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Although lithium (LIT) therapy is key in managing bipolar disorder long-term, prolonged use significantly contributes to acquired Nephrogenic Diabetes Insipidus (NDI). This study examined whether combining Silymarin (SIL) with Vitamin C (Vit C) enhances protection against lithium-induced nephrotoxicity in rats, comparing their individual antioxidant effects as well. Rats subjected to Li exposure were provided with a standard commercial diet supplemented with 80 mmol LiCl per kilogram for 28 days. Concurrently, SIL and Vit C were administered orally at dosages of 200 and 100 mg/kg body weight, respectively, throughout the 28 days. The study assessed levels of reactive oxygen species (ROS), glutathione (GSH), and malondialdehyde (MDA), as well as the enzyme activity of superoxide dismutase (SOD), to evaluate the protective effects of SIL and Vit C against oxidative stress. Aquaporin-2 (AQP2) levels in kidney tissues were evaluated using immunohistochemistry and ELISA. Serum and urine parameters (sodium, potassium, creatinine, blood urea nitrogen [BUN], and urea) and serum lithium levels were also measured. Lithium-induced nephrotoxicity showed increased renal toxicity markers and decreased antioxidant enzyme activity. SIL administration significantly reduced markers of kidney tissue toxicity, increased antioxidant enzyme activities, regulated the aforementioned physiological parameters in blood and urine, and downregulated AQP2 expression in the kidney. However, Vit C administration did not demonstrate a significant protective effect against lithium-induced renal toxicity. These findings indicate that SIL effectively protects against lithium-induced nephrotoxicity, whereas Vitamin C does not exhibit this protective effect.

A 9-month-old male presented with vomiting and dehydration with mild hypernatremia in the context of failure to thrive. He was later diagnosed with nephrogenic diabetes insipidus (NDI) during this hospitalisation and was also found to have eosinophilic esophagitis (EoE). He has since been growing well after EoE and NDI were properly managed. Molecular genetic testing revealed an unreported deletion in AQP2 which was deemed pathogenic and of autosomal dominant inheritance when correlated with his clinical findings and family history. This case report describes the clinical course of this patient in comparison to his family members and reviews current literature on autosomal dominant NDI caused by AQP2 mutations., (© 2024 Asian Pacific Society of Nephrology.)

Background: Oryeongsan (Wulingsan, Goreisan) has long been used for the treatment of impaired body fluid metabolism. However, the action mechanisms have not been clearly defined. Recently, effects of Oryeongsan on the body fluid and Na + metabolism and the action mechanisms have been shown more clearly. The present review focuses on the recent findings on the effects of Oryeongsan in the cardio-renal system in relation with body fluid metabolism and action mechanisms leading to a decrease in blood pressure in animal models of hypertension., Methods: The new and recent findings were searched by using searching systems including PubMed-NCBI and Google-Scholar., Results: Oryeongsan induced an increase in glomerular filtration rate, and natriuresis and diuresis with a decreased osmolality and resulted in a contraction of the body fluid and Na + balance. These findings were associated with a suppression of abundance of Na + - H + -exchanger isoform 3 expression and V 2 receptor/aquaporin2 water channel signaling pathway in the kidney. Further, treatment with Oryeongsan accentuated atrial natriuretic peptide secretion in the atria from spontaneously hypertensive rats in which the secretion was suppressed. In addition, Oryeongsan ameliorated impaired vasodilation in spontaneously hypertensive rats., Conclusion: The effects of Oryeongsan in the kidney, atria, and vessel were accompanied by a suppression of AT 1 receptor and concurrent accentuation of abundance of AT 2 /Mas receptors expression and modulation of the natriuretic peptide system in these organs from hypertensive rats. The review shows multiple sites of action of Oryeongsan and mechanisms involved in the regulation of volume and pressure homeostasis in the body., Competing Interests: The authors declare that they have no conflicts of interest., (© 2024 Korea Institute of Oriental Medicine. Published by Elsevier B.V.)

High sugar-sweetened beverage intake has been related to human kidney disease and metabolic alterations. We determine the impact of high sucrose intake from pregnancy until early postnatal days and post-weaning on kidneys from adult male offspring rats. Wistar female rats were mated and assigned into two groups: one control drinking tap water (CM) and another drinking 5 % sucrose diluted in water (SM). Two offspring per mother were randomly allocated into two experimental groups at weaning. One had free access to simple water (CO) and the other to 5 % sucrose (SO) for 14 weeks. After treatment, levels of relative aquaporin-2 (AQP2), glomerulosclerosis index (GI), collecting tube area, perirenal fat, blood creatinine, and blood ureic nitrogen concentration (BUN) were determined. Two-way ANOVA followed by Bonferroni post-hoc test was used, considering P ≤ 0.05 as a significant statistical difference. Sucrose consumption during gestation/lactation and interaction increased AQP2 expression in the renal cortex and BUN concentration. In contrast, gestation/lactation consumption increased collecting tube area, post-weaning consumption favored perirenal fat, and finally, gestation/lactation, post-weaning, and the interaction caused glomerulosclerosis. Our results suggest that the consumption of sucrose water during gestation/lactation or post-weaning or combination triggers pathological changes in the kidneys of adult rats., Competing Interests: Declaration of competing interest The authors declare no conflicts of interest, financial or otherwise., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Aims: Sevoflurane is an ether-based inhalational anaesthetic that induces and maintains general anaesthesia. Our study aimed to detect sevoflurane-induced nephrogenic diabetes insipidus using data mining algorithms (DMAs) and molecular docking. The FAERS database was analysed using OpenVigil 2.1 for disproportionality analysis., Methods: We analysed FAERS data from 2004 to 2022 to determine the incidence of nephrogenic diabetes insipidus associated with sevoflurane. Reporting odds ratios (RORs) and proportional reporting ratios (PRRs) with 95% confidence intervals were calculated. We also used molecular docking with AutoDock Vina to examine sevoflurane's binding affinity to relevant receptors., Results: A total of 554 nephrogenic diabetes insipidus cases were reported in FAERS, of which 2.5% (14 cases) were associated with sevoflurane. Positive signals were observed for sevoflurane with ROR of 76.012 (95% CI: 44.67-129.35) and PRR of 75.72 (χ 2 : 934.688). Of the 14 cases, 50% required hospitalization, 14% resulted in death, and the remaining cases were categorized as other outcomes. Molecular docking analysis showed that sevoflurane exhibited high binding affinity towards AQP2 (4NEF) and AVPR2 (6U1N) with docking scores of -4.9 and -5.3, respectively., Conclusions: Sevoflurane use is significantly associated with the incidence of nephrogenic diabetes insipidus. Healthcare professionals should be cautious when using this medication and report any adverse events to regulatory agencies. Further research is needed to validate these findings and identify risk factors while performing statistical adjustments to prevent false-positives. Clinical monitoring is crucial to validate potential adverse effects of sevoflurane., (© 2023 The Authors. British Journal of Clinical Pharmacology published by John Wiley & Sons Ltd on behalf of British Pharmacological Society.)

Nephrogenic syndrome of inappropriate antidiuresis (NSIAD) is a rare X-linked disease caused by gain-of-function mutations of arginine vasopressin receptor 2 (V2R). Patients with NSIAD are characterized by the inability to excrete a free water load and by inappropriately increased urinary osmolality despite very low levels of plasma vasopressin, resulting in euvolaemic hyponatraemia. To dissect the signalling downstream V2R constitutively active variants, Flp-In T-REx Madin-Darby canine kidney (FTM) cells, stably transfected with V2R mutants (R137L, R137C and F229V) and AQP2-wt or non-phosphorylatable AQP2-S269A/AQP2-S256A, were used as cellular models. All three activating V2R mutations presented constitutive plasma membrane expression of AQP2-wt and significantly higher basal water permeability. In addition, V2R-R137L/C showed significantly higher activity of Rho-associated kinase (ROCK), a serine/threonine kinase previously suggested to be involved in S269-AQP2 phosphorylation downstream of these V2R mutants. Interestingly, FTM cells expressing V2R-R137L/C mutants and AQP2-S269A showed a significant reduction in AQP2 membrane abundance and a significant reduction in ROCK activity, indicating the crucial importance of S269-AQP2 phosphorylation in the gain-of-function phenotype. Conversely, V2R-R137L/C mutants retained the gain-of-function phenotype when AQP2-S256A was co-expressed. In contrast, cells expressing the F229V mutant and the non-phosphorylatable AQP2-S256A had a significant reduction in AQP2 membrane abundance along with a significant reduction in basal osmotic water permeability, indicating a crucial role of Ser256 for this mutant. These data indicate that the constitutive AQP2 trafficking associated with the gain-of-function V2R-R137L/C mutants causing NSIAD is protein kinase A independent and requires an intact Ser269 in AQP2 under the control of ROCK phosphorylation. KEY POINTS: Nephrogenic syndrome of inappropriate antidiuresis is caused by two constitutively active variant phenotypes of AVPR2, one sensitive to vaptans (V2R-F229V) and the other vaptan resistant (V2R-R137C/L). In renal cells, all three activating arginine vasopressin receptor 2 (V2R) variants display constitutive AQP2 plasma membrane expression and high basal water permeability. In cells expressing V2R-R137L/C mutants, disruption of the AQP2-S269 phosphorylation site caused the loss of the gain-of-function phenotype, which, in contrast, was retained in V2R-F229V-expressing cells. Cells expressing the V2R-F229V mutant were instead sensitive to disruption of the AQP2-S256 phosphorylation site. The serine/threonine kinase Rho-associated kinase (ROCK) was found to be involved in AQP2-S269 phosphorylation downstream of the V2R-R137L/C mutants. These findings might have clinical relevance for patients with nephrogenic syndrome of inappropriate antidiuresis., (© 2023 The Authors. The Journal of Physiology published by John Wiley & Sons Ltd on behalf of The Physiological Society.)

