in Kir5.1 −/− mice. Finally, in contrast to WT mice, electrophysiological recording of K + channels in the DCT basolateral membrane of Kir5.1 −/− mice revealed that, even though Kir5.1 is absent, there is an increased K + conductance caused by the decreased pH sensitivity of the remaining homomeric Kir4.1 channels. In conclusion, disruption of Kcnj16 induces a severe renal phenotype that, apart from hypokalemia, is the opposite of the phenotype seen in SeSAME/EAST syndrome. These results highlight the important role that Kir5.1 plays as a pH-sensitive regulator of salt transport in the DCT, and the implication of these results for the correct genetic diagnosis of renal tubulopathies is discussed.