GDF15, an Emerging Player in Renal Physiology and Pathophysiology.

These last years, the growth factor GDF15 has emerged as a key element in many different biological processes. It has been established as being produced in response to many pathological states and is now referred to as a stress-related hormone. Regarding kidney functions, GDF15 has been involved in different pathologies such as chronic kidney disease, diabetic nephropathy, renal cancer, and so on. Interestingly, recent studies also revealed a role of GDF15 in the renal homeostatic mechanisms allowing to maintain constant, as far as possible, the plasma parameters such as pH and K+ values. In this review, we recapitulate the role of GDF15 in physiological and pathological context by focusing our interest on its renal effect.

Putting bicarbonate on the spot: pharmacological insights for CFTR correction in the airway epithelium.

Introduction: Cystic fibrosis (CF) is caused by defective Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) proteins. CFTR controls chloride (Cl-) and bicarbonate (HCO3 -) transport into the Airway Surface Liquid (ASL). We investigated the impact of F508del-CFTR correction on HCO3 - secretion by studying transepithelial HCO3 - fluxes. Methods: HCO3 - secretion was measured by pH-stat technique in primary human respiratory epithelial cells from healthy subjects (WT) and people with CF (pwCF) carrying at least one F508del variant. Its changes after CFTR modulation by the triple combination VX445/661/770 and in the context of TNF-α+IL-17 induced inflammation were correlated to ASL pH and transcriptional levels of CFTR and other HCO3 - transporters of airway epithelia such as SLC26A4 (Pendrin), SLC26A9 and NBCe1. Results: CFTR-mediated HCO3 - secretion was not detected in F508del primary human respiratory epithelial cells. It was rescued up to ∼ 80% of the WT level by VX-445/661/770. In contrast, TNF-α+IL-17 normalized transepithelial HCO3 - transport and increased ASL pH. This was related to an increase in SLC26A4 and CFTR transcript levels. VX-445/661/770 induced an increase in pH only in the context of inflammation. Effects on HCO3 - transport were not different between F508del homozygous and F508del compound heterozygous CF airway epithelia. Conclusion: Our studies show that correction of F508del-CFTR HCO3 - is not sufficient to buffer acidic ASL and inflammation is a key regulator of HCO3 - secretion in CF airways. Prediction of the response to CFTR modulators by theratyping should take into account airway inflammation.

GDF15 mediates renal cell plasticity in response to potassium depletion in mice.

OBJECTIVE: To understand the mechanisms involved in the response to a low-K+ diet (LK), we investigated the role of the growth factor GDF15 and the ion pump H,K-ATPase type 2 (HKA2) in this process. METHODS: Male mice of different genotypes (WT, GDF15-KO, and HKA2-KO) were fed an LK diet for different periods of time. We analyzed GDF15 levels, metabolic and physiological parameters, and the cellular composition of collecting ducts. RESULTS: Mice fed an LK diet showed a 2-4-fold increase in plasma and urine GDF15 levels. Compared to WT mice, GDF15-KO mice rapidly developed hypokalemia due to impaired renal adaptation. This is related to their 1/ inability to increase the number of type A intercalated cells (AIC) and 2/ absence of upregulation of H,K-ATPase type 2 (HKA2), the two processes responsible for K+ retention. Interestingly, we showed that the GDF15-mediated proliferative effect on AIC was dependent on the ErbB2 receptor and required the presence of HKA2. Finally, renal leakage of K+ induced a reduction in muscle mass in GDF15-KO mice fed LK diet. CONCLUSIONS: In this study, we showed that GDF15 and HKA2 are linked and play a central role in the response to K+ restriction by orchestrating the modification of the cellular composition of the collecting duct.

Airway surface hyperviscosity and defective mucociliary transport by IL-17/TNF-α are corrected by ß-adrenergic stimulus.

The fluid covering the surface of airway epithelia represents a first barrier against pathogens. The chemical and physical properties of the airway surface fluid are controlled by the activity of ion channels and transporters. In cystic fibrosis (CF), loss of CFTR chloride channel function causes airway surface dehydration, bacterial infection, and inflammation. We investigated the effects of IL-17A plus TNF-α, 2 cytokines with relevant roles in CF and other chronic lung diseases. Transcriptome analysis revealed a profound change with upregulation of several genes involved in ion transport, antibacterial defense, and neutrophil recruitment. At the functional level, bronchial epithelia treated in vitro with the cytokine combination showed upregulation of ENaC channel, ATP12A proton pump, ADRB2 ß-adrenergic receptor, and SLC26A4 anion exchanger. The overall result of IL-17A/TNF-α treatment was hyperviscosity of the airway surface, as demonstrated by fluorescence recovery after photobleaching (FRAP) experiments. Importantly, stimulation with a ß-adrenergic agonist switched airway surface to a low-viscosity state in non-CF but not in CF epithelia. Our study suggests that CF lung disease is sustained by a vicious cycle in which epithelia cannot exit from the hyperviscous state, thus perpetuating the proinflammatory airway surface condition.