Aquaporin-2 (AQP2) is a member of the aquaporin water channel family. In the kidney, AQP2 is expressed in collecting duct principal cells where it facilitates water reabsorption in response to antidiuretic hormone (arginine vasopressin, AVP). AVP induces the redistribution of AQP2 from intracellular vesicles and its incorporation into the plasma membrane. The plasma membrane insertion of AQP2 represents the crucial step in AVP-mediated water reabsorption. Dysregulation of the system preventing the AQP2 plasma membrane insertion causes diabetes insipidus (DI), a disease characterised by an impaired urine concentrating ability and polydipsia. There is no satisfactory treatment of DI available. This review discusses kinases that control the localisation of AQP2 and points out potential kinase-directed targets for the treatment of DI., (© 2023 Max Delbrück Center for Molecular Medicine in the Helmholtz Association. The Journal of Physiology published by John Wiley & Sons Ltd on behalf of The Physiological Society.)

High concentrations of urinary calcium counteract vasopressin action via the activation of the Calcium-Sensing Receptor (CaSR) expressed in the luminal membrane of the collecting duct cells, which impairs the trafficking of aquaporin-2 (AQP2). In line with these findings, we provide evidence that, with respect to wild-type mice, CaSR knock-in (KI) mice mimicking autosomal dominant hypocalcaemia, display a significant decrease in the total content of AQP2 associated with significantly higher levels of AQP2 phosphorylation at Ser261, a phosphorylation site involved in AQP2 degradation. Interestingly, KI mice also had significantly higher levels of phosphorylated p38MAPK, a downstream effector of CaSR and known to phosphorylate AQP2 at Ser261. Moreover, ATF1 phosphorylated at Ser63, a transcription factor downstream of p38MAPK, was significantly higher in KI. In addition, KI mice had significantly higher levels of AQP2-targeting miRNA137 consistent with a post-transcriptional downregulation of AQP2. In vivo treatment of KI mice with the calcilytic JTT-305, a CaSR antagonist, increased AQP2 expression and reduced AQP2-targeting miRNA137 levels in KI mice. Together, these results provide direct evidence for a critical role of CaSR in impairing both short-term vasopressin response by increasing AQP2-pS261, as well as AQP2 abundance, via the p38MAPK-ATF1-miR137 pathway. KEY POINTS: Calcium-Sensing Receptor (CaSR) activating mutations are the main cause of autosomal dominant hypocalcaemia (ADH) characterized by inappropriate renal calcium excretion leading to hypocalcaemia and hypercalciuria. Current treatments of ADH patients with parathyroid hormone, although improving hypocalcaemia, do not improve hypercalciuria or nephrocalcinosis. In vivo treatment with calcilytic JTT-305/MK-5442 ameliorates most of the ADH phenotypes of the CaSR knock-in mice including hypercalciuria or nephrocalcinosis and reverses the downregulation of the vasopressin-sensitive aquaporin-2 (AQP2) expression, providing direct evidence for a critical role of CaSR in impairing vasopressin response. The beneficial effect of calcilytic in reducing the risk of renal calcification may occur in a parathyroid hormone-independent action through vasopressin-dependent inhibition of cAMP synthesis in the thick ascending limb and in the collecting duct. The amelioration of most of the abnormalities in calcium metabolism including hypercalciuria, renal calcification, and AQP2-mediated osmotic water reabsorption makes calcilytic a good candidate as a novel therapeutic agent for ADH., (© 2024 The Authors. The Journal of Physiology © 2024 The Physiological Society.)

The advent of modern quantitative protein mass spectrometry techniques around the turn of the 21st century has contributed to a revolution in biology referred to as 'systems biology'. These methods allow identification and quantification of thousands of proteins in a biological specimen, as well as detection and quantification of post-translational protein modifications including phosphorylation. Here, we discuss these methodologies and show how they can be applied to understand the effects of the peptide hormone vasopressin to regulate the molecular water channel aquaporin-2. The emerging picture provides a detailed framework for understanding the molecular mechanisms involved in water balance disorders., (Published 2022. This article is a U.S. Government work and is in the public domain in the USA.)

Despite continuous medical advancements, traumatic brain injury (TBI) remains a leading cause of death and disability worldwide. Consequently, there is a pursuit for biomarkers that allow non-invasive monitoring of patients after cranial trauma, potentially improving clinical management and reducing complications and mortality. Aquaporins (AQPs), which are crucial for transmembrane water transport, may be significant in this context. This study included 48 patients, with 27 having acute (aSDH) and 21 having chronic subdural hematoma (cSDH). Blood plasma samples were collected from the participants at three intervals: the first sample before surgery, the second at 15 h, and the third at 30 h post-surgery. Plasma concentrations of AQP1, AQP2, AQP4, and AQP9 were determined using the sandwich ELISA technique. CT scans were performed on all patients pre- and post-surgery. Correlations between variables were examined using Spearman's nonparametric rank correlation coefficient. A strong correlation was found between aquaporin 2 levels and the volume of chronic subdural hematoma and midline shift. However, no significant link was found between aquaporin levels (AQP1, AQP2, AQP4, and AQP9) before and after surgery for acute subdural hematoma, nor for AQP1, AQP4, and AQP9 after surgery for chronic subdural hematoma. In the chronic SDH group, AQP2 plasma concentration negatively correlated with the midline shift measured before surgery (Spearman's ρ -0.54; p = 0.017) and positively with hematoma volume change between baseline and 30 h post-surgery (Spearman's ρ 0.627; p = 0.007). No statistically significant correlation was found between aquaporin plasma levels and hematoma volume for AQP1, AQP2, AQP4, and AQP9 in patients with acute SDH. There is a correlation between chronic subdural hematoma volume, measured radiologically, and serum AQP2 concentration, highlighting aquaporins' potential as clinical biomarkers.

The dietary approach to stop hypertension (DASH) diet combines the antihypertensive effect of a low sodium and high potassium diet. In particular, the potassium component of the diet acts as a switch in the distal convoluted tubule to reduce sodium reabsorption, similar to a diuretic but without the side effects. Previous trials to understand the mechanism of the DASH diet were based on animal models and did not characterize changes in human ion channel protein abundance. More recently, protein cargo of urinary extracellular vesicles (uEVs) has been shown to mirror tissue content and physiological changes within the kidney. We designed an inpatient open label nutritional study transitioning hypertensive volunteers from an American style diet to DASH diet to examine physiological changes in adults with stage 1 hypertension otherwise untreated (Sacks FM, Svetkey LP, Vollmer WM, Appel LJ, Bray GA, Harsha D, Obarzanek E, Conlin PR, Miller ER 3rd, Simons-Morton DG, Karanja N, Lin PH; DASH-Sodium Collaborative Research Group. N Engl J Med 344: 3-10, 2001). Urine samples from this study were used for proteomic characterization of a large range of pure uEVs (small to large) to reveal kidney epithelium changes in response to the DASH diet. These samples were collected from nine volunteers at three time points, and mass spectrometry identified 1,800 proteins from all 27 samples. We demonstrated an increase in total SLC12A3 [sodium-chloride cotransporter (NCC)] abundance and a decrease in aquaporin-2 (AQP2) in uEVs with this mass spectrometry analysis, immunoblotting revealed a significant increase in the proportion of activated (phosphorylated) NCC to total NCC and a decrease in AQP2 from day 5 to day . This data demonstrates that the human kidney's response to nutritional interventions may be captured noninvasively by uEV protein abundance changes. Future studies need to confirm these findings in a larger cohort and focus on which factor drove the changes in NCC and AQP2, to which degree NCC and AQP2 contributed to the antihypertensive effect and address if some uEVs function also as a waste pathway for functionally inactive proteins rather than mirroring protein changes. 11 Numerous studies link DASH diet to lower blood pressure, but its mechanism is unclear. Urinary extracellular vesicles (uEVs) offer noninvasive insights, potentially replacing tissue sampling. Transitioning to DASH diet alters kidney transporters in our stage 1 hypertension cohort: AQP2 decreases, NCC increases in uEVs. This aligns with increased urine volume, reduced sodium reabsorption, and blood pressure decline. Our data highlight uEV protein changes as diet markers, suggesting some uEVs may function as waste pathways. We analyzed larger EVs alongside small EVs, and NCC in immunoblots across its molecular weight range.NEW & NOTEWORTHY Numerous studies link DASH diet to lower blood pressure, but its mechanism is unclear. Urinary extracellular vesicles (uEVs) offer noninvasive insights, potentially replacing tissue sampling. Transitioning to DASH diet alters kidney transporters in our stage 1 hypertension cohort: AQP2 decreases, NCC increases in uEVs. This aligns with increased urine volume, reduced sodium reabsorption, and blood pressure decline. Our data highlight uEV protein changes as diet markers, suggesting some uEVs may function as waste pathways. We analyzed larger EVs alongside small EVs, and NCC in immunoblots across its molecular weight range.

