Search

Aldosterone is responsible for maintaining volume and potassium homeostasis. Although high salt consumption should suppress aldosterone production, individuals with hyperaldosteronism lose this regulation, leading to a state of high aldosterone despite dietary sodium consumption. The present study examines the effects of elevated aldosterone, with or without high salt consumption, on the expression of key Na+ transporters and remodelling in the distal nephron. Epithelial sodium channel (ENaC) α‐subunit expression was increased with aldosterone regardless of Na+ intake. However, ENaC β‐ and γ‐subunits unexpectedly increased at both a transcript and protein level with aldosterone when high salt was present. Expression of total and phosphorylated Na+Cl− cotransporter (NCC) significantly increased with aldosterone, in association with decreased blood [K+], but the addition of high salt markedly attenuated the aldosterone‐dependent NCC increase, despite equally severe hypokalaemia. We hypothesized this was a result of differences in distal convoluted tubule length when salt was given with aldosterone. Imaging and measurement of the entire pNCC‐positive tubule revealed that aldosterone alone caused a shortening of this segment, although the tubule had a larger cross‐sectional diameter. This was not true when salt was given with aldosterone because the combination was associated with a lengthening of the tubule in addition to increased diameter, suggesting that differences in the pNCC‐positive area are not responsible for differences in NCC expression. Together, our results suggest the actions of aldosterone, and the subsequent changes related to hypokalaemia, are altered in the presence of high dietary Na+. Key points: Aldosterone regulates volume and potassium homeostasis through effects on transporters in the kidney; its production can be dysregulated, preventing its suppression by high dietary sodium intake.Here, we examined how chronic high sodium consumption affects aldosterone's regulation of sodium transporters in the distal nephron.Our results suggest that high sodium consumption with aldosterone is associated with increased expression of all three epithelial sodium channel subunits, rather than just the alpha subunit.Aldosterone and its associated decrease in blood [K+] lead to an increased expression of Na‐Cl cotransporter (NCC); the addition of high sodium consumption with aldosterone partially attenuates this NCC expression, despite similarly low blood [K+].Upstream kinase regulators and tubule remodelling do not explain these results. [ABSTRACT FROM AUTHOR]

Yinzheng Li,1 Shulin Ma,1 Zheng Wang,1 Mengxia Shi,1 Rui Zeng,1– 4,* Ying Yao1,5,* 1Department of Nephrology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430030, People’s Republic of China; 2Key Laboratory of Organ Transplantation, Ministry of Education, Wuhan, 430030, People’s Republic of China; 3NHC Key Laboratory of Organ Transplantation, Wuhan, 430030, People’s Republic of China; 4Key Laboratory of Organ Transplantation, Chinese Academy of Medical Sciences, Wuhan, 430030, People’s Republic of China; 5Department of Nutrition, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430030, People’s Republic of China*These authors contributed equally to this workCorrespondence: Ying Yao; Rui Zeng, Department of Nephrology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430030, People’s Republic of China, Tel +86 13720379867 ; +86 15002726366, Email yaoyingkk@126.com; zengrui@tjh.tjmu.edu.cnPurpose: The distal nephron of kidney plays a pivotal role in advancing acute kidney injury (AKI). Understanding the role of distal nephrons in AKI and identifying markers of injured distal nephrons are critical to comprehending the mechanism of renal injury and identifying novel therapeutic targets.Methods: We analyzed single-cell RNA sequencing (scRNA-seq) data from mice with AKI induced by ischemia-reperfusion (IR), unilateral ureteral obstruction (UUO), cisplatin (CP), sodium oxalate (SO) and lipopolysaccharide (LPS). Additionally, we analyzed renal transcriptomics samples for AKI. Subsequently, we validated the effectiveness of targeting the biomarker Gclc in vitro and in vivo through metabolomics and immunofluorescence.Results: The LOH-Inj and DCT-Inj subtypes were identified through scRNA-seq. Compared to normal distal nephrons, the injured distal nephrons exhibited higher levels of ferroptosis, pro-inflammation, and fibrosis. The expression of ferroptosis-related gene Gclc were high in various AKI models. Furthermore, Gclc was exclusively expressed in the distal nephron and upregulated in the injury subtype. To confirm our findings, we suppressed GCLC expression in the kidneys, resulting to aggravated IR-induced AKI. Inhibition of Gclc promoted damage to primarily renal tubular epithelial cells by promoting inflammatory infiltration, inhibiting glutathione metabolism and exacerbating oxidative stress.Conclusion: Our research findings suggest that Gclc is a potential marker for injured distal nephron.Keywords: Gclc, distal nephron, acute kidney injury, single-cell sequencing

Benjamin T Enslow1, James D Stockand1, Jonathan M Berman2 1UT Health, San Antonio, TX, USA; 2New York Institute of Technology College of Osteopathic Medicine at Arkansas State University, Jonesboro, AR, USACorrspondence: Jonathan M BermanNew York Institute of Technology at Arkansas State University, Jonesboro, AR 72467, USATel +1 870 680 8896Email jberma03@nyit.eduAbstract: Liddle’s syndrome is a genetic disorder characterized by hypertension with hypokalemic metabolic alkalosis, hyporeninemia and suppressed aldosterone secretion that often appears early in life. It results from inappropriately elevated sodium reabsorption in the distal nephron. Liddle’s syndrome is caused by mutations to subunits of the Epithelial Sodium Channel (ENaC). Among other mechanisms, such mutations typically prevent ubiquitination of these subunits, slowing the rate at which they are internalized from the membrane, resulting in an elevation of channel activity. A minority of Liddle’s syndrome mutations, though, result in a complementary effect that also elevates activity by increasing the probability that ENaC channels within the membrane are open. Potassium-sparing diuretics such as amiloride and triamterene reduce ENaC activity, and in combination with a reduced sodium diet can restore normotension and electrolyte imbalance in Liddle’s syndrome patients and animal models. Liddle’s syndrome can be diagnosed clinically by phenotype and confirmed through genetic testing. This review examines the clinical features of Liddle’s syndrome, the differential diagnosis of Liddle’s syndrome and differentiation from other genetic diseases with similar phenotype, and what is currently known about the population-level prevalence of Liddle’s syndrome. This review gives special focus to the molecular mechanisms of Liddle’s syndrome.Keywords: ENaC, Liddle’s syndrome, hypertension, blood pressure, distal nephron

