The association between diabetes insipidus (DI) and chronic dietary potassium deprivation is well known but it remains uncertain how the disorder develops and whether it is influenced by the sexual dimorphism in potassium handling. Here, we determine the plasma potassium (PK) threshold for DI in male and female mice and ascertain if the DI is initiated by polydipsia, or a central or nephrogenic defect. C57BL6J mice were randomized to a control diet or to graded reductions in dietary K+ for 8 days, and kidney function and transporters involved in water balance were characterized. We found male and female mice develop polyuria and secondary polydipsia. Altered water balance coincides with a decrease in AQP2 phosphorylation and apical localization despite increased levels of the vasopressin surrogate marker, copeptin. No change in the protein abundance of the urea transporter, UT-A1, was observed. NKCC2 decreased only in males. DDAVP treatment failed to reverse water diuresis in K+-restricted mice. These findings indicate that even small fall in PK is associated with nephrogenic DI (NDI), coincident with the development of altered AQP2 regulation, implicating low PK as a causal trigger of NDI. We found PK decreased more in females, and consequently females were more prone to develop NDI. Together these data indicate that AQP2 regulation is disrupted by a small decrease in PK and the response is influenced by sexual dimorphism in potassium handling. These findings provide new insights into the mechanisms linking water and potassium balances, and support defining the disorder as "Potassium-Dependent NDI."

Background: The urinary potassium excretion machinery is upregulated with increasing dietary potassium, but the role of accompanying dietary anions remains inadequately characterized. Poorly absorbable anions, including HCO3-, are thought to increase K+ secretion through a transepithelial voltage effect. Here we test if they also influence the potassium secretion machinery. Methods: Wild-type mice, aldosterone synthase knockout (AS-KO), or pendrin knockout mice were randomized to control, high KCl, or high KHCO3 diets. The potassium secretory capacity was assessed in balance studies. Protein abundance, modification, and localization of the potassium-secretory transporters were evaluated by western blot and confocal microscopy. Results: Feeding the high KHCO3 diet increased urinary K+ excretion and the trans-tubular K+ gradient significantly more than the high KCl diet, coincident with more pronounced upregulation of ENaC and ROMK and apical localization in the distal nephron. Studies in AS-KO mice revealed that the enhanced effects of bicarbonate were aldosterone-independent. The KHCO3 diet also uniquely increased the BK potassium channel b4 subunit, stabilizing BKa on the apical membrane, the Cl/HCO3 exchanger, pendrin, and the apical KCl cotransporter (KCC3a), all of which are expressed specifically in pendrin-positive IC cells. Studies in pendrin KO mice revealed that pendrin was required to increase K+excretion with the KHCO3 diet. In summary, bicarbonate stimulates potassium excretion beyond a poorly-absorbable anion effect, upregulating ENaC and ROMK in principal cells and BK, pendrin, and KCC3a in PP-ICs. The adaptive mechanism prevents hyperkalemia and alkalosis with the consumption of alkaline-ash-rich diets but may drive potassium wasting and hypokalemia in alkalosis.

We recently discovered that DCT1 cell‐specific expression of constitutively active SPAK (CA‐SPAK) drives a remodeling process of the entire distal nephron that includes atrophy of the aldosterone‐sensitive distal nephron and parallel inhibition of ROMK and ENaC (Grimm et al, JASN ’17). The mechanism by which genetic activation of NCC drives downstream nephron remodeling remains elusive but observations that PGE2 inhibits ROMK and ENaC ex vivo raises that paracrine signaling may be involved (Jin et al, AJPR 2007; Guan et al, JCI 1998). Here, we tested the hypothesis that activation of NCC causes atrophy of the CNT through increased PGE2 synthesis and release. We found increased levels of PGE2 in kidney cortical homogenates of CA‐SPAK mice compared to control mice on a control diet. Feeding wild‐type mice (WT) a low potassium diet (LKD) to physiologically activate NCC resulted in a comparable elevation of PGE2 and ASDN remodeling response, characterized by a decrease in ROMK/ENaC. Administration of the NCC‐blocking diuretic, hydrochlorothiazide, to CA‐SPAK mice restored PGE2 to control levels but had no effect on PGE2 of control or SPAK KO mice. Thus, PGE2 is elevated in response to super activation of NCC. PGE2 synthesis is controlled by specific prostaglandin E synthase isoforms. As revealed in western blot analysis, the microsomal prostaglandin E synthase‐1 (mPGES1) was selectively elevated in the kidney cortex of CA‐SPAK mice and WT mice on LKD compared to control mice, and this response was blocked by thiazide diuretics. Localization of mPGES1 by quantitative microscopy surprisingly revealed that increased protein abundance of mPGES1 was confined to the CNT and CCD of CA‐SPAK or the DCT2 and CNT of LKD treated mice. No significant changes in mPGES1 were observed in the DCT1. In conclusion, these studies identify PGE2 as a possible remodeling factor that is released from the late DCT and ASDN in response to NCC activation. We speculate that decreased sodium delivery to the CNT and CCD when NCC is activated drives the response; similar to how low NaCl drives PGE2 synthesis in the macula densa. The paracrine/autocrine pathway provides a novel means to communicate potassium sensing in the DCT to potassium secretion in the ASDN by driving tubule remodeling of specific segments of the distal tubule. Support or Funding Information NIH, NIDDK T32 and Fondation LeDucq