Purpose: Intermittent fasting (IF) has been shown to induce a well-organized adaptive defense against stress inside the cells, which increases the production of anti-oxidant defenses, repair of DNA, biogenesis of mitochondria, and genes that combat inflammation. So, the goal of the current investigation was to identify the effects of IF on rats with adriamycin (ADR)-induced nephropathy and any potential underlying mechanisms., Methods: Four groups of 40 mature Sprague-Dawley male rats were allocated as follow; control, fasting, ADR, and ADR plus fasting. After 8 weeks of ADR administration urine, blood samples and kidneys were taken for assessment of serum creatinine (Cr), BUN, urinary proteins, indicators of oxidative damage (malondialdehyde (MDA), reduced glutathione (GSH) and Catalase (CAT) levels), histopathological examinations, immunohistochemical examinations for caspase-3, Sirt1, aquaporin2 (AQP2) and real time PCR for antioxidant genes; Nrf2, HO-1 in kidney tissues., Results: IF significantly improved serum creatinine, BUN and urinary protein excretion, oxidative stress (low MDA with high CAT and GSH), in addition to morphological damage to the renal tubules and glomeruli as well as caspase-3 production during apoptosis. Moreover, IF stimulates significantly the expression of Sirt1 and Nrf2/HO-1 and AQP2., Conclusion: AQP2, Sirt1, Nrf2/HO-1 signaling may be upregulated and activated by IF, which alleviates ADR nephropathy. Enhancing endogenous antioxidants, reducing apoptosis and tubulointerstitial damage, and maintaining the glomerular membrane's integrity are other goals., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Background: Restoration of blood supply is a desired goal for the treatment of acute ischemic stroke. However, the restoration often leads to cerebral ischemia-reperfusion injury (CIR/I), which greatly increases the risk of non-neural organ damage. In particular, the acute kidney injury might be one of the most common complications., Aims: The study aimed to understand the damage occurred and the potential molecular mechanisms., Methods: The study was explored on the CIR/I rats generated by performing middle cerebral artery occlusion/reperfusion (MCAO/Reperfusion). The rats were evaluated with injury on the brains, followed by the non-neural organs including kidneys, livers, colons and stomachs. They were examined further with histopathological changes, and gene expression alterations by using RT-qPCR of ten aquaporins ( Aqps ) subtypes including Aqp1 ～ Aqp9 and Aqp11 . Furthermore, the Aqps expression profiles were constructed for each organ and analyzed by performing Principle Component Analysis. In addition, immunohistochemistry was explored to look at the protein expression of Aqp1 , Aqp2 , Aqp3 and Aqp4 in the rat kidneys., Results: There was a prominent down-regulation profile in the MCAO/Reperfusion rat kidneys. The protein expression of Aqp1, Aqp2, Aqp3 and Aqp4 was decreased in the kidneys of the MCAO/Reperfusion rats. We suggested that the kidney was in the highest risk to be damaged following the CIR/I. Down-regulation of Aqp2, Aqp3 and Aqp4 was involved in the acute kidney injury induced by the CIR/I., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2024 The Authors.)

Aquaporins (AQPs) constitute a wide family of water channels implicated in all kind of physiological processes. Zinc is the second most abundant trace element in the human body and a few studies have highlighted regulation of AQP0 and AQP4 by zinc. In the present work, we addressed the putative regulation of AQPs by zinc cations in silico through molecular dynamics simulations of human AQP0, AQP2, AQP4, and AQP5. Our results align with other scales of study and several in vitro techniques, hence strengthening the reliability of this regulation by zinc. We also described two distinct putative molecular mechanisms associated with the increase or decrease in AQPs' water permeability after zinc binding. In association with other studies, our work will help deciphering the interaction networks existing between zinc and channel proteins.

The shuttling of renal collecting duct aquaporin-2 (AQP2) between intracellular vesicles and the apical plasma membrane is paramount for regulation of renal water reabsorption. The binding of the circulating antidiuretic hormone arginine vasopressin (AVP) to the basolateral AVP receptor increases intracellular cAMP, which ultimately leads to AQP2 plasma membrane accumulation via a dual effect on AQP2 vesicle fusion with the apical plasma membrane and reduced AQP2 endocytosis. This AQP2 plasma membrane accumulation increases water reabsorption and consequently urine concentration. Conventional fluorescent microscopy provides a lateral resolution of ∼250 nm, which is insufficient to resolve the AQP2-containing endosomes/vesicles. Therefore, detailed information regarding the AQP2 vesicular population is still lacking. Newly established 4.5x Expansion Microscopy (ExM) can increase resolution to 60-70 nm. Using 4.5x ExM, we detected AQP2 vesicles/endosomes as small as 79 nm considering an average expansion factor of 4.3 for endosomes. Using different markers of the endosomal system provided detailed information of the cellular AQP2 itinerary upon changes in endogenous cAMP levels. Before cAMP elevation, AQP2 colocalized with early and recycling, but not late endosomes. Forskolin-induced cAMP increase was characterized by AQP2 insertion into the plasma membrane and AQP2 withdrawal from large perinuclear endosomes as well as some localization to lysosomal compartments. Forskolin washout promoted AQP2 endocytosis where AQP2 localized to not only early and recycling endosomes but also late endosomes and lysosomes indicating increased AQP2 degradation. Thus, our results show that 4.5 ExM is an attractive approach to obtain detailed information regarding AQP2 shuttling. NEW & NOTEWORTHY Renal aquaporin-2 (AQP2) imaged by expansion microscopy provides unprecedented 3-D information regarding the AQP2 itinerary in response to changes in cellular cAMP.

Aquaporin-2 (AQP2) water channels are proteins that are recycled between intracellular vesicles and the apical plasma membrane in renal collecting ducts. Lipopolysaccharide-responsive beige-like anchor protein (LRBA) is a protein kinase A (PKA) anchoring protein that creates compartmentalized PKA signalling responsible for AQP2 phosphorylation. In response to increased plasma osmolality, vasopressin/cyclic adenosine monophosphate (cAMP)/PKA signalling phosphorylates AQP2, promoting AQP2 trafficking into the apical plasma membrane and increasing water reabsorption from urine. However, the molecular mechanisms by which LRBA mediates vasopressin-induced AQP2 phosphorylation remain unknown. To investigate AQP2 intracellular localization and phosphorylation status in vivo, a density gradient ultracentrifugation technique was combined with an in situ proximity ligation assay, super-resolution structured illumination microscopy and immunoelectron microscopy. Most of the AQP2 was localized on the recycling endosome in the presence of tolvaptan, a vasopressin type 2 receptor (V2R) antagonist. Desmopressin, a V2R agonist, phosphorylated AQP2, translocating it from the recycling endosome to the apical plasma membrane. In contrast, LRBA was constitutively localized at the recycling endosome. Therefore, LRBA and AQP2 were well colocalized in the absence of vasopressin stimulation. The loss of LRBA/PKA signalling by Lrba knockout impaired vasopressin-induced AQP2 phosphorylation, resulting in AQP2 retention at the recycling endosome. Defective AQP2 trafficking caused low urinary concentrating ability in Lrba -/- mice. The LRBA-PKA complex created compartmentalized PKA signalling at the recycling endosome, which facilitated AQP2 phosphorylation in response to vasopressin. KEY POINTS: Membrane proteins are continuously internalized into the endosomal system via endocytosis, after which they are either recycled back to the plasma membrane or degraded at the lysosome. In T cells, lipopolysaccharide-responsive beige-like anchor protein (LRBA) binds directly to the cytotoxic T lymphocyte antigen 4 (CTLA-4), a checkpoint immune molecule, to prevent CTLA-4 lysosomal degradation and promote its vesicle recycling. LRBA has different physiological functions in renal collecting ducts. LRBA and aquaporin-2 (AQP2) water channels were colocalized on the recycling endosome in vivo in the absence of the anti-diuretic hormone vasopressin. LRBA promoted vasopressin-induced AQP2 trafficking, increasing water reabsorption from urine via AQP2. LRBA determined renal responsiveness to vasopressin at recycling endosomes. LRBA is a ubiquitously expressed anchor protein. LRBA signalosomes might regulate membrane trafficking of several constitutively recycled proteins at recycling endosomes., (© 2023 The Authors. The Journal of Physiology © 2023 The Physiological Society.)

Final urine volume and concentration are defined by water reabsorption through the water channel proteins aquaporin (AQP)-2, -3 and -4 in the collecting duct. However, the transcriptional regulation of these AQPs is not well understood. The Hippo/Yes-associated protein 1 (YAP) pathway plays an important role in organ size control and tissue homeostasis. When the Hippo pathway including the Mst1/Mst2 kinases is inhibited, YAP is activated and functions as a transcription co-activator. Our previous work revealed a pathological role of tubular YAP activation in chronic kidney disease, but the physiological role of YAP in the kidney remains to be established. Here, we found that tubule-specific Yap knockout mice showed increased urine output and decreased urinary osmolality. Decreases in Aqp2, -3 and -4 mRNA and protein abundance in the kidney were evident in Yap knockout mice. Analysis of Mst1/Mst2 double knockout and Mst1/Mst2/Yap triple knockout mice showed that expression of Aqp2 and Aqp4 but not Aqp3 was dependent on YAP. Furthermore, YAP was recruited to the promoters of the Aqp2 and Aqp4 genes and stimulated their transcription. Interestingly, YAP was found to interact with transcription factors GATA2, GATA3 and NFATc1. These three factors promoted Aqp2 transcription in a YAP dependent manner in collecting duct cells. These three factors also promoted Aqp4 transcription whereas only GATA2 and GATA3 enhanced Aqp3 transcription. Thus, our results suggest that YAP promotes Aqp2 and Aqp4 transcription, interacts with GATA2, GATA3 and NFATc1 to control Aqp2 expression, while Aqp-2, -3 and -4 exploit overlapping mechanisms for their baseline transcriptional regulation., (Copyright © 2022 International Society of Nephrology. All rights reserved.)