Acidosis-induced activation of distal nephron principal cells triggers Gdf15 secretion and adaptive proliferation of intercalated cells.

AIM: Type A intercalated cells of the renal collecting duct participate in the maintenance of the acid/base balance through their capacity to adapt proton secretion to homeostatic requirements. We previously showed that increased proton secretion stems in part from the enlargement of the population of proton secreting cells in the outer medullary collecting duct through division of fully differentiated cells, and that this response is triggered by growth/differentiation factor 15. This study aimed at deciphering the mechanism of acid load-induced secretion of Gdf15 and its mechanism of action. METHODS: We developed an original method to evaluate the proliferation of intercalated cells and applied it to genetically modified or pharmacologically treated mice under basal and acid-loaded conditions. RESULTS: Gdf15 is secreted by principal cells of the collecting duct in response to the stimulation of vasopressin receptors. Vasopressin-induced production of cAMP triggers activation of AMP-stimulated kinases and of Na,K-ATPase, and induction of p53 and Gdf15. Gdf15 action on intercalated cells is mediated by ErbB2 receptors, the activation of which triggers the expression of cyclin d1, of p53 and anti-proliferative genes, and of Egr1. CONCLUSION: Acidosis-induced proliferation of intercalated cells results from a cross talk with principal cells which secrete Gdf15 in response to their stimulation by vasopressin. Thus, vasopressin is a major determinant of the collecting duct cellular homeostasis as it promotes proliferation of intercalated cells under acidosis conditions and of principal cells under normal acid-base status.

Increased colonic K+ excretion through inhibition of the H,K-ATPase type 2 helps reduce plasma K+ level in a murine model of nephronic reduction.

Hyperkalemia is frequently observed in patients at the end-stage of chronic kidney disease (CKD), and has possible harmful consequences on cardiac function. Many strategies are currently used to manage hyperkalemia, one consisting of increasing fecal K+ excretion through the administration of cation-exchange resins. In this study, we explored another more specific method of increasing intestinal K+ secretion by inhibiting the H,K-ATPase type 2 (HKA2), which is the main colonic K+ reabsorptive pathway. We hypothetised that the absence of this pump could impede the increase of plasma K+ levels following nephronic reduction (N5/6) by favoring fecal K+ secretion. In N5/6 WT and HKA2KO mice under normal K+ intake, the plasma K+ level remained within the normal range, however, a load of K+ induced strong hyperkalemia in N5/6 WT mice (9.1 ± 0.5 mM), which was significantly less pronounced in N5/6 HKA2KO mice (7.9 ± 0.4 mM, p < 0.01). This was correlated to a higher capacity of HKA2KO mice to excrete K+ in their feces. The absence of HKA2 also increased fecal Na+ excretion by inhibiting its colonic ENaC-dependent absorption. We also showed that angiotensin-converting-enzyme inhibitor like enalapril, used to treat hypertension during CKD, induced a less severe hyperkalemia in N5/6 HKA2KO than in N5/6 WT mice. This study therefore provides the proof of concept that the targeted inhibition of HKA2 could be a specific therapeutic maneuver to reduce plasma K+ levels in CKD patients.

H,K-ATPase type 2 regulates gestational extracellular compartment expansion and blood pressure in mice.

The modifications of the hemodynamic system and hydromineral metabolism are physiological features characterizing a normal gestation. Thus, the ability to expand plasma volume without increasing the level of blood pressure is necessary for the correct perfusion of the placenta. The kidney is essential in this adaptation by reabsorbing avidly sodium and fluid. In this study, we observed that the H,K-ATPase type 2 (HKA2), an ion pump expressed in kidney and colon and already involved in the control of the K+ balance during gestation, is also required for the correct plasma volume expansion and to maintain normal blood pressure. Indeed, compared with WT pregnant mice that exhibit a 1.6-fold increase of their plasma volume, pregnant HKA2-null mice (HKA2KO) only modestly expand their extracellular volume (×1.2). The renal expression of the epithelial Na channel (ENaC) α- and γ-subunits and that of the pendrin are stimulated in gravid WT mice, whereas the Na/Cl- cotransporter (NCC) expression is downregulated. These modifications are all blunted in HKA2KO mice. This impeded renal adaptation to gestation is accompanied by the development of hypotension in the pregnant HKA2KO mice. Altogether, our results showed that the absence of the HKA2 during gestation leads to an "underfilled" situation and has established this transporter as a key player of the renal control of salt and potassium metabolism during gestation.