With no lysine kinase-4 (WNK4) belongs to a serine-threonine kinase family characterized by the atypical positioning of its catalytic lysine. Despite the fact that WNK4 has been found in many tissues, the majority of its study has revolved around its function in the kidney, specifically as a positive regulator of the thiazide-sensitive NaCl cotransporter (NCC) in the distal convoluted tubule of the nephron. This is explained by the description of gain-of-function mutations in the gene encoding WNK4 that causes familial hyperkalemic hypertension. This disease is mainly driven by increased downstream activation of the Ste20/SPS1-related proline-alanine-rich kinase/oxidative stress responsive kinase-1-NCC pathway, which increases salt reabsorption in the distal convoluted tubule and indirectly impairs renal K+ secretion. Here, we review the large volume of information that has accumulated about different aspects of WNK4 function. We first review the knowledge on WNK4 structure and enumerate the functional domains and motifs that have been characterized. Then, we discuss WNK4 physiological functions based on the information obtained from in vitro studies and from a diverse set of genetically modified mouse models with altered WNK4 function. We then review in vitro and in vivo evidence on the different levels of regulation of WNK4. Finally, we go through the evidence that has suggested how different physiological conditions act through WNK4 to modulate NCC activity. [ABSTRACT FROM AUTHOR]

Summary: Magnesium (Mg2+) homeostasis depends on active transcellular Mg2+ reuptake from urine in distal convoluted tubules (DCTs) via the Mg2+ channel TRPM6, whose activity has been proposed to be regulated by EGF. Calcium (Ca2+) homeostasis depends on paracellular reabsorption in the thick ascending limbs of Henle (TALs). KCTD1 promotes terminal differentiation of TALs/DCTs, but how its deficiency affects urinary Mg2+ and Ca2+ reabsorption is unknown. Here, this study shows that DCT1-specific KCTD1 inactivation leads to hypomagnesemia despite normal TRPM6 levels because of reduced levels of the sodium chloride co-transporter NCC, whereas Mg2+ homeostasis does not depend on EGF. Moreover, KCTD1 deficiency impairs paracellular urinary Ca2+ and Mg2+ reabsorption in TALs because of reduced NKCC2/claudin-16/-19 and increased claudin-14 expression, leading to hypocalcemia and consequently to secondary hyperparathyroidism and progressive metabolic bone disease. Thus, KCTD1 regulates urinary reabsorption of Mg2+ and Ca2+ by inducing expression of NCC in DCTs and NKCC2/claudin-16/-19 in TALs.

The discovery of new genetic mutations that cause hypertension has illuminated previously unrecognized physiological pathways. One such regulatory pathway was identified when mutations in with no lysine kinase (WNK)4, Kelch-like 3 (KLHL3), and cullin 3 (CUL3) were shown to cause the disease familial hyperkalemic hypertension (FHHt). Mutations in all three genes upregulate the NaCl cotransporter (NCC) due to an impaired ability to degrade WNK protein through the cullin-RING-ligase (CRL) ubiquitin-proteasome system. The CUL3 FHHt mutations cause the most severe phenotype, yet the precise mechanism by which these mutations cause the disease has not been established and current proposed models are controversial. New data have identified a possible novel mechanism involving dysregulation of CUL3 activity by the COP9 signalosome (CSN). The CSN interaction with mutant CUL3 is diminished, causing hyperneddylation of the CRL. Recent work has shown that direct renal CSN impairment mimics some aspects of the CUL3 mutation, including lower KLHL3 abundance and activation of the WNK-NCC pathway. Furthermore, in vitro and in vivo studies of CSN inhibition have shown selective degradation of CRL substrate adaptors via auto-ubiquitination, allowing substrate accumulation. In this review, we will focus on recent research that highlights the role of the CSN role in CUL3 mutations that cause FHHt. We will also highlight how these results inform other recent studies of CSN dysfunction. [ABSTRACT FROM AUTHOR]

Hypertension is common in the general population. Management of hypertensive patients at risk of hyperkalemia is challenging due to potential life-threatening complications such as cardiac arrest. Chronic hyperkalemia is often associated with impaired renal ability to excrete excessive potassium ions (K+). This may refer to chronic kidney disease or certain pharmacological interventions, including broadly used renin–angiotensin–aldosterone system and calcineurin inhibitors. Understanding the intrinsic mechanisms permitting kidney adaptations to hyperkalemia is critical for choosing therapeutic strategies. Valuable insights were obtained from the analysis of familial hyperkalemic hypertension (FHHt) syndrome, which became a classic model for coincidence of high blood pressure and hyperkalemia. FHHt can be caused by mutations in several genes, all of them resulting in excessive activity of with-no-lysine kinases (WNKs) in the distal nephron of the kidney. WNKs have been increasingly recognized as key signalling enzymes in the regulation of renal sodium ions (Na+) and K+ handling, enabling adaptive responses to systemic shifts of potassium homoeostasis consequent to variations in dietary potassium intake or disease. The WNK signalling pathway recruits a complex protein network mediating catalytic and non-catalytic effects of distinct WNK isoforms on relevant Na+- or K+-transporting proteins. In this review article, we summarize recent progress in understanding WNK signalling. An update of available models for renal adaptation to hyperkalemic conditions is presented. Consequences for blood pressure regulation are discussed. Pharmacological targeting of WNKs or their substrates offers promising options to manage hypertension while preventing hyperkalemia. [ABSTRACT FROM AUTHOR]