Serum sodium disorders are generally a marker of water balance in the body. Thus, hypernatremia is most often caused by an overall deficit of total body water. Other unique circumstances may lead to excess salt, without an impact on the body's total water volume. Hypernatremia is commonly acquired in both the hospital and community. As hypernatremia is associated with increased morbidity and mortality, treatment should be initiated promptly. In this review, we will discuss the pathophysiology and management of the main types of hypernatremia, which can be categorized as either a loss of water or gain of sodium that can be mediated by renal or extrarenal mechanisms., (Copyright © 2022 National Kidney Foundation, Inc. Published by Elsevier Inc. All rights reserved.)

SAP97 is a member of the MAGUK family of proteins, but unlike other MAGUK proteins that are selectively expressed in the CNS, SAP97 is also expressed in peripheral organs, like the heart and kidneys. SAP97 has several protein binding cassettes, and this review will describe their involvement in creating SAP97-anchored multiprotein networks. SAP97-anchored networks localized at the inner leaflet of the cell membrane play a major role in trafficking and targeting of membrane G protein-coupled receptors (GPCR), channels, and structural proteins. SAP97 plays a major role in compartmentalizing voltage gated sodium and potassium channels to specific cellular compartments of heart cells. SAP97 undergoes extensive alternative splicing. These splice variants give rise to different SAP97 isoforms that alter its cellular localization, networking, signaling and trafficking effects. Regarding GPCR, SAP97 binds to the β 1 -adrenergic receptor and recruits AKAP5/PKA and PDE4D8 to create a multiprotein complex that regulates trafficking and signaling of cardiac β 1 -AR. In the kidneys, SAP97 anchored networks played a role in trafficking of aquaporin-2 water channels. Cardiac specific ablation of SAP97 (SAP97-cKO) resulted in cardiac hypertrophy and failure in aging mice. Similarly, instituting transverse aortic constriction (TAC) in young SAP97 c-KO mice exacerbated TAC-induced cardiac remodeling and dysfunction. These findings highlight a critical role for SAP97 in the pathophysiology of a number of cardiac and renal diseases, suggesting that SAP97 is a relevant target for drug discovery., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Aquaporins (AQPs) are water channels widely distributed in living organisms and involved in many pathophysiologies as well as in cell volume regulations (CVR). In the present study, based on the structural homology existing between mineralocorticoid receptors (MRs), glucocorticoid receptors (GRs), cholesterol consensus motif (CCM) and the extra-cellular vestibules of AQPs, we investigated the binding of corticosteroids on the AQP family through in silico molecular dynamics simulations of AQP2 interactions with cortisol. We propose, for the first time, a putative AQPs corticosteroid binding site (ACBS) and discussed its conservation through structural alignment. Corticosteroids can mediate non-genomic effects; nonetheless, the transduction pathways involved are still misunderstood. Moreover, a growing body of evidence is pointing toward the existence of a novel membrane receptor mediating part of these rapid corticosteroids' effects. Our results suggest that the naturally produced glucocorticoid cortisol inhibits channel water permeability. Based on these results, we propose a detailed description of a putative underlying molecular mechanism. In this process, we also bring new insights on the regulatory function of AQPs extra-cellular loops and on the role of ions in tuning the water permeability. Altogether, this work brings new insights into the non-genomic effects of corticosteroids through the proposition of AQPs as the membrane receptor of this family of regulatory molecules. This original result is the starting point for future investigations to define more in-depth and in vivo the validity of this functional model.

We previously reported the novel finding that β3-AR is functionally expressed in the renal tubule and shares its cellular localization with the vasopressin receptor AVPR2, whose physiological stimulation triggers antidiuresis by increasing the plasma membrane expression of the water channel AQP2 and the NKCC2 symporter in renal cells. We also showed that pharmacologic stimulation of β3-AR is capable of triggering antidiuresis and correcting polyuria, in the knockout mice for the AVPR2 receptor, the animal model of human X-linked nephrogenic diabetes insipidus (XNDI), a rare genetic disease still missing a cure. Here, to demonstrate that the same response can be evoked in humans, we evaluated the effect of treatment with the β3-AR agonist mirabegron on AQP2 and NKCC2 trafficking, by evaluating their urinary excretion in a cohort of patients with overactive bladder syndrome, for the treatment of which the drug is already approved. Compared to baseline, treatment with mirabegron significantly increased AQP2 and NKCC2 excretion for the 12 weeks of treatment. This data is a step forward in corroborating the hypothesis that in patients with XNDI, treatment with mirabegron could bypass the inactivation of AVPR2, trigger antidiuresis and correct the dramatic polyuria which is the main hallmark of this disease.

Serine proteases (SPs) play physiological roles in the kidney. We previously reported that a synthetic SP inhibitor, camostat mesilate (CM), suppressed sodium reabsorption in the renal tubule and showed natriuretic effects in aldosterone-infused rats. Here, we aimed to explore novel physiological roles of SPs in the renal tubule and understand the mechanism of actions of SP inhibitors, by administering CM to healthy rats. Sprague-Dawley rats were classified into control and CM (subcutaneous sustained-release pellet) groups and sacrificed on day 7. CM significantly increased urine volumes by approximately two-fold in a urinary sodium- and osmolyte excretion-independent manner, indicating the occurrence of free water excretion. Serum vasopressin, potassium, and calcium levels and the osmolality in the renal medulla, which all affect free water reabsorption in the renal tubule, remained unchanged after CM administration. CM decreased urinary exosomal AQP2 excretion, suggesting suppression of AQP2 activity in the collecting duct. These changes were reversed by desmopressin infusion. Water diuresis caused by CM was independent of its action on prostasin or TMPRSS4. Our results revealed the association of SP inhibition with free water handling and demonstrated that CM administration exerted diuretic effects with AQP2 downregulation, suggesting SP inhibitors as a new class of aquaretic drugs., Competing Interests: Declaration of competing interest The authors indicated no potential conflicts of interest., (Copyright © 2022 The Authors. Production and hosting by Elsevier B.V. All rights reserved.)

The Growth Arrest-Specific protein 1 (Gas1) has been recently described in kidney as an endogenous inhibitor of cell proliferation in mesangial cells and with an important role in the maintenance of nephron progenitor cells. Furthermore, the expression of Gas1 was demonstrated in NCAM + progenitor parietal cells of Bowman's capsule. Thus, the aim of this study was to analyze the expression of Gas1 in the collecting ducts (CD) of healthy rats and to examine whether high glucose levels modify its expression during the early stages of diabetes in STZ-treated rats. Immunofluorescence reveals that principal cells AQP2 + express Gas1 in both healthy and diabetic conditions. Western blot from enriched fractions of medullary CD suggests that diabetes promotes the increase of Gas1. AQP2 + cells are also positive for the expression of CD24 and CD1133 in diabetic rats. In addition, diabetes modifies the cell morphology in the CD and favors the increase of principal cells (AQP2+/Gas1+), induces a significant decrease of intercalated cells (V-ATPase+/Gas1-) and the presence of intermediate cells (Gas1+/V-ATPase+) which express both principal and intercalated cell markers. The expression of Gas1 in the distal tubules was also determined by immunofluorescence, western blot and ELISA in diabetic rats. The results identify Gas1 as a specific marker of principal cells in healthy and diabetic rats and suggest that diabetes promotes the expression of Gas1. Gas1 may have an important role in the maintenance and differentiation to principal cells in the CD during early stages of diabetes., (© 2022. The Author(s), under exclusive licence to Springer Nature B.V.)

Introduction: Phosphatase and tensin (PTEN) is a multifunctional gene associated with the normal development and physiological function of various tissues including the kidney. However, its role in renal tubular reabsorption function has not been well elucidated., Methods: We generated a renal tubule-specific Pten knockout mouse model by crossing Pten fl/fl mice with Ksp-Cre transgenic mice, evaluated the effect of Pten loss on renal tubular function, and investigated the underlying mechanisms., Results: Pten loss resulted in abnormal renal structure and function and water retention in multiple organs. Our results also demonstrated that aquaporin-2 (AQP2), an important water channel protein, was upregulated and concentrated on the apical plasma membrane of collecting duct cells, which could be responsible for the impaired water balance in Pten loss mice. The regulation of Pten loss on AQP2 was mediated by protein kinase B (AKT) activation., Conclusions: Our results reveal a connection between PTEN gene inactivation and water retention, suggesting the importance of PTEN in normal kidney development and function., Competing Interests: The authors have no conflicts of interest to declare., (Copyright © 2022 by The Author(s). Published by S. Karger AG, Basel.)