Adrenal adaptation in potassium-depleted men: role of progesterone?

BACKGROUND: In rodents, the stimulation of adrenal progesterone is necessary for renal adaptation under potassium depletion. Here, we sought to determine the role of progesterone in adrenal adaptation in potassium-depleted healthy human volunteers and compared our findings with data collected in patients with Gitelman syndrome (GS), a salt-losing tubulopathy. METHODS: Twelve healthy young men were given a potassium-depleted diet for 7 days at a tertiary referral medical centre (NCT02297048). We measured by liquid chromatography coupled to tandem mass spectroscopy plasma steroid concentrations at Days 0 and 7 before and 30 min after treatment with tetracosactide. We compared these data with data collected in 10 GS patients submitted to tetracosactide test. RESULTS: The potassium-depleted diet decreased plasma potassium in healthy subjects by 0.3 ± 0.1 mmol/L, decreased plasma aldosterone concentration by 50% (P = 0.0332) and increased plasma 17-hydroxypregnenolone concentration by 45% (P = 0.0232) without affecting other steroids. CYP17 activity, as assessed by 17-hydroxypregnenolone/pregnenolone ratio, increased by 60% (P = 0.0389). As compared with healthy subjects, GS patients had 3-fold higher plasma concentrations of aldosterone, 11-deoxycortisol (+30%) and delta 4-androstenedione (+14%). Their post-tetracosactide progesterone concentration was 2-fold higher than that of healthy subjects and better correlated to plasma potassium than to plasma renin. CONCLUSION: The increase in 17-hydroxypregnenolone concentration after mild potassium depletion in otherwise healthy human subjects suggests that 17 hydroxylation of pregnenolone prevents the increase in progesterone observed in potassium-depleted mice. The unexpected over-response of non-mineralocorticoid steroids to tetracosactide in GS subjects suggests that the adrenal system not only adapts to sodium depletion but may also respond to hypokalaemia.

Author Correction: Airway surface liquid acidification initiates host defense abnormalities in Cystic Fibrosis.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

ANP-stimulated Na+ secretion in the collecting duct prevents Na+ retention in the renal adaptation to acid load.

We have recently reported that type A intercalated cells of the collecting duct secrete Na+ by a mechanism coupling the basolateral type 1 Na+-K+-2Cl- cotransporter with apical type 2 H+-K+-ATPase (HKA2) functioning under its Na+/K+ exchange mode. The first aim of the present study was to evaluate whether this secretory pathway is a target of atrial natriuretic peptide (ANP). Despite hyperaldosteronemia, metabolic acidosis is not associated with Na+ retention. The second aim of the present study was to evaluate whether ANP-induced stimulation of Na+ secretion by type A intercalated cells might account for mineralocorticoid escape during metabolic acidosis. In Xenopus oocytes expressing HKA2, cGMP, the second messenger of ANP, increased the membrane expression, activity, and Na+-transporting rate of HKA2. Feeding mice with a NH4Cl-enriched diet increased urinary excretion of aldosterone and induced a transient Na+ retention that reversed within 3 days. At that time, expression of ANP mRNA in the collecting duct and urinary excretion of cGMP were increased. Reversion of Na+ retention was prevented by treatment with an inhibitor of ANP receptors and was absent in HKA2-null mice. In conclusion, paracrine stimulation of HKA2 by ANP is responsible for the escape of the Na+-retaining effect of aldosterone during metabolic acidosis.

Airway surface liquid acidification initiates host defense abnormalities in Cystic Fibrosis.