The temporal response of changes in renal sodium reabsorption during increased renal sympathetic nerve activity has not been investigated. Central hypovolemia by application of lower-body negative-pressure (LBNP) elicits baroreceptor mediated sympathetic reflexes to maintain arterial blood pressure. We hypothesized, that during 90 min LBNP, the renal sodium retention would increase rapidly, remain increased during intervention, and return to baseline immediately after end of intervention. 30 young, healthy, sodium loaded, non-obese males were exposed to −15 mmHg LBNP, −30 mmHg LBNP, −15 mmHg LBNP + renin blockade or time-control (0 mmHg LBNP) for 90 min. Urine was collected every 15 min during 90 min of intervention and 60 min of recovery to identify a possible relation between time of intervention and renal response. All intervention groups exhibited a comparable reduction in distal sodium excretion at the end of the intervention (P = 0.46 between groups; −15 mmHg: −3.1 ± 0.9 %, −30 mmHg: −2.9 ± 0.6 %, −15 mmHg + aslikiren: −1.8 ± 0.6 %). −15 mmHg+Aliskiren resulted in a slower onset, but all groups exhibited a continued reduction in sodium excretion after 1 h of recovery despite return to baseline of renin, aldosterone, diuresis and cardiovascular parameters. Sympathetic stimulation for 90 min via LBNP at −30 mmHg LBNP compared to −15 mmHg did not result in a greater response in fractional Na+ excretion, suggesting that additional baroreceptor unloading did not cause further increases in renal sodium reabsorption. Changes in distal Na+ excretion were linear with respect to time (dose) of intervention, but seem to exhibit a saturation-like effect at a level around 4 %. The lack of recovery after 1 h is also a new finding that warrants further investigation. [ABSTRACT FROM AUTHOR]

In the renal collecting ducts, chloride reabsorption occurs through both transcellular and paracellular pathways. Recent literature highlights a functional interplay between both pathways. We recently showed that in polarized inner medullary collecting duct cells, expression of the basolateral kidney anion exchanger 1 (kAE1) results in a decreased transepithelial electrical resistance (TEER), in a claudin-4 dependent pathway. Claudin-4 is a paracellular sodium blocker and chloride pore. Here, we show that kAE1 expression in mouse inner medullary collecting duct cells triggers WNK4, SPAK and claudin-4 phosphorylation. Expression of a functionally dead kAE1 E681Q mutant has no effect on phosphorylation of these proteins. Expression of a catalytically inactive WNK4 D321A or chloride-insensitive WNK4 L319F mutant abolishes kAE1 effect on TEER, supporting a contribution of WNK4 to the process. We propose that variations of the cytosolic pH and chloride concentration upon kAE1 expression alter WNK4 kinase activity and tight junction properties., Competing Interests: The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Emmanuelle Cordat reports financial support was provided by 10.13039/501100000024Canadian Institutes of Health Research. Rawad Lashhab reports financial support was provided by 10.13039/501100000038Natural Sciences and Engineering Research Council of Canada. Maria Chavez-Canales reports financial support was provided by 10.13039/501100003141CONACyT Mexicon. Maria Chavez-Canales reports financial support was provided by PAPIIT UNAM., (© 2023 Published by Elsevier Ltd.)

The kidney is essential for systemic calcium homeostasis. Urinary calcium excretion can be viewed as an integrative renal response to endocrine and local stimuli. The extracellular calcium-sensing receptor (CaSR) elicits a number of adaptive reactions to increased plasma Ca2+ levels including the control of parathyroid hormone release and regulation of the renal calcium handling. Calcium reabsorption in the distal nephron of the kidney is functionally coupled to sodium transport. Apart from Ca2+ transport systems, CaSR signaling affects relevant distal Na+-(K+)-2Cl− cotransporters, NKCC2 and NCC. NKCC2 and NCC are activated by a kinase cascade comprising with-no-lysine [K] kinases (WNKs) and two homologous Ste20-related kinases, SPAK and OSR1. Gain-of-function mutations within the WNK-SPAK/OSR1-NKCC2/NCC pathway lead to renal salt retention and hypertension, whereas loss-of-function mutations have been associated with salt-losing tubulopathies such as Bartter or Gitelman syndromes. A Bartter-like syndrome has been also described in patients carrying gain-of-function mutations in the CaSR gene. Recent work suggested that CaSR signals via the WNK-SPAK/OSR1 cascade to modulate salt reabsorption along the distal nephron. The review presented here summarizes the latest progress in understanding of functional interactions between CaSR and WNKs and their potential impact on the renal salt handling and blood pressure.

Autosomal dominant mutations in cullin-3 (Cul3) cause the most severe form of familial hyperkalemic hypertension (FHHt). Cul3 mutations cause skipping of exon 9, which results in an internal deletion of 57 amino acids from the CUL3 protein (CUL3-Δ9). The precise mechanism by which this altered form of CUL3 causes FHHt is controversial. CUL3 is a member of the cullin-RING ubiquitin ligase family that mediates ubiquitination and thus degradation of cellular proteins, including with-no-lysine [K] kinases (WNKs). In CUL3-Δ9-mediated FHHt, proteasomal degradation of WNKs is abrogated, leading to overactivation of the WNK targets sterile 20/SPS-1 related proline/alanine-rich kinase and oxidative stressresponse kinase-1, which directly phosphorylate and activate the thiazide-sensitive Na+-Cl- cotransporter. Several groups have suggested different mechanisms by which CUL3-Δ9 causes FHHt. The majority of these are derived from in vitro data, but recently the Kurz group (Schumacher FR, Siew K, Zhang J, Johnson C, Wood N, Cleary SE, Al Maskari RS, Ferryman JT, Hardege I, Figg NL, Enchev R, Knebel A, O’Shaughnessy KM, Kurz T. EMBO Mol Med 7: 1285–1306, 2015) described the first mouse model of CUL3-Δ9-mediated FHHt. Analysis of this model suggested that CUL3-Δ9 is degraded in vivo, and thus Cul3 mutations cause FHHt by inducing haploinsufficiency. We recently directly tested this model but found that other dominant effects of CUL3-Δ9 must contribute to the development of FHHt. In this review, we focus on our current knowledge of CUL3-Δ9 action gained from in vitro and in vivo models that may help unravel this complex problem. [ABSTRACT FROM AUTHOR]