Water homeostasis is tightly regulated by the kidneys via the process of urine concentration. During reduced water intake, the antidiuretic hormone arginine vasopressin (AVP) binds to the vasopressin receptor type II (V2R) in the kidney to enhance countercurrent multiplication and medullary osmolality, and increase water reabsorption via aquaporin-2 (AQP2) water channels. The importance of this AVP, V2R, and AQP2 axis is highlighted by low urine osmolality and polyuria in people with various water balance disorders, including nephrogenic diabetes insipidus (NDI). ELF5 and nuclear factor of activated T cells 5 (NFAT5) are two transcription factors proposed to regulate Aqp2 expression, but their role is poorly defined. Here we generated two novel mouse lines with principal cell (PC)-specific deletion of ELF5 or NFAT5 and phenotyped them in respect to renal water handling. ELF5-deficient mice (ELF5 PC-KO ) had a very mild phenotype, with no clear differences in AQP2 abundance, and mild differences in renal water handling and maximal urinary concentrating capacity. In contrast, NFAT5 (NFAT5 PC-KO ) mice had significantly higher water intake and their 24 h urine volume was almost 10-fold greater than controls. After challenging with dDAVP or 8 h water restriction, NFAT5 PC-KO mice were unable to concentrate their urine, demonstrating that they suffer from NDI. The abundance of AQP2, other AQPs, and the urea transporter UT-A1 were greatly decreased in NFAT5 PC-KO mice. In conclusion, NFAT5 is a major regulator of not only Aqp2 gene transcription, but also other genes important for water homeostasis and its absence leads to the development of NDI., (© 2022 The Authors. The FASEB Journal published by Wiley Periodicals LLC on behalf of Federation of American Societies for Experimental Biology.)

Protein kinase A (PKA) directly phosphorylates aquaporin-2 (AQP2) water channels in renal collecting ducts to reabsorb water from urine for the maintenance of systemic water homeostasis. More than 50 functionally distinct PKA-anchoring proteins (AKAPs) respectively create compartmentalized PKA signaling to determine the substrate specificity of PKA. Identification of an AKAP responsible for AQP2 phosphorylation is an essential step toward elucidating the molecular mechanisms of urinary concentration. PKA activation by several compounds is a novel screening strategy to uncover PKA substrates whose phosphorylation levels were nearly perfectly correlated with that of AQP2. The leading candidate in this assay proved to be an AKAP termed lipopolysaccharide-responsive and beige-like anchor protein (LRBA). We found that LRBA colocalized with AQP2 in vivo, and Lrba knockout mice displayed a polyuric phenotype with severely impaired AQP2 phosphorylation. Most of the PKA substrates other than AQP2 were adequately phosphorylated by PKA in the absence of LRBA, demonstrating that LRBA-anchored PKA preferentially phosphorylated AQP2 in renal collecting ducts. Furthermore, the LRBA-PKA interaction, rather than other AKAP-PKA interactions, was robustly dissociated by PKA activation. AKAP-PKA interaction inhibitors have attracted attention for their ability to directly phosphorylate AQP2. Therefore, the LRBA-PKA interaction is a promising drug target for the development of anti-aquaretics.

Aim: This study investigated whether enhanced histone acetylation, achieved by inhibiting histone deacetylases (HDACs), could prevent decreased aquaporin-2 (AQP2) expression during hypokalaemia., Methods: Male Wistar rats were fed a potassium-free diet with or without 4-phenylbutyric acid (4-PBA) or the selective HDAC3 inhibitor RGFP966 for 4 days. Primary renal inner medullary collecting duct (IMCD) cells and immortalized mouse cortical collecting duct (mpkCCD) cells were cultured in potassium-deprivation medium with or without HDAC inhibitors., Results: 4-PBA increased the levels of AQP2 mRNA and protein in the kidney inner medullae in hypokalaemic (HK) rats, which was associated with decreased urine output and increased urinary osmolality. The level of acetylated H3K27 (H3K27ac) protein was decreased in the inner medullae of HK rat kidneys; this decrease was mitigated by 4-PBA. The H3K27ac levels were decreased in IMCD and mpkCCD cells cultured in potassium-deprivation medium. Decreased H3K27ac in the Aqp2 promoter region was associated with reduced Aqp2 mRNA levels. HDAC3 protein expression was upregulated in mpkCCD and IMCD cells in response to potassium deprivation, and the binding of HDAC3 to the Aqp2 promoter was also increased. RGFP966 increased the levels of H3K27ac and AQP2 proteins and enhanced binding between H3K27ac and AQP2 in mpkCCD cells. Furthermore, RGFP966 reversed the hypokalaemia-induced downregulation of AQP2 and H3K27ac and alleviated polyuria in rats. RGFP966 increased interstitial osmolality in the kidney inner medullae of HK rats but did not affect urinary cAMP levels., Conclusion: HDAC inhibitors prevented the downregulation of AQP2 induced by potassium deprivation, probably by enhancing H3K27 acetylation., (© 2022 Scandinavian Physiological Society. Published by John Wiley & Sons Ltd.)

Arginine vasopressin (AVP) stimulates the concentration of renal urine by increasing the principal cell expression of aquaporin-2 (AQP2) water channels. Prostaglandin E 2 (PGE 2 ) and prostaglandin 2α (PGF 2α ) increase the water absorption of the principal cell without AVP, but PGE 2 decreases it in the presence of AVP. The underlying mechanism of this paradoxical response was investigated here. Mouse cortical collecting duct (mkpCCD c14 ) cells mimic principal cells as they endogenously express AQP2 in response to AVP. PGE 2 increased AQP2 abundance without desmopressin (dDAVP), while in the presence of dDAVP, PGE 2 , and PGF 2α reduced AQP2 abundance. dDAVP increased the cellular PGD 2 and PGE 2 release and decreased the PGF 2α release. MpkCCD cells expressed mRNAs for the receptors of PGE 2 (EP1/EP4), PGF 2 (FP), and TxB 2 (TP). Incubation with dDAVP increased the expression of EP1 and FP but decreased the expression of EP4. In the absence of dDAVP, incubation of mpkCCD cells with an EP4, but not EP1/3, agonist increased AQP2 abundance, and the PGE 2 -induced increase in AQP2 was blocked with an EP4 antagonist. Moreover, in the presence of dDAVP, an EP1/3, but not EP4, agonist decreased the AQP2 abundance, and the addition of EP1 antagonists prevented the PGE 2 -mediated downregulation of AQP2. Our study shows that in mpkCCD c14 cells, reduced EP4 receptor and increased EP1/FP receptor expression by dDAVP explains the differential effects of PGE 2 and PGF 2α on AQP2 abundance with or without dDAVP. As the V2R and EP4 receptor, but not the EP1 and FP receptor, can couple to Gs and stimulate the cyclic adenosine monophosphate (cAMP) pathway, our data support a view that cells can desensitize themselves for receptors activating the same pathway and sensitize themselves for receptors of alternative pathways., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Deen, Boone, Schweer, Olesen, Carmone, Wetzels, Fenton and Kortenoeven.)

The cAMP-dependent aquaporin-2 (AQP2) redistribution from intracellular vesicles into the plasma membrane of renal collecting duct principal cells induces water reabsorption and fine-tunes body water homeostasis. However, the mechanisms controlling the localization of AQP2 are not understood in detail. Using immortalized mouse medullary collecting duct (MCD4) and primary rat inner medullary collecting duct (IMCD) cells as model systems, we here discovered a key regulatory role of Aurora kinase A (AURKA) in the control of AQP2. The AURKA-selective inhibitor Aurora-A inhibitor I and novel derivatives as well as a structurally different inhibitor, Alisertib, prevented the cAMP-induced redistribution of AQP2. Aurora-A inhibitor I led to a depolymerization of actin stress fibers, which serve as tracks for the translocation of AQP2-bearing vesicles to the plasma membrane. The phosphorylation of cofilin-1 (CFL1) inactivates the actin-depolymerizing function of CFL1. Aurora-A inhibitor I decreased the CFL1 phosphorylation, accounting for the removal of the actin stress fibers and the inhibition of the redistribution of AQP2. Surprisingly, Alisertib caused an increase in actin stress fibers and did not affect CFL1 phosphorylation, indicating that AURKA exerts its control over AQP2 through different mechanisms. An involvement of AURKA and CFL1 in the control of the localization of AQP2 was hitherto unknown.

Regulated vesicle exocytosis is a key response to extracellular stimuli in diverse physiological processes, including hormone regulated short-term urine concentration. In the renal collecting duct, the water channel aquaporin-2 (AQP2) localizes to the apical plasma membrane as well as to small, subapical vesicles. In response to stimulation with the antidiuretic hormone, arginine vasopressin, aquaporin-2-containing vesicles fuse with the plasma membrane, which increases collecting duct water reabsorption and thus, urine concentration. The nanoscale size of these vesicles has limited analysis of their three-dimensional (3D) organization. Using a cell system combined with 3D superresolution microscopy, we provide the first direct analysis of the 3D network of aquaporin-2-containing exocytic vesicles in a cell culture system. We show that aquaporin-2 vesicles are 43 ± 3 nm in diameter, a size similar to synaptic vesicles, and that one fraction of AQP2 vesicles localized with the subcortical F-actin layer and the other localized in between the F-actin layer and the plasma membrane. Aquaporin-2 vesicles associated with F-actin and this association were enhanced in a serine 256 phospho-mimic of aquaporin-2, whose phosphorylation is a key event in antidiuretic hormone-mediated aquaporin-2 vesicle exocytosis.

