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Role of Kir5.1 (Kcnj16) Channel in the Control of Potassium Homeostasis in the Kidney

The inwardly‐rectifying potassium (K+) channels (Kir), Kir4.1/Kir5.1 (Kcnj10/Kcnj16) are critical channels responsible for the recycling of K+ in the distal nephron as well as regulation of basolateral membrane potential. Human mutations in KCNJ10 and KCNJ16 are associated with renal pathologies characterized by salt‐wasting, renal tubulopathy and hypokalemia (EAST/SeSAME and Gitelman syndromes) as well as cardiac phenotypes such as Brugada syndrome). The Kir4.1/Kir5.1 heteromeric channel is expressed on the basolateral side of distal convoluted tubule (DCT) cells and the principal cells of the cortical collecting duct (CCDs). Immunohistochemistry analysis of Dahl salt‐sensitive (SS) rats showed increased Kir5.1 abundance in the CCD compared to the DCT. Moreover, expression of Kir5.1 in both segments was significantly upregulated after 3 weeks of high salt diet (4% NaCl), showing a direct correlation between the development of salt sensitive hypertension and basolateral K+ channel function. To further explore the role of this channel in salt‐sensitive hypertension, we have created a loss‐of‐function mutation in Kir5.1 in the Dahl SS rat (SS‐Kcnj16em1Mcwi) using zinc‐finger nucleases. Overall, Kir5.1 loss‐of‐function mutation in the SS rat induced a renal phenotype characterized by hypokalemia (2.1±0.2 vs 4.3±0.1 mmol/L (mutant vs SS)), low aldosterone level (245.4±26.5 vs 1050±100 pg/ml), salt wasting (FEK+ 30.3±3.1 vs 5.8±0.6; FENa+ 0.42±0.06 vs 0.19±0.03; FEMg2+ 15.8±1.3 vs 6.1±0.9 %) and decreased mean arterial blood pressure (91.3±1.8 vs 104.7±5.5 mmHg). Furthermore, the absence of the Kir5.1 from distal nephron resulted in a compensatory increase of Kir4.1 protein abundance. IHC analysis of Kir4.1 localization revealed a rerouting from its expected location in the basolateral membrane of CCD and DCT tubules to the cytosol instead. We also found that loss‐of‐function mutation of Kir5.1 triggered upregulation of both total and phosphorylated forms of the NCC and NKCC2. In contrast, we were not able to identify any changes in the expression of ENaC subunits. Increasing the salt content of the diet from 0.4 to 4% triggered sodium and potassium excretion in Kir5.1 mutants, which resulted in a rapid reduction of blood K+ to critically low values (1.36±0.04 vs 2.1±0.1mmo/L (HS vs LS)). Na+ blood concentrations (141.6±0.5 vs 139.2±1.5 mmo/L) were unchanged between these two groups. This severe hypokalemia resulted in high mortality rate with most animals dying within 24 hours. These data demonstrate the critical role of renal basolateral Kir4.1/Kir5.1 channels in potassium handling, and identify them as a potential pharmacological target for the treatment of the imbalanced potassium homeostasis.