Key points Hypercalcaemia can occur under various pathological conditions, such as primary hyperparathyroidism, malignancy or granulomatosis, and it induces natriuresis and polyuria in various species via an unknown mechanism., A previous study demonstrated that hypercalcaemia induced by vitamin D in rats increased endothelin (ET)-1 expression in the distal nephron, which suggests the involvement of the ET system in hypercalcaemia-induced effects., In the present study, we demonstrate that, during vitamin D-induced hypercalcaemia, the activation of ET system by increased ET-1 is responsible for natriuresis but not for polyuria., Vitamin D-treated hypercalcaemic mice showed a blunted response to amiloride, suggesting that epithelial sodium channel function is inhibited., We have identified an original pathway that specifically mediates the effects of vitamin D-induced hypercalcaemia on sodium handling in the distal nephron without affecting water handling., Abstract Acute hypercalcaemia increases urinary sodium and water excretion; however, the underlying molecular mechanism remains unclear. Because vitamin D-induced hypercalcaemia increases the renal expression of endothelin (ET)-1, we hypothesized that ET-1 mediates the effects of hypercalcaemia on renal sodium and water handling. Hypercalcaemia was induced in 8-week-old, parathyroid hormone-supplemented, male mice by oral administration of dihydrotachysterol (DHT) for 3 days. DHT-treated mice became hypercalcaemic and displayed increased urinary water and sodium excretion compared to controls. mRNA levels of ET-1 and the transcription factors CCAAT-enhancer binding protein β and δ were specifically increased in the distal convoluted tubule and downstream segments in DHT-treated mice. To examine the role of the ET system in hypercalcaemia-induced natriuresis and polyuria, mice were treated with the ET-1 receptor antagonist macitentan, with or without DHT. Mice treated with both macitentan and DHT displayed hypercalcaemia and polyuria similar to that in mice treated with DHT alone; however, no increase in urinary sodium excretion was observed. To identify the affected sodium transport mechanism, we assessed the response to various diuretics in control and DHT-treated hypercalcaemic mice. Amiloride, an inhibitor of the epithelial sodium channel (ENaC), increased sodium excretion to a lesser extent in DHT-treated mice compared to control mice. Mice treated with either macitentan+DHT or macitentan alone had a similar response to amiloride. In summary, vitamin D-induced hypercalcaemia increases the renal production of ET-1 and decreases ENaC activity, which is probably responsible for the rise in urinary sodium excretion but not for polyuria. [ABSTRACT FROM AUTHOR]

Kelch-like 3 (KLHL3) is a component of an E3 ubiquitin ligase complex that regulates blood pressure by targeting With-No-Lysine (WNK) kinases for degradation. Mutations in KLHL3 cause constitutively increased renal salt reabsorption and impaired K + secretion, resulting in hypertension and hyperkalemia. Although clinical studies have shown that dietary K + intake affects blood pressure, the mechanisms have been obscure. In this study, we demonstrate that the KLHL3 ubiquitin ligase complex is involved in the low-K + -mediated activation of Na-Cl cotransporter (NCC) in the kidney. In the distal convoluted tubules of mice eating a low-K + diet, we found increased KLHL3 phosphorylation at S433 (KLHL3 S433-P ), a modification that impairs WNK binding, and also reduced total KLHL3 levels. These changes are accompanied by the accumulation of the target substrate WNK4, and activation of the downstream kinases SPAK (STE20/SPS1-related proline-alanine-rich protein kinase) and OSR1 (oxidative stress-responsive 1), resulting in NCC phosphorylation and its accumulation at the plasma membrane. Increased phosphorylation of S433 was explained by increased levels of active, phosphorylated protein kinase C (but not protein kinase A), which directly phosphorylates S433. Moreover, in HEK cells expressing KLHL3 and WNK4, we showed that the activation of protein kinase C by phorbol 12-myristate 13-acetate induces KLHL3 S433-P and increases WNK4 levels by abrogating its ubiquitination. These data demonstrate the role of KLHL3 in low-K + -mediated induction of NCC; this physiologic adaptation reduces distal electrogenic Na + reabsorption, preventing further renal K + loss but promoting increased blood pressure. [ABSTRACT FROM AUTHOR]

The distal nephron is a heterogeneous part of the nephron composed by six different cell types, forming the epithelium of the distal convoluted (DCT), connecting, and collecting duct. To dissect the function of these cells, knockout models specific for their unique cell marker have been created. However, since this part of the nephron develops at the border between the ureteric bud and the metanephric mesenchyme, the specificity of the single cell markers has been recently questioned. Here, by mapping the fate of the aquaporin 2 (AQP2) and Na+-Cl- cotransporter (NCC)-positive cells using transgenic mouse lines expressing the yellow fluorescent protein fluorescent marker, we showed that the origin of the distal nephron is extremely composite. Indeed, AQP2-expressing precursor results give rise not only to the principal cells, but also to some of the A- and B-type intercalated cells and even to cells of the DCT. On the other hand, some principal cells and B-type intercalated cells can develop from NCC-expressing precursors. In conclusion, these results demonstrate that the origin of different cell types in the distal nephron is not as clearly defined as originally thought. Importantly, they highlight the fact that knocking out a gene encoding for a selective functional marker in the adult does not guarantee cell specificity during the overall kidney development. Tools allowing not only cell-specific but also time-controlled recombination will be useful in this sense. [ABSTRACT FROM AUTHOR]