Acquired forms of nephrogenic diabetes insipidus (NDI) include lithium (Li)-induced and hypokalemia-induced NDI. Both forms are associated with AQP2 downregulation and collecting duct (CD) cellular remodeling. Statins are cholesterol-lowering drugs appearing to increase AQP2 membrane-translocation and improve urine concentration in other NDI models. We have investigated if statins are able to prevent or rescue the Li-induced changes in mice and in a mouse cortical CD cell line (mCCD c1l ). Biotinylation assays showed that acute (1hr) atorvastatin, simvastatin, or fluvastatin increased AQP2 membrane accumulation in mCCD c1l cells showing that the cell line responds to acute statin treatment. To see whether chronic statin treatment abolish the Li effects, mCCD c1l cells were treated with 48 h Li, combined Li/atorvastatin or combined Li/simvastatin. Li reduced AQP2, but combined Li/atorvastatin or Li/simvastatin did not prevent AQP2 downregulation. In mice, chronic (21 days) Li increased urine output and reduced urine osmolality, but combined Li/atorvastatin did not prevent these effects. In inner medulla (IM), Li reduced total AQP2 and increased pS261-AQP2. Combined Li/atorvastatin did not abolish these changes. Atorvastatin did not prevent a Li-induced increase in intercalated cells and proliferation in IM. In mice with already established NDI, atorvastatin had no effect on the Li-induced changes either. Mice subjected to 14 days of potassium-deficient diet developed polyuria and AQP2 downregulation in IM. Co-treatment with atorvastatin did not prevent this. In conclusion, atorvastatin does not appear to be able to prevent or rescue Li-NDI or to prevent hypokalemic-induced NDI., (© 2021 The Authors. Physiological Reports published by Wiley Periodicals LLC on behalf of The Physiological Society and the American Physiological Society.)

Background: Congenital nephrogenic diabetes insipidus (NDI) is primarily caused by loss-of-function mutations in the vasopressin type 2 receptor (V2R). Renal unresponsiveness to the antidiuretic hormone vasopressin impairs aquaporin-2 (AQP2) water channel activity and water reabsorption from urine, resulting in polyuria. Currently available symptomatic treatments inadequately reduce patients' excessive amounts of urine excretion, threatening their quality of life. In the past 25 years, vasopressin/cyclic adenosine monophosphate (cAMP)/protein kinase A (PKA) has been believed to be the most important signaling pathway for AQP2 activation. Although cAMP production without vasopressin is the reasonable therapeutic strategy for congenital NDI caused by V2R mutations, the efficacy of candidate drugs on AQP2 activation is far less than that of vasopressin., Results: Intracellular distribution and activity of PKA are largely controlled by its scaffold proteins, A-kinase anchoring proteins (AKAPs). Dissociating the binding of AKAPs and PKA significantly increased PKA activity in the renal collecting ducts and activated AQP2 phosphorylation and trafficking. Remarkably, the AKAPs-PKA disruptor FMP-API-1 increased transcellular water permeability in isolated renal collecting ducts to the same extent as vasopressin. Moreover, derivatives of FMP-API-1 possessed much more high potency. FMP-API-1/27 is the first low-molecular-weight compound to be discovered that can phosphorylate AQP2 more effectively than preexisting drug candidates., Conclusion: AKAP-PKA disruptors are a promising therapeutic target for congenital NDI. In this article, we shall discuss the pathophysiological roles of PKA and novel strategies to activate PKA in renal collecting ducts., (© 2021. The Author(s).)

Aquaporin-2 plays a vital role in the human kidney as a water passage channel. Any disorder with its function can cause water imbalance and consequently disease in humans, especially nephrogenic diabetes insipidus (NDI). For this reason, an accurate understanding of its performance can be useful for therapeutic purposes. In this article, we investigate the gating mechanism induced by spontaneous fluctuations in aquaporin-2's (AQP2) channels in the palmitoyl-oleoyl-phosphatidyl-ethanolamine lipid bilayer by molecular dynamics. Our results show that the selectivity filter (SF) in AQP2 is also a gating site depending on the side-chain conformation of His172. The important role of His172 in modulating the wide and narrow conformations has been further investigated by the simulation of the H172G mutant. The osmotic permeability values of all four monomers are in the range of wide state and the average is very close to that of the wide channel formed by wild-type AQP2. Moreover, by calculating the osmotic permeability and the potential of mean force of each of the AQP2 monomers for wide/narrow states of the SF, it is seen that the SF at its narrow conformation can induce a much larger energy barrier for water molecules permeation, hindering the transport of water molecules remarkably. The reason for the discrepancy among osmotic permeabilities of different monomers of aquaporins is revealed by investigating the osmotic permeability of each monomer through the wide/narrow states of their SF., (© 2021 Wiley Periodicals LLC.)

Background: MicroRNAs (miRNAs), formed by cleavage of pre-microRNA by the endoribonuclease Dicer, are critical modulators of cell function by post-transcriptionally regulating gene expression., Methods: Selective ablation of Dicer in AQP2-expressing cells (Dicer AQP2Cre+ mice) was used to investigate the role of miRNAs in the kidney collecting duct of mice., Results: The mice had severe polyuria and nephrogenic diabetes insipidus, potentially due to greatly reduced AQP2 and AQP4 levels. Although epithelial sodium channel levels were decreased in cortex and increased in inner medulla, amiloride-sensitive sodium reabsorption was equivalent in Dicer AQP2Cre+ mice and controls. Small-RNA sequencing and proteomic analysis revealed 31 and 178 significantly regulated miRNAs and proteins, respectively. Integrated bioinformatic analysis of the miRNAome and proteome suggested alterations in the epigenetic machinery and various transcription factors regulating AQP2 expression in Dicer AQP2Cre+ mice. The expression profile and function of three miRNAs (miR-7688-5p, miR-8114, and miR-409-3p) whose predicted targets were involved in epigenetic control (Phf2, Kdm5c, and Kdm4a) or transcriptional regulation (GATA3, GATA2, and ELF3) of AQP2 were validated. Luciferase assays could not demonstrate direct interaction of AQP2 or the three potential transcription factors with miR-7688-5p, miR-8114, and miR-409-3p. However, transfection of respective miRNA mimics reduced AQP2 expression. Chromatin immunoprecipitation assays demonstrated decreased Phf2 and significantly increased Kdm5c interactions at the Aqp2 gene promoter in Dicer AQP2Cre+ mice, resulting in decreased RNA Pol II association., Conclusions: Novel evidence indicates miRNA-mediated epigenetic regulation of AQP2 expression., (Copyright © 2021 by the American Society of Nephrology.)

The molecular mechanisms underlying chronic kidney disease (CKD) are poorly understood and treatment options are limited, a situation underpinning the need for elucidating the causative molecular mechanisms and for identifying innovative treatment options. It is emerging that cyclic 3',5'-adenosine monophosphate (cAMP) signalling occurs in defined cellular compartments within nanometre dimensions in processes whose dysregulation is associated with CKD. cAMP compartmentalization is tightly controlled by a specific set of proteins, including A-kinase anchoring proteins (AKAPs) and phosphodiesterases (PDEs). AKAPs such as AKAP18, AKAP220, AKAP-Lbc and STUB1, and PDE4 coordinate arginine-vasopressin (AVP)-induced water reabsorption by collecting duct principal cells. However, hyperactivation of the AVP system is associated with kidney damage and CKD. Podocyte injury involves aberrant AKAP signalling. cAMP signalling in immune cells can be local and slow the progression of inflammatory processes typical for CKD. A major risk factor of CKD is hypertension. cAMP directs the release of the blood pressure regulator, renin, from juxtaglomerular cells, and plays a role in Na + reabsorption through ENaC, NKCC2 and NCC in the kidney. Mutations in the cAMP hydrolysing PDE3A that cause lowering of cAMP lead to hypertension. Another major risk factor of CKD is diabetes mellitus. AKAP18 and AKAP150 and several PDEs are involved in insulin release. Despite the increasing amount of data, an understanding of functions of compartmentalized cAMP signalling with relevance for CKD is fragmentary. Uncovering functions will improve the understanding of physiological processes and identification of disease-relevant aberrations may guide towards new therapeutic concepts for the treatment of CKD., (© 2021 The Authors. Acta Physiologica published by John Wiley & Sons Ltd on behalf of Scandinavian Physiological Society.)

Congenital nephrogenic diabetes insipidus (NDI) is a rare genetic disorder characterized by renal inability to concentrate urine. We utilized a multicenter strategy to investigate the genotype and phenotype in a cohort of Chinese children clinically diagnosed with NDI from 2014 to 2019. Ten boys from nine families were identified with mutations in AVPR2 or AQP2 along with dehydration, polyuria-polydipsia, and severe hypernatremia. Genetic screening confirmed the diagnosis of seven additional relatives with partial or subclinical NDI. Protein structural analysis revealed a notable clustering of diagnostic mutations in the transmembrane region of AVPR2 and an enrichment of diagnostic mutations in the C-terminal region of AQP2 . The pathogenic variants are significantly more likely to be located inside the domain compared with population variants. Through the structural analysis and in silico prediction, the eight mutations identified in this study were presumed to be disease-causing. The most common treatments were thiazide diuretics and non-steroidal anti-inflammatory drugs (NSAIDs). Emergency treatment for hypernatremia dehydration in neonates should not use isotonic saline as a rehydration fluid. Genetic analysis presumably confirmed the diagnosis of NDI in each patient in our study. We outlined methods for the early identification of NDI through phenotype and genotype, and outlined optimized treatment strategies., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Liao, Xiang, Li, Fang, Fang, Zhang, Cao, Zhai, Chen, Xu, Liu, Tang, Liu, Wang, Luan, Shen, Chen, Jiang, Ma, Xu and Rao.)