Cystic fibrosis (CF) is caused by defective Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) protein. Morbidity is mainly due to early airway infection. We hypothesized that S. aureus clearance during the first hours of infection was impaired in CF human Airway Surface Liquid (ASL) because of a lowered pH. The ASL pH of human bronchial epithelial cell lines and primary respiratory cells from healthy controls (WT) and patients with CF was measured with a pH microelectrode. The antimicrobial capacity of airway cells was studied after S. aureus apical infection by counting surviving bacteria. ASL was significantly more acidic in CF than in WT respiratory cells. This was consistent with a defect in bicarbonate secretion involving CFTR and SLC26A4 (pendrin) and a persistent proton secretion by ATP12A. ASL demonstrated a defect in S. aureus clearance which was improved by pH normalization. Pendrin inhibition in WT airways recapitulated the CF airway defect and increased S. aureus proliferation. ATP12A inhibition by ouabain decreased bacterial proliferation. Antimicrobial peptides LL-37 and hBD1 demonstrated a pH-dependent activity. Normalizing ASL pH might improve innate airway defense in newborns with CF during onset of S. aureus infection. Pendrin activation and ATP12A inhibition could represent novel therapeutic strategies to normalize pH in CF airways.

Increased expression of ATP12A proton pump in cystic fibrosis airways.

Proton secretion mediated by ATP12A protein on the surface of the airway epithelium may contribute to cystic fibrosis (CF) lung disease by favoring bacterial infection and airway obstruction. We studied ATP12A in fresh bronchial samples and in cultured epithelial cells. In vivo, ATP12A expression was found almost exclusively at the apical side of nonciliated cells of airway epithelium and in submucosal glands, with much higher expression in CF samples. This could be due to bacterial infection and inflammation, since treating cultured cells with bacterial supernatants or with IL-4 (a cytokine that induces goblet cell hyperplasia) increased the expression of ATP12A in nonciliated cells. This observation was associated with upregulation and translocation of ATP1B1 protein from the basal to apical epithelial side, where it colocalizes with ATP12A. ATP12A function was evaluated by measuring the pH of the apical fluid in cultured epithelia. Under resting conditions, CF epithelia showed more acidic values. This abnormality was minimized by inhibiting ATP12A with ouabain. Following treatment with IL-4, ATP12A function was markedly increased, as indicated by strong acidification occurring under bicarbonate-free conditions. Our study reveals potentially novel aspects of ATP12A and remarks its importance as a possible therapeutic target in CF and other respiratory diseases.

Glucagon revisited: Coordinated actions on the liver and kidney.

Glucagon secretion is stimulated by a low plasma glucose concentration. By activating glycogenolysis and gluconeogenesis in the liver, glucagon contributes to maintain a normal glycemia. Glucagon secretion is also stimulated by the intake of proteins, and glucagon contributes to amino acid metabolism and nitrogen excretion. Amino acids are used for gluconeogenesis and ureagenesis, two metabolic pathways that are closely associated. Intriguingly, cyclic AMP, the second messenger of glucagon action in the liver, is released into the bloodstream becoming an extracellular messenger. These effects depend not only on glucagon itself but on the actual glucagon/insulin ratio because insulin counteracts glucagon action on the liver. This review revisits the role of glucagon in nitrogen metabolism and in disposal of nitrogen wastes. This role involves coordinated actions of glucagon on the liver and kidney. Glucagon influences the transport of fluid and solutes in the distal tubule and collecting duct, and extracellular cAMP influences proximal tubule reabsorption. These combined effects increase the fractional excretion of urea, sodium, potassium and phosphates. Moreover, the simultaneous actions of glucagon and extracellular cAMP are responsible, at least in part, for the protein-induced rise in glomerular filtration rate that contributes to a more efficient excretion of protein-derived end products.

Versatility of NaCl transport mechanisms in the cortical collecting duct.

The cortical collecting duct (CCD) forms part of the aldosterone-sensitive distal nephron and plays an essential role in maintaining the NaCl balance and acid-base status. The CCD epithelium comprises principal cells as well as different types of intercalated cells. Until recently, transcellular Na+ transport was thought to be restricted to principal cells, whereas (acid-secreting) type A and (bicarbonate-secreting) type B intercalated cells were associated with the regulation of acid-base homeostasis. This review describes how this traditional view has been upended by several discoveries in the past decade. A series of studies has shown that type B intercalated cells can mediate electroneutral NaCl reabsorption by a mechanism involving Na+-dependent and Na+-independent Cl-/[Formula: see text] exchange, and that is energetically driven by basolateral vacuolar H+-ATPase pumps. Other research indicates that type A intercalated cells can mediate NaCl secretion, through a bumetanide-sensitive pathway that is energized by apical H+,K+-ATPase type 2 pumps operating as Na+/K+ exchangers. We also review recent findings on the contribution of the paracellular route to NaCl transport in the CCD. Last, we describe cross-talk processes, by which one CCD cell type impacts Na+/Cl- transport in another cell type. The mechanisms that have been identified to date demonstrate clearly the interdependence of NaCl and acid-base transport systems in the CCD. They also highlight the remarkable versatility of this nephron segment.