Potassium Kir4.1/5.1 channels are abundantly expressed at the basolateral membrane of principal cells in the cortical collecting duct (CCD), where they are thought to modulate transport rates by controlling transepithelial voltage. Insulin and insulin-like growth factor-1 (IGF-1) stimulate apically localized epithelial sodium channels (ENaC) to augment sodium reabsorption in the CCD. However, little is known about their actions on potassium channels localized at the basolateral membrane. In this study, we implemented patch-clamp analysis in freshly isolated murine CCD to assess the effect of these hormones on Kir4.1/5.1 at both single channel and cellular levels. We demonstrated that K+-selective conductance via Kir4.1/ 5.1 is the major contributor to the macroscopic current recorded from the basolateral side in principal cells. Acute treatment with 10 μM amiloride (ENaC blocker), 100 nM tertiapin-Q (TPNQ; ROMK inhibitor), and 100 μM ouabain (Na+-K+-ATPase blocker) failed to produce a measurable effect on the macroscopic current. In contrast, Kir4.1 inhibitor nortriptyline (100 μM), but not fluoxetine (100 μM), virtually abolished whole cell K+-selective conductance. Insulin (100 nM) markedly increased the open probability of Kir4.1/ 5.1 and nortriptyline-sensitive whole cell current, leading to significant hyperpolarization of the basolateral membrane. Inhibition of the phosphatidylinositol 3-kinase cascade with LY294002 (20 μM) abolished action of insulin on Kir4.1/5.1. IGF-1 had similar stimulatory actions on Kir4.1/5.1-mediated conductance only when applied at a higher (500 nM) concentration and was ineffective at 100 nM. We concluded that both insulin and, to a lesser extent, IGF-1 activate Kir4.1/5.1 channel activity and open probability to hyperpolarize the basolateral membrane, thereby facilitating Na+ reabsorption in the CCD. [ABSTRACT FROM AUTHOR]

Gordon Syndrome (GS) is a rare familial hypertension syndrome with a characteristic hyperkalaemia which distinguishes it from other syndromic forms of hypertension that typically cause hypokalaemia. Patients with GS respond to aggressive salt-restriction or relatively small doses of thiazide diuretics, which suggests that activation of the thiazide-sensitive Na/Cl cotransporter (NCC) in the distal nephron is to blame. However, the mechanism has proved to be complex. In 2001, mutations in genes encoding two serine/threonine kinases, WNK1 and WNK4, were identified as causing GS. However, it took several years to appreciate that these kinases operated in a cascade with downstream serine/threonine kinases (SPAK and OSR1) actually phosphorylating and activating NCC and the closely related cotransporters NKCC1 and NKCC2. The hyperkalaemia in GS arises from an independent action of WNK1/WNK4 to reduce cell-surface expression of ROMK, the secretory K-channel in the collecting ducts. However, mutations in WNK1/4 are present in a small minority of GS families, and further genes have emerged ( CUL3 and KLHL3) that code for Cullin-3 (a scaffold protein in an ubiquitin-E3 ligase) and an adaptor protein, Kelch3, respectively. These new players regulate the ubiquitination and proteasomal degradation of WNK kinases, thereby adding to the complex picture we now have of NCC regulation in the distal nephron. [ABSTRACT FROM AUTHOR]

These studies test whether three disease-causing mutations in genes (SCNN1A and SCNN1G) encoding subunits of the epithelial Na+ channel, ENaC, affect the biophysical and gating properties of this important renal ion channel. The S562P missense mutation in αENaC and the K106_S108delinsN mutation in cENaC are associated with pseudohypoaldosteronism type 1 (PHA1). The N530S missense mutation in cENaC causes Liddle's syndrome. Incorporation of S562P into aENaC and K106_S108N into γENaC resulted in significant decreases in macroscopic ENaC currents. Conversely, incorporation of N530S into γENaC increased macroscopic ENaC current. The S562P substitution resulted in a nonfunctional channel. The K106_S108N mutation produced a functional channel having a normal macroscopic current-voltage relation, there was a slight but significant decrease in unitary conductance and a marked decrease in single-channel open probability. The N530S substitution increased single-channel open probability having no effect on the macroscopic current-voltage relation or unitary conductance of the channel. These findings are consistent with mutation of residues at 562 in αENaC and 530 in cENaC, and a 30 splice site in SCNN1G (318-1 G→A; K106_108SdelinsN) resulting in aberrant ENaC activity due to changes in the biophysical and gating properties of the channel. Such changes likely contribute to the cellular mechanism underpinning the PHA1 and Liddle's syndrome caused by these mutations in ENaC subunits. [ABSTRACT FROM AUTHOR]

The aldosterone-sensitive distal nephron (ASDN) exhibits axial heterogeneity in structure and function from the distal convoluted tubule to the medullary collecting duct. Ion and water transport is primarily divided between the cortex and medulla of the ASDN, respectively. Transcellular transport in this segment is highly regulated in health and disease and is integrated across different cell types. We currently lack an inexpensive, high-yield, and tractable technique to harvest and culture cells for the study of gene expression and physiological properties of mouse cortical ASDN. To address this need, we harvested tubules bound to Dolichos biflorus agglutinin lectin-coated magnetic beads from the kidney cortex and characterized these cell preparations. We determined that these cells are enriched for markers of distal convoluted tubule, connecting tubule, and cortical collecting duct, including principal and intercalated cells. In primary culture, these cells develop polarized monolayers with high resistance (1,000-1,500 Ω * cm²) and maintain expression and activity of key channels. These cells demonstrate an amiloride-sensitive short-circuit current that can be enhanced with aldosterone and maintain measurable potassium and anion secretion. Our method can be easily adopted to study the biology of the ASDN and to investigate phenotypic differences between wild-type and transgenic mouse models. [ABSTRACT FROM AUTHOR]

The kidney continuously adapts daily renal excretion of NaCl to match dietary intakes in order to maintain the NaCl content of the body, and keep vascular volume constant. Any situation that leads to NaCl retention favors a rise in blood pressure. The aldosterone-sensitive distal nephron, which contains two main types of cells, principal (PC) and intercalated (IC) cells, is an important site for the final regulation of urinary Na excretion. Research over the past 20 years established a paradigm in which PCs are the exclusive site of Na absorption while ICs are solely dedicated to acid-base transport. Recent studies have revealed the unexpected importance of ICs for NaCl reabsorption. Here, we review the mechanisms of Na and Cl transport in the aldosterone-sensitive distal nephron, with emphasis on the role of ICs in maintaining NaCl balance and normal blood pressure. [ABSTRACT FROM AUTHOR]