Emerging evidence is showing that apelin plays an important role in regulating salt and water balance by counteracting the antidiuretic action of vasopressin (AVP). However, the underlying mechanism remains unknown. Here, we hypothesized that (pro) renin receptor (PRR)/soluble prorenin receptor (sPRR) might mediate the diuretic action of apelin in the distal nephron. During water deprivation (WD), the urine concentrating capability was impaired by an apelin peptide, apelin-13, accompanied by the suppression of the protein expression of aquaporin 2 (AQP2), NKCC2, PRR/sPRR, renin and nuclear β-catenin levels in the kidney. The upregulated expression of AQP2 or PRR/sPRR both induced by AVP and 8-Br-cAMP was blocked by apelin-13, PKA inhibitor (H89), or β-catenin inhibitor (ICG001). Interestingly, the blockage of apelin-13 on AVP-induced AQP2 protein expression was reversed by exogenous sPRR. Together, the present study has defined the cyclic adenosine monophosphate (cAMP)/protein kinase A (PKA)/sPRR pathway in the CD as the molecular target of the diuretic action of apelin., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Chen, Xu, Hu, Deng, Qiu, Mo, Du and Yang.)

Vasopressin is the key hormone involved in water conservation and regulation of water balance, essential for life. In the renal collecting duct, vasopressin binds to the V2 receptor, increasing water permeability through activation of aquaporin-2 redistribution to the luminal membrane. This mechanism promotes rapid water reabsorption, important for immediate survival; however, only recently it has become clear that long-term adverse effects are associated with alterations of the vasopressin-aquaporin-2 pathway, leading to several syndromes associated with water balance disorders. The kidney resistance to the vasopressin action may cause severe dehydration for patients and, conversely, nonosmotic release of vasopressin is associated with water retention and increasing the circulatory blood volume. This chapter discusses the relevance of the altered vasopressin-aquaporin-2 pathway in some diseases associated with water balance disorders, including congenital nephrogenic diabetes insipidus, syndrome of inappropriate secretion of antidiuretic hormone, nephrogenic syndrome of inappropriate antidiuresis, and autosomal dominant polycystic kidney disease. The emerging picture suggests that targeting the vasopressin-AQP2 axis can provide therapeutic benefits in those patients., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Background: Primary nephrogenic diabetes insipidus (NDI) is a rare disorder and little is known about treatment practices and long-term outcome., Methods: Paediatric and adult nephrologists contacted through European professional organizations entered data in an online form., Results: Data were collected on 315 patients (22 countries, male 84%, adults 35%). Mutation testing had been performed in 270 (86%); pathogenic variants were identified in 258 (96%). The median (range) age at diagnosis was 0.6 (0.0-60) years and at last follow-up 14.0 (0.1-70) years. In adults, height was normal with a mean (standard deviation) score of -0.39 (±1.0), yet there was increased prevalence of obesity (body mass index >30 kg/m2; 41% versus 16% European average; P < 0.001). There was also increased prevalence of chronic kidney disease (CKD) Stage ≥2 in children (32%) and adults (48%). Evidence of flow uropathy was present in 38%. A higher proportion of children than adults (85% versus 54%; P < 0.001) received medications to reduce urine output. Patients ≥25 years were less likely to have a university degree than the European average (21% versus 35%; P = 0.003) but full-time employment was similar. Mental health problems, predominantly attention-deficit hyperactivity disorder (16%), were reported in 36% of patients., Conclusion: This large NDI cohort shows an overall favourable outcome with normal adult height and only mild to moderate CKD in most. Yet, while full-time employment was similar to the European average, educational achievement was lower, and more than half had urological and/or mental health problems., (© The Author(s) 2020. Published by Oxford University Press on behalf of ERA-EDTA. All rights reserved.)

As a rare hereditary disease, congenital nephrogenic diabetes insipidus (NDI) is clinically characterized by polyuria with hyposthenuria and polydipsia. NDI results from collecting duct principal cell hyporesponsiveness or insensitivity to the antidiuretic action of arginine vasopressin (AVP). The principal cell-specific water channel aquaporin-2 (AQP2) plays an essential role in water reabsorption along osmotic gradients. The capacity to accumulate AQP2 in the apical plasma membrane in response to decreased fluid volume or increased plasma osmolality is critically regulated by the antidiuretic hormone AVP and its receptor 2 (AVPR2). Mutations in AVPR2 result in X-linked recessive NDI, the most common form of inherited NDI. Genetic defects in AQP2 cause autosomal recessive or dominant NDI. In this review, we provide an updated overview of the genetic and molecular mechanisms of congenital NDI, with a focus on the potential disease-causing mutations in AVPR2 and AQP2 , the molecular defects in the AVPR2 and AQP2 mutants, post-translational modifications (i.e., phosphorylation, ubiquitination, and glycosylation) and various protein-protein interactions that regulate phosphorylation, ubiquitination, tetramerization, trafficking, stability, and degradation of AQP2.

Background: Chronic kidney diseases (CKD) are usually associated with dyslipidemia. Statin therapy has been primarily recommended for the prevention of cardiovascular risk in patients with CKD; however, the effects of statins on kidney disease progression remain controversial. This study aims to investigate the effects of statin treatment on renal handling of water in patients and in animals on a high-fat diet. Methods: Retrospective cohort patient data were reviewed and the protein expression levels of aquaporin-2 (AQP2) and NLRP3 inflammasome adaptor ASC were examined in kidney biopsy specimens. The effects of statins on AQP2 and NLRP3 inflammasome components were examined in nlrp3 -/- mice, 5/6 nephroectomized (5/6Nx) rats with a high-fat diet (HFD), and in vitro . Results: In the retrospective cohort study, serum cholesterol was negatively correlated to eGFR and AQP2 protein expression in the kidney biopsy specimens. Statins exhibited no effect on eGFR but abolished the negative correlation between cholesterol and AQP2 expression. Whilst nlrp3 +/+ mice showed an increased urine output and a decreased expression of AQP2 protein after a HFD, which was moderately attenuated in nlrp3 deletion mice with HFD. In 5/6Nx rats on a HFD, atorvastatin markedly decreased the urine output and upregulated the protein expression of AQP2. Cholesterol stimulated the protein expression of NLRP3 inflammasome components ASC, caspase-1 and IL-1β, and decreased AQP2 protein abundance in vitro , which was markedly prevented by statins, likely through the enhancement of ASC speck degradation via autophagy. Conclusion: Serum cholesterol level has a negative correlation with AQP2 protein expression in the kidney biopsy specimens of patients. Statins can ameliorate cholesterol-induced inflammation by promoting the degradation of ASC speck, and improve the expression of aquaporin in the kidneys of animals on a HFD., Competing Interests: Competing Interests: The authors have declared that no competing interest exists., (© The author(s).)

Today, plant-based therapies have been attracted attention to overcome diabetes complications. This study was an attempt to evaluate whether antidiabetic and nephroprotective effects of Stevia Rebaudiana Bertoni (SRB) can be exerted via upregulation of GLUT-4, SNAP23, and Stx4 in skeletal muscles or modulation of AQP2 mRNA expression and antioxidant signaling pathway activity (Nrf2/Keap1) in kidneys. To achieve this aim, diabetes was induced via STZ-nicotinamide (STZ-NA). Diabetes increased the level of Blood Urea Nitrogen (BUN), serum creatinine, Fasting Blood Sugar (FBS), and Keap1 mRNA expression, which was coincide with reduction in mRNA levels of Nrf2, GLUT4, SNAP23, and Stx4. SRB and metformin compensate mentioned variables. However, SRB extract was more effective than metformin to increase the levels of GLUT4 and Nrf2 mRNA. It seems that SRB might attenuate the diabetic complications via manipulating the glucose uptake components in peripheral tissues and might exert the nephroprotective effects by modulation of AQP2, and Nrf2/Keap1 mRNA expression. PRACTICAL APPLICATIONS: Synthetic antidiabetic drugs have been only partially successful in controlling the diabetic complications. Moreover, use of these drugs is associated with a number of adverse effects. Over the past few years, a renewed attention has been paid to the prevention and treatment of diabetes using medicinal plants and functional foods. SRB that have been known as natural sweetener for centuries, is a such natural agent that has high source of various phytochemicals with antidiabetic, renal protective, antitumor, and antioxidant properties. In the current study, possible molecular mechanisms of insulin-mimetic and nephroprotective effects of SRB extract was evaluated in diabetic rats. Due to powerful antihyperglycemic and nephroprotective effects of SRB extract that were showed in this study and previous studies, hence the fact that SRB is to be highlighted for future research as a new therapeutic agent for diabetes., (© 2020 Wiley Periodicals LLC.)