H,K-ATPase type 2 contributes to salt-sensitive hypertension induced by K(+) restriction.

In industrialized countries, a large part of the population is daily exposed to low K(+) intake, a situation correlated with the development of salt-sensitive hypertension. Among many processes, adaptation to K(+)-restriction involves the stimulation of H,K-ATPase type 2 (HKA2) in the kidney and colon and, in this study, we have investigated whether HKA2 also contributes to the determination of blood pressure (BP). By using wild-type (WT) and HKA2-null mice (HKA2 KO), we showed that after 4 days of K(+) restriction, WT remain normokalemic and normotensive (112 ± 3 mmHg) whereas HKA2 KO mice exhibit hypokalemia and hypotension (104 ± 2 mmHg). The decrease of BP in HKA2 KO is due to the absence of NaCl-cotransporter (NCC) stimulation, leading to renal loss of salt and decreased extracellular volume (by 20 %). These effects are likely related to the renal resistance to vasopressin observed in HKA2 KO that may be explained, in part by the increased production of prostaglandin E2 (PGE2). In WT, the stimulation of NCC induced by K(+)-restriction is responsible for the elevation in BP when salt intake increases, an effect blunted in HKA2-null mice. The presence of an activated HKA2 is therefore required to limit the decrease in plasma [K(+)] but also contributes to the development of salt-sensitive hypertension.

The renal cortical collecting duct: a secreting epithelium?

KEY POINTS: The cortical collecting duct (CCD) plays an essential role in sodium homeostasis by fine-tuning the amount of sodium that is excreted in the urine. Ex vivo, the microperfused CCD reabsorbs sodium in the absence of lumen-to-bath concentration gradients. In the present study, we show that, in the presence of physiological lumen-to-bath concentration gradients, and in the absence of endocrine, paracrine and neural regulation, the mouse CCD secretes sodium, which represents a paradigm shift. This secretion occurs via the paracellular route, as well as a transcellular pathway that is energized by apical H+ /K+ -ATPase type 2 pumps operating as Na+ /K+ exchangers. The newly identified transcellular secretory pathway represents a physiological target for the regulation of sodium handling and for anti-hypertensive therapeutic agents. ABSTRACT: In vitro microperfusion experiments have demonstrated that cortical collecting ducts (CCDs) reabsorb sodium via principal and type B intercalated cells under sodium-depleted conditions and thereby contribute to sodium and blood pressure homeostasis. However, these experiments were performed in the absence of the transepithelial ion concentration gradients that prevail in vivo and determine paracellular transport. The present study aimed to characterize Na+ , K+ and Cl- fluxes in the mouse CCD in the presence of physiological transepithelial concentration gradients. For this purpose, we combined in vitro measurements of ion fluxes across microperfused CCDs of sodium-depleted mice with the predictions of a mathematical model. When NaCl transport was inhibited in all cells, CCDs secreted Na+ and reabsorbed K+ ; Cl- transport was negligible. Removing inhibitors of type A and B intercalated cells increased Na+ secretion in wild-type (WT) mice but not in H+ /K+ -ATPase type 2 (HKA2) knockout mice. Further inhibition of basolateral NaCl entry via the Na+ -K+ -2Cl- cotransporter in type A intercalated cells reduced Na+ secretion in WT mice to the levels observed in HKA2-/- mice. With no inhibitors, WT mouse CCDs still secreted Na+ and reabsorbed K+ . In vivo, HKA2-/- mice excreted less Na+ than WT mice after switching to a high-salt diet. Taken together, our results indicate that type A intercalated cells secrete Na+ via basolateral Na+ -K+ -2Cl- cotransporters in tandem with apical HKA2 pumps. They also suggest that the CCD can mediate overall Na+ secretion, and that its ability to reabsorb NaCl in vivo depends on the presence of acute regulatory factors.

H-K-ATPase type 2: relevance for renal physiology and beyond.