Despite similar stimulatory actions on the epithelial sodium channel (ENaC)-mediated sodium reabsorption in the distal tubule, insulin promotes kaliuresis, whereas insulin-like growth factor-1 (IGF-1) causes a reduction in urinary potassium levels. The factors contributing to this phenomenon remain elusive. Electrogenic distal nephron ENaC-mediated Na+ transport establishes driving force for Cl- reabsorption and K+ secretion. Using patch-clamp electrophysiology, we document that a Cl- channel is highly abundant on the basolateral plasma membrane of intercalated cells in freshly isolated mouse cortical collecting duct (CCD) cells. The channel has characteristics attributable to the ClC-K2: slow gating kinetics, conductance ~10 pS, voltage independence, Cl->NO3- anion selectivity, and inhibition/ activation by low/high pH, respectively. IGF-1 (100 and 500 nM) acutely stimulates ClC-K2 activity in a reversible manner. Inhibition of PI3-kinase (PI3-K) with LY294002 (20 μM) abrogates activation of ClC-K2 by IGF-1. Interestingly, insulin (100 nM) reversibly decreases ClC-K2 activity in CCD cells. This inhibitory action is independent of PI3-K and is mediated by stimulation of a mitogenactivated protein kinase-dependent cascade. We propose that IGF-1, by stimulating ClC-K2 channels, promotes net Na+ and Cl- reabsorption, thus reducing driving force for potassium secretion by the CCD. In contrast, inhibition of ClC-K2 by insulin favors coupling of Na+ reabsorption with K+ secretion at the apical membrane contributing to kaliuresis. [ABSTRACT FROM AUTHOR]

Sodium absorption by the distal part of the nephron, i.e., the distal convoluted tubule, the connecting tubule, and the collecting duct, plays a major role in the control of homeostasis by the kidney. In this part of the nephron, sodium transport can either be electroneutral or electrogenic. The study of electrogenic Na absorption, which is mediated by the epithelial sodium channel (ENaC), has been the focus of considerable interest because of its implication in sodium, potassium, and acid-base homeostasis. However, recent studies have highlighted the crucial role played by electroneutral NaCl absorption in the regulation of the body content of sodium chloride, which in turn controls extracellular fluid volume and blood pressure. Here, we review the identification and characterization of the NaCl cotransporter (NCC), the molecule accounting for the main part of electroneutral NaCl absorption in the distal nephron, and its regulators. We also discuss recent work describing the identification of a novel 'NCC-like' transport system mediated by pendrin and the sodium-driven chloride/bicarbonate exchanger (NDCBE) in the β-intercalated cells of the collecting system. [ABSTRACT FROM AUTHOR]

It is recognized that dopamine promotes natriuresis by inhibiting multiple transporting systems in the proximal tubule. In contrast, less is known about the molecular targets of dopamine actions on water-electrolyte transport in the cortical collecting duct (CCD). Epithelial cells in the CCD are exposed to dopamine, which is synthesized locally or secreted from sympathetic nerve endings. Basolateral K+ channels in the distal renal tubule are critical for K+ recycling and controlling basolateral membrane potential to establish the driving force for Na+ reabsorption. Here, we demonstrate that Kir4.1 and Kir5.1 are highly expressed in the mouse kidney cortex and are localized to the basolateral membrane of the CCD. Using patch-clamp electrophysiology in freshly isolated CCDs, we detected highly abundant 40-pS and scarce 20-pS single channel conductances, most likely representing Kir4.1/5.1 and Kir4.1 channels, respectively. Dopamine reversibly decreased the open probability of both channels, with a relatively greater action on the Kir4.1/5.1 heterodimer. This effect was mediated by D2-like but not D1-like dopamine receptors. PKC blockade abolished the inhibition of basolateral K+ channels by dopamine. Importantly, dopamine significantly decreased the amplitude of Kir4.1/5.1 and Kir4.1 unitary currents. Consistently, dopamine induced an acute depolarization of basolateral membrane potential, as directly monitored using currentclamp mode in isolated CCDs. Therefore, we demonstrate that dopamine inhibits basolateral Kir4.1/5.1 and Kir4.1 channels in CCD cells via stimulation of D2-like receptors and subsequently PKC. This leads to depolarization of the basolateral membrane and a decreased driving force for Na+ reabsorption in the distal renal tubule. [ABSTRACT FROM AUTHOR]

Ca-activated K channels (BK), which are stimulated by high distal nephron flow, are utilized during high-K conditions to remove excess K. Because BK predominantly reside with BK-β4 in acid/base-transporting intercalated cells (IC), we determined whether BK-β4 knockout mice (β4KO) exhibit deficient K excretion when consuming a high-K alkaline diet (HK-alk) vs. high-K chloride diet (HK-Cl). When wild type (WT) were placed on HK-alk, but not HK-Cl, renal BK-β4 expression increased (Western blot). When WT and β4KO were placed on HK-Cl, plasma K concentration ([K]) was elevated compared with control K diets; however, K excretion was not different between WT and β4KO. When HK-alk was consumed, the plasma [K] was lower and K clearance was greater in WT compared with β4KO. The urine was alkaline in mice on HK-alk; however, urinary pH was not different between WT and β4KO. Immunohistochemical analysis of pendrin and V-ATPase revealed the same increases in β-IC, comparing WT and β4KO on HK-alk. We found an amiloride-sensitive reduction in Na excretion in β4KO, compared with WT, on HK-alk, indicating enhanced Na reabsorption as a compensatory mechanism to secrete K. Treating mice with an alkaline, Na-deficient, high-K diet (LNaHK) to minimize Na reabsorption exaggerated the defective K handling of β4KO. When WT on LNaHK were given NH4Cl in the drinking water, K excretion was reduced to the magnitude of β4KO on LNaHK. These results show that WT, but not β4KO, efficiently excretes K on HK-alk but not on HK-Cl and suggest that BK-α/β4-mediated K secretion is promoted by bicarbonaturia. [ABSTRACT FROM AUTHOR]