Aberrant activation of with-no-lysine kinase (WNK)-STE20/SPS1-related proline-alanine-rich protein kinase (SPAK) kinase signaling in the distal convoluted tubule (DCT) causes unbridled activation of the thiazide-sensitive sodium chloride cotransporter (NCC), leading to familial hyperkalemic hypertension (FHHt) in humans. Studies in FHHt mice engineered to constitutively activate SPAK specifically in the DCT (CA-SPAK mice) revealed maladaptive remodeling of the aldosterone sensitive distal nephron (ASDN), characterized by decrease in the potassium excretory channel, renal outer medullary potassium (ROMK), and epithelial sodium channel (ENaC), that contributes to the hyperkalemia. The mechanisms by which NCC activation in DCT promotes remodeling of connecting tubule (CNT) are unknown, but paracrine communication and reduced salt delivery to the ASDN have been suspected. Here, we explore the involvement of prostaglandin E2 (PGE2). We found that PGE2 and the terminal PGE2 synthase, mPGES1, are increased in kidney cortex of CA-SPAK mice, compared to control or SPAK KO mice. Hydrochlorothiazide (HCTZ) reduced PGE2 to control levels, indicating increased PGE2 synthesis is dependent on increased NCC activity. Immunolocalization studies revealed mPGES1 is selectively increased in the CNT of CA-SPAK mice, implicating low salt-delivery to ASDN as the trigger. Salt titration studies in an in vitro ASDN cell model, mouse CCD cell (mCCD-CL1), confirmed PGE2 synthesis is activated by low salt, and revealed that response is paralleled by induction of mPGES1 gene expression. Finally, inhibition of the PGE2 receptor, EP1, in CA-SPAK mice partially restored potassium homeostasis as it partially rescued ROMK protein abundance, but not ENaC. Together, these data indicate low sodium delivery to the ASDN activates PGE2 synthesis and this inhibits ROMK through autocrine activation of the EP1 receptor. These findings provide new insights into the mechanism by which activation of sodium transport in the DCT causes remodeling of the ASDN.

Hypokalemia can induce diabetes insipidus (DI), characterized by loss of free‐water reabsorption, loss of urine concentrating ability, polyuria and polydipsia. Although the disorder has been described for decades, the pathophysiological mechanisms remain poorly understood. In addition, it still unclear if there is a sexual dimorphism. In this study, we characterized hypokalemia‐induced DI over a spectrum of plasma potassium levels in mice fed a range of low potassium diets (LKD) between 0% and 0.11% K+, over 8 days compared to control diet (1%K+). We observed that 0%K+ and 0.026%K+ diets both induced hypokalemia, polyuria and a decrease in urine osmolality starting after 5 and 6 days, respectively. However, the 0.056%K diet induced only mild DI starting at day 7. Interestingly, the 0.11%K diet did not result in DI. We then assessed the DI in males and females mice fed with LKD (0%K+) or control diet for 8 days. After 3 days of LKD, both males and females exhibited decrease in urine osmolality which was more sever in females (−21%) than in males (−12%). Profound diuresis was observed at day 5 and it was more enhanced in females (5.2 +/−0.58 mL/day) than in males (3.0 +/−0.43 mL/day), which might be a consequence of the more severe hypokalemia in females compared to males (2.6 +/−0.29 versus 3.5 +/−0.08 mmol/L) after 8 days of LKD. Moreover, it has been shown that the urinary concentration defect occurs mainly in the inner medullary collecting duct (IMCD), involving a reduction in AQP2 channel expression. Recent studies localized AQP2 protein in the autophagosome in the IMCD of DI models suggesting the autophagic degradation of AQP2 under hypokalemia‐induced DI (Khositseth S., Sci rep 2015)(Kim WY., Sci rep 2019). We confirmed and extended this observation, finding that after 8 days of LKD, the ATG12‐Atg5‐Atg16 complex, the LC3II/LC3I ratio, and P62 were all increased in the medulla but no difference was detected by western blot in the cortex. However, we observed that AQP2 levels were more severely decreased in the cortex than in the medulla of the hypokalemic versus control mice suggesting that pathways other than autophagy are at play. Together, the data indicate: i) a tight relationship between loss of the renal concentrating ability and development of hypokalemia, ii) hypokalemia in response to dietary potassium deficiency is more sever in female mice than males, and as a consequence, females are more prone to develop DI than males, iii) the autophagy is induced in the medulla of the hypokalemic mice, iv) The hypokalemia‐induced decrease in AQP2 is correlated with induction of autophagy in the medulla but not in the cortex. Support or Funding Information NIH