Objectives Congenital nephrogenic diabetes insipidus (NDI) is a rare hereditary disorder which is characterized by unresponsiveness to arginine vasopressin (AVP) in collecting ducts and leads to polyuria and polydipsia. The wide clinical spectrum of congenital NDI can cause difficulties in early diagnosis. We aimed to evaluate clinical prognosis of children with congenital NDI in long-term period. Methods Nineteen children with congenital NDI followed up in Pediatric Nephrology Department were enrolled to the study. This study is a single-center retrospective study, which reports clinical follow-up and genetic results of children with congenital NDI. Results Presenting symptoms of patients were mostly dehydration and fever due to polyuria and polydipsia. Four male patients had bilateral nonobstructive hydroureteronephrosis (HUN) and neurogenic bladder which requires clean intermittent catheterization (CIC). One patient had intracranial calcification which is a rarely seen complication in congenital NDI due to recurrent hypernatremic dehydration and severe brain dehydration. The causative mutations were identified in all patients. The identified mutations in six of them (31.6%) were hemizygous mutations in AVPR2 gene and homozygous mutations of AQP2 gene in the rest 13 cases (68.4%). More than that, four of these mutations (two in AVPR2 and two in AQP2) were novel mutations. Noncompliance with the treatments is associated with high risk of morbidity due to neurogenic bladder and chronic kidney disease (CKD). Conclusions The prognosis of congenital NDI is good when diagnosis can be made early and treatment is started immediately. Genetic counseling and prenatal testing for hereditary diseases are recommended especially in regions with relatively higher rates of consanguineous marriages.

NSIAD is a rare X-linked condition, caused by activating mutations in the AVPR2 gene coding for the vasopressin V2 receptor (V2R) associated with hyponatremia, despite undetectable plasma vasopressin levels. We have recently provided in vitro evidence that, compared to V2R-wt, expression of activating V2R mutations R137L, R137C and F229V cause a constitutive redistribution of the AQP2 water channel to the plasma membrane, higher basal water permeability and significantly higher basal levels of p256-AQP2 in the F229V mutant but not in R137L or R137C. In this study, V2R mutations were expressed in collecting duct principal cells and the associated signalling was dissected. V2R-R137L and R137C mutants had significantly higher basal pT269-AQP2 levels -independently of S256 and PKA-which were reduced to control by treatment with Rho kinase (ROCK) inhibitor. Interestingly, ROCK activity was found significantly higher in V2R-R137L along with activation of the Gα12/13-Rho-ROCK pathway. Of note, inhibition of ROCK reduced the basal elevated osmotic water permeability to control. To conclude, our data demonstrate for the first time that the gain-of-function mutation of the V2R, R137L causing NSIAD, signals through an alternative PKA-independent pathway that increases AQP2 membrane targeting through ROCK-induced phosphorylation at S/T269 independently of S256 of AQP2.

Vasopressin regulates renal water excretion by binding to a G α s-coupled receptor (V2R) in collecting duct cells, resulting in increased water permeability through regulation of the aquaporin-2 (AQP2) water channel. This action is widely accepted to be associated with cAMP-mediated activation of protein kinase A (PKA). Here, we use phosphoproteomics in collecting duct cells in which PKA has been deleted (CRISPR-Cas9) to identify PKA-independent responses to vasopressin. The results show that V2R-mediated vasopressin signaling is predominantly, but not entirely, PKA-dependent. Upregulated sites in PKA-null cells include Ser256 of AQP2, which is critical to regulation of AQP2 trafficking. In addition, phosphorylation changes in the protein kinases Stk39 (SPAK) and Prkci (an atypical PKC) are consistent with PKA-independent regulation of these protein kinases. Target motif analysis of the phosphopeptides increased in PKA-null cells indicates that vasopressin activates one or more members of the AMPK/SNF1-subfamily of basophilic protein kinases. In vitro phosphorylation assays using recombinant, purified SNF1-subfamily kinases confirmed postulated target specificities. Of interest, measured IBMX-dependent cAMP levels were an order of magnitude higher in PKA-null than in PKA-intact cells, indicative of a PKA-dependent feedback mechanism. Overall, the findings support the conclusion that V2-receptor mediated signaling in collecting duct cells is in part PKA-independent., (Published 2020. This article is a U.S. Government work and is in the public domain in the USA.)

Aldosterone indirectly regulates water reabsorption in the distal tubule by regulating sodium reabsorption. However, the direct effect of aldosterone on vasopressin-regulated water and urea permeability in the rat inner medullary collecting duct (IMCD) has not been tested. We investigated whether aldosterone regulates osmotic water permeability in isolated perfused rat IMCDs. Adding aldosterone (500 nM) to the bath significantly decreased osmotic water permeability in the presence of vasopressin (50 pM) in both male and female rat IMCDs. Aldosterone significantly decreased aquaporin-2 (AQP2) phosphorylation at S256 but did not change it at S261. Previous studies show that aldosterone can act both genomically and non-genomically. We tested the mechanism by which aldosterone attenuates osmotic water permeability. Blockade of gene transcription with actinomycin D did not reverse aldosterone-attenuated osmotic water permeability. In addition to AQP2, the urea transporter UT-A1 contributes to vasopressin-regulated urine concentrating ability. We tested aldosterone-regulated urea permeability in vasopressin-treated IMCDs. Blockade of gene transcription did not reverse aldosterone-attenuated urea permeability. In conclusion, aldosterone directly regulates water reabsorption through a non-genomic mechanism. Aldosterone-attenuated water reabsorption may be related to decreased trafficking of AQP2 to the plasma membrane. There may be a sex difference apparent in the inhibitory effect of aldosterone on water reabsorption in the inner medullary collecting duct. This study is the first to show a direct effect of aldosterone to inhibit vasopressin-stimulated osmotic water permeability and urea permeability in perfused rat IMCDs.

Congenital nephrogenic diabetes insipidus (CNDI) is a rare inherited disorder, mostly caused by antidiuretic hormone receptor type 2 (ADHR2) gene mutations, which are inherited as X-linked traits. Less than 10% of cases are due to mutations in the aquaporin-2 (AQP2) gene, inherited in autosomal recessive or dominant manner. We report the case of two adult sisters, of 30 and 27 years of age, diagnosed in early infancy with X-linked CNDI. The patients' sex and family history did not fit in well with this diagnosis, so we sequenced the coding regions of the ADHR2 and AQP2 genes. As expected, no mutations were found in the ADHR2 gene, while we found a compound heterozygosis for two different mutations in the AQP2 gene. A missense mutation (c. 439G>A, p.Ala147Thr), an already known cause of CNDI, and a novel missense putative mutation of an adenine to cytosine at position 551 (c.551A>C), resulting in the substitution of asparagine with threonine at amino acid position 184 (p.Asn184Thr). This second mutation changes a fundamental extracellular Asn-Pro-Ala motif (NPA) of the AQP2 protein, inhibiting its function. Its pathogenicity has been confirmed by in silico predictions and is in line with comparable alterations to the intracellular NPA motif of the AQP2 protein., (Copyright by Società Italiana di Nefrologia SIN, Rome, Italy.)

Aquaporins have been shown to be regulated by phosphorylation of serine residues, but the possible role of tyrosine residues phosphorylation has not been evaluated. Changes in the localization of aquaporin 2 (AQP2) in the queen endometrium have been related to serum progesterone levels. The aim of this study was to determine whether these AQP2-localization changes are mediated by variations in its tyrosine phosphorylation levels. Twelve queens were included in the study and divided into (a) non-macroscopically pregnant with low levels of progesterone; (b) non-macroscopically pregnant with high levels of progesterone; (c) 30 days of pregnancy; and (d) 60 days of pregnancy. Samples from endometrium and placental transference zone were obtained, immunoprecipitated and analysed by immunoblotting to determine the abundance of AQP2 and its relative levels of tyrosine phosphorylation. No significant differences in the tyrosine phosphorylation levels of immunoprecipated-AQP2 were observed between groups. We can thus conclude that changes in the localization of AQP2 in the queen endometrium are not modulated by tyrosine phosphorylation., (© 2020 Blackwell Verlag GmbH.)

Arginine-vasopressin (AVP) facilitates water reabsorption in renal collecting duct principal cells through regulation of the water channel aquaporin-2 (AQP2). The hormone binds to vasopressin V2 receptors (V2R) on the surface of the cells and stimulates cAMP synthesis. The cAMP activates protein kinase A (PKA), which initiates signaling that causes an accumulation of AQP2 in the plasma membrane of the cells facilitating water reabsorption from primary urine and fine-tuning of body water homeostasis. AVP-mediated PKA activation also causes an increase in the AQP2 protein abundance through a mechanism that involves dephosphorylation of AQP2 at serine 261 and a decrease in its poly-ubiquitination. However, the signaling downstream of PKA that controls the localization and abundance of AQP2 is incompletely understood. We carried out an siRNA screen targeting 719 kinase-related genes, representing the majority of the kinases of the human genome and analyzed the effect of the knockdown on AQP2 by high-content imaging and biochemical approaches. The screening identified 13 hits whose knockdown inhibited the AQP2 accumulation in the plasma membrane. Amongst the candidates was the so far hardly characterized cyclin-dependent kinase 18 (CDK18). Our further analysis revealed a hitherto unrecognized signalosome comprising CDK18, an E3 ubiquitin ligase, STUB1 (CHIP), PKA and AQP2 that controls the localization and abundance of AQP2. CDK18 controls AQP2 through phosphorylation at serine 261 and STUB1-mediated ubiquitination. STUB1 functions as an A-kinase anchoring protein (AKAP) tethering PKA to the protein complex and bridging AQP2 and CDK18. The modulation of the protein complex may lead to novel concepts for the treatment of disorders which are caused or are associated with dysregulated AQP2 and for which a satisfactory treatment is not available, e.g., hyponatremia, liver cirrhosis, diabetes insipidus, ADPKD or heart failure., Competing Interests: The authors declare no conflict of interest.