Autosomal recessive distal renal tubular acidosis (dRTA) is a severe disorder of acid-base homeostasis, often accompanied by sensorineural deafness. We and others have previously shown that mutations in the tissue-restricted a4 and B1 subunits of the H+-ATPase underlie this syndrome. Here, we describe an Atp6v0a4 knockout mouse, which lacks the a4 subunit. Using β-galactosidase as a reporter for the null gene, developmental a4 expression was detected in developing bone, nose, eye, and skin, in addition to that expected in kidney and inner ear. By the time of weaning, Atp6v0a4-/- mice demonstrated severe metabolic acidosis, hypokalemia, and early nephrocalcinosis. Null mice were hypocitraturic, but hypercalciuria was absent. They were severely hearing-impaired, as shown by elevated auditory brainstem response thresholds and absent endocochlear potential. They died rapidly unless alkalinized. If they survived weaning with alkali supplementation, treatment could later be withdrawn, but -/- animals remained acidotic with alkaline urine. They also had an impaired sense of smell. Heterozygous animals were biochemically normal until acid-challenged, when they became more acidotic than +/+ animals. This mouse model recapitulates the loss of H+-ATPase function seen in human disease and can provide additional insights into dRTA and the physiology of the a4 subunit. [ABSTRACT FROM AUTHOR]

Background. The epithelial calcium channel (ECaC) (TRPV5) and the Cl−/HCO3− exchanger pendrin (SLC26A4) are expressed on the apical membrane of tubular cells in the distal nephron and play essential roles in calcium re-absorption and bicarbonate secretion, respectively, in the kidney. Methods. A combination of functional and molecular biology techniques were employed to examine the role of pendrin deletion in calcium excretion. Results. Here, we demonstrate that deletion of pendrin causes acidic urine [urine pH 4.9 in knockout (KO) versus 5.9 in wild-type (WT) mice, P < 0.03)] and downregulates the calcium-absorbing molecules ECaC and Na/Ca exchanger in the kidney, as shown by northern hybridization, immunoblot analysis and/or immunofluorescent labeling. These changes were associated with a ∼100% increase in 24-h urine calcium excretion in pendrin null mice. Subjecting the pendrin WT and KO mice to oral bicarbonate loading for 12 days increased the urine pH to ∼8 in both genotypes, normalized the expression of ECaC and Na/Ca exchanger and reduced the urine calcium excretion in pendrin-null mice to levels comparable to WT mice. Conclusions. We suggest that pendrin dysfunction should be suspected and investigated in humans with an otherwise unexplained acidic urine and hypercalciuria. [ABSTRACT FROM PUBLISHER]

Renin expression in principal cells of collecting ducts (CD) is upregulated in angiotensin II (ANG II)-dependent hypertensive rats; however, it remains unclear whether increased CD-derived renin undergoes tubular secretion. Accordingly, urinary levels of renin (uRen), angiotensinogen (uAGT), and ANG II (uANG II) were measured in chronic ANG II-infused Sprague-Dawley rats (80 ng/min for 14 days, n = 10) and sham-operated rats (n = 10). Systolic blood pressure increased in the ANG II rats by day 5 and continued to increase throughout the study (day 13; ANG II: 175 ± 10 vs. sham: 116 ± 2 mmHg; P < 0.05). ANG II infusion increased renal cortical and medullary ANG II levels (cortical ANG II: 606 ± 72 vs. 247 ± 43 fmol/g; P < 0.05; medullary ANG II: 2,066 ± 116 vs. 646 ± 36 fmol/g; P < 0.05). Although plasma renin activity (PRA) was suppressed in the ANG II-infused rats (0.3 ± 0.2 vs. 5.5 ± 1.8 ng ANG I·ml-1·h-1; P < 0.05), renin content in renal medulla was increased (12,605 ± 1,343 vs. 7,956 ± 765 ng ANG I·h-1·mg-1; P <0.05). Excretion of uAGT and uANG II increased in the ANG II rats [uAGT: 1,107 ± 106 vs. 60 ± 26 ng/day; P < 0.0001; uANG II: 3,813 ± 431 vs. 2,080 ± 361 fmol/day; P < 0.05]. By day 13, despite suppression of PRA, urinary prorenin content increased in ANG II rats [15.7 ± 3 vs. 2.6 ± 1 X 10-3 enzyme Units excreted (EUE)/day, P < 0.011 as was the excretion rate of renin (8.6 ± 2 X 10-6 EUE/day) compared with sham (2.8 ± 1 X 10-6 EUE/day; P < 0.05). Urinary renin and prorenin protein levels examined by Western blot were augmented ∼10-fold in the ANG IT-infused rats. Concomitant AT1 receptor blockade with candesartan prevented the increase. Thus, in ANG II-dependent hypertensive rats with marked PRA suppression, increased urinary levels of renin and prorenin reflect their augmented secretion by CD cells into the luminal fluid. The greater availability of renin and AGT in the urine reflects the capability for intratubular ANG II formation which stimulates sodium reabsorption in distal nephron segments. [ABSTRACT FROM AUTHOR]