H-K-ATPase type 2 (HKA2), also known as the "nongastric" or "colonic" H-K-ATPase, is broadly expressed, and its presence in the kidney has puzzled experts in the field of renal ion transport systems for many years. One of the most important and robust characteristics of this transporter is that it is strongly stimulated after dietary K(+) restriction. This result prompted many investigators to propose that it should play a role in allowing the kidney to efficiently retain K(+) under K(+) depletion. However, the apparent absence of a clear renal phenotype in HKA2-null mice has led to the idea that this transporter is an epiphenomenon. This review summarizes past and recent findings regarding the functional, structural and physiological characteristics of H-K-ATPase type 2. The findings discussed in this review suggest that, as in the famous story, the ugly duckling of the X-K-ATPase family is actually a swan.

A link between fertility and K+ homeostasis: role of the renal H,K-ATPase type 2.

Renal K(+) retention is activated during pregnancy through a mechanism unknown to date. Here, we showed that the renal stimulation of H,K-ATPase type 2 (HKA2), whose expression was recently identified to be progesterone-dependent, is part of the mechanism favoring K(+) accumulation during gestation. Moreover, investigation of the gestational phenotype of HKA2-null mice compared to their wild-type (WT) littermate revealed a decrease in fertility (gestation was successful in 33 % of HKA2-null mice vs. 83 % of WT mice) and in litter size (6.5 ± 0.6 and 7.8 ± 0.4 fetuses per litter, respectively). We also observed that urinary K(+) excretion decreased by 20 % and plasma K(+) concentration rose slightly (11 %) in WT mice during gestation (relative to basal conditions). In contrast, the renal excretion of K(+) and plasma K(+) levels in HKA2-null mice remained constant during gestation, whereas fecal K(+) excretion increased. As a consequence, HKA2-null mice did not accumulate K(+) in their extracellular compartment as efficiently as WT mice did. Finally, the link between inefficient K(+) balance adaptations and gestational complications was established when we observed that these complications could be reversed with an increased K(+) uptake. Altogether, these results define a novel physiological role for the HKA2 transporter and uncover a link between K(+) metabolism and fertility.

Expression profile of nuclear receptors along male mouse nephron segments reveals a link between ERRß and thick ascending limb function.

The nuclear receptor family orchestrates many functions related to reproduction, development, metabolism, and adaptation to the circadian cycle. The majority of these receptors are expressed in the kidney, but their exact quantitative localization in this ultrastructured organ remains poorly described, making it difficult to elucidate the renal function of these receptors. In this report, using quantitative PCR on microdissected mouse renal tubules, we established a detailed quantitative expression map of nuclear receptors along the nephron. This map can serve to identify nuclear receptors with specific localization. Thus, we unexpectedly found that the estrogen-related receptor ß (ERRß) is expressed predominantly in the thick ascending limb (TAL) and, to a much lesser extent, in the distal convoluted tubules. In vivo treatment with an ERR inverse agonist (diethylstilbestrol) showed a link between this receptor family and the expression of the Na⁺,K⁺-2Cl⁻ cotransporter type 2 (NKCC2), and resulted in phenotype presenting some similarities with the Bartter syndrom (hypokalemia, urinary Na⁺ loss and volume contraction). Conversely, stimulation of ERRß with a selective agonist (GSK4716) in a TAL cell line stimulated NKCC2 expression. All together, these results provide broad information regarding the renal expression of all members of the nuclear receptor family and have allowed us to identify a new regulator of ion transport in the TAL segments.

Circadian expression of H,K-ATPase type 2 contributes to the stability of plasma K⁺ levels.

Maintenance by the kidney of stable plasma K(+) values is crucial, as plasma K(+) controls muscle and nerve activity. Since renal K(+) excretion is regulated by the circadian clock, we aimed to identify the ion transporters involved in this process. In control mice, the renal mRNA expression of H,K-ATPase type 2 (HKA2) is 25% higher during rest compared to the activity period. Conversely, under dietary K(+) restriction, HKA2 expression is ∼40% higher during the activity period. This reversal suggests that HKA2 contributes to the circadian regulation of K(+) homeostasis. Compared to their wild-type (WT) littermates, HKA2-null mice fed a normal diet have 2-fold higher K(+) renal excretion during rest. Under K(+) restriction, their urinary K(+) loss is 40% higher during the activity period. This inability to excrete K(+) "on time" is reflected in plasma K(+) values, which vary by 12% between activity and rest periods in HKA2-null mice but remain stable in WT mice. Analysis of the circadian expression of HKA2 regulators suggests that Nrf2, but not progesterone, contributes to its rhythmicity. Therefore, HKA2 acts to maintain the circadian rhythm of urinary K(+) excretion and preserve stable plasma K(+) values throughout the day.