Na+ is commonly designed as the culprit of salt-sensitive hypertension but several studies suggest that abnormal Cl- transport is in fact the triggering mechanism. This review focuses on the regulation of blood pressure (BP) by pendrin, an apical Cl-/HCO3- exchanger which mediates HCO3- secretion and transcellular Cl- transport in type B intercalated cells (B-ICs) of the distal nephron. Studies in mice showed that it is required not only for acid-base regulation but also for BP regulation as pendrin knock-out mice develop hypotension when submitted to NaCl restriction and are resistant to aldosterone-induced hypertension. Pendrin contributes to these processes by two mechanisms. First, pendrin-mediated Cl- transport is coupled with Na+ reabsorption by the Na+-dependent Cl-/HCO3- exchanger NDCBE to mediate NaCl reabsorption in B-ICs. Second, pendrin activity regulates Na+ reabsorption by the adjacent principal cells, possibly by interaction with the ATP-mediated paracrine signalling recently identified between ICs and principal cells. Interestingly, the water channel AQP5 was recently found to be expressed at the apical side of B-ICs, in the absence of a basolateral water channel, and pendrin and AQP5 membrane expressions are both inhibited by K+ depletion, suggesting that pendrin and AQP5 could cooperate to regulate cell volume, a potent stimulus of ATP release. Copyright © 2011 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

The Cl-/HCO3- exchanger pendrin (SLC26A4, PDS) and the thiazide-sensitive NaCl cotransporter NCC (SLC12A3) are expressed on the apical membranes of distal nephron segments and mediate salt absorption, with pendrin working in tandem with the epithelial Na channel (ENaC) and NCC working by itself. Pendrin is expressed on the apical membrane of intercalated cells in late distal convoluted tubule (DCT), connecting tubule (CNT) and the cortical collecting duct (CCD) whereas the thiazide-sensitive NaCl cotransporter NCC is primarily detected on the apical membrane of DCT cells. Recent studies indicate that pendrin expression is increased in kidneys of NCC knockout mice, raising the possibility that pendrin and NCC can compensate for loss of the other by increasing their expression and activity. Current investigations in our laboratories demonstrate that pendrin plays an important role in compensatory salt absorption in response to the loop diuretics and the thiazide derivatives. These studies further demonstrate that whereas single deletion of pendrin or NCC does not cause salt wasting in mutant mice under baseline conditions, double knockout of pendrin and NCC causes profound polyuria and polydipsia, along with salt wasting under basal conditions. As a result, animals develop significant dehydration. We propose that pharmacologic inhibition of pendrin and NCC can provide a novel and strong diuretic regimen for patients with fluid overload, including those with congestive heart failure, nephrotic syndrome or renal failure. Copyright © 2011 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

Increased flow in the distal nephron induces K secretion through the large-conductance, calcium-activated K channel (BK), which is primarily expressed in intercalated cells (IC). Since flow also increases ATP release from IC, we hypothesized that purinergic signaling has a role in shear stress (π; 10 dynes/cm²) -induced, BK-dependent, K efflux. We found that 10 μM ATP led to increased IC Ca concentration, which was significantly reduced in the presence of the P² receptor blocker suramin or calcium-free buffer. ATP also produced BK-dependent K efflux, and IC volume decrease. Suramin inhibited T-induced K efflux, suggesting that K efflux is at least partially dependent on purinergic signaling. BK-β4 small interfering (si) RNA, but not nontarget siRNA, decreased ATP secretion and both ATP-dependent and π-induced K efflux. Similarly, carbenoxolone (25 μM), which blocks connexins, putative AlP pathways, blocked T-induced K efflux and ATP secretion. Compared with BK-β4-/- mice, wild-type mice with high distal flows exhibited significantly more urinary ATP excretion. These data demonstrate coupled electrochemical efflux between K and ATP as part of the mechanism for π-induced ATP release in IC. [ABSTRACT FROM AUTHOR]

The fine regulation of Na+ and K+ transport takes place in the cortical distal nephron. It is well established that K+ secretion occurs through apical K+ channels: the ROMK and the Ca2+ - and voltage-dependent maxi-K. Previously, we identified the voltage-gated Kv1.3 channel in the inner medulla of the rat kidney (Escobar LI, Martínez-Téllez JC, Salas M, Castilla SA, Carrisoza R, Tapia D, Vázquez M, Bargas J, Bolívar JJ. Am J Physiol Cell Physiol 286: C965-C974, 2004). To examine the role of Kv1.3 in the renal regulation of K+ homeostasis, we characterized the effect of dietary K+ on the molecular and functional expression of this channel. We performed real-time-PCR and immunoblot assays in kidneys from rats fed a control (CK; 1.2% wt/wt) or high-K+ (HK; 10% wt/wt) diet for 5-15 days. Kv1.3 mRNA and protein expression did not change with HK in the whole kidney. However, dietary K+ loading provoked a change in the cellular distribution of Kv1.3 from the cytoplasm to apical membranes. Immunolocalization of Kv1.3 detected the channel exclusively in the intercalated cells. We investigated whether Kv1.3 mediated K+ transport in microperfused cortical collecting ducts (CCDs). The HK diet led to an increase in net K+ transport from 7.4 ± 1.1 (CK) to 11.4 ± 1.0 (HK) pmol·min-1mm-1. Luminal margatoxin, a specific blocker of Kv1.3, decreased net K+ secretion in HK CCDs to 6.0 ± 1.6 pmol·min-1mm-1. Our data provide the first evidence that Kv1.3 channels participate in K+ secretion and that apical membrane localization of Kv1.3 is enhanced in the intercalated cells by dietary K+ loading. [ABSTRACT FROM AUTHOR]

Apical release of ATP and UTP can activate P2Y2 receptors in the aldosterone-sensitive distal nephron (ASDN) and inhibit the open probability (Po) of the epithelial sodium channel (ENaC). Little is known, however, about the regulation and physiological relevance of this system. Patch-clamp studies in freshly isolated ASDN provide evidence that increased dietary Na+ intake in wild-type mice lowers ENaC Po, consistent with a contribution to Na+ homeostasis, and is associated with increased urinary concentrations of UTP and the ATP hydrolytic product, ADP. Genetic deletion of P2Y2 receptors in mice (P2Y2 -/-; littermates to wild-type mice) or inhibition of apical P2Yreceptor activation in wild-type mice prevents dietary Na+-induced lowering of ENaC Po. Although they lack suppression of ENaC Po by dietary NaCl, P2Y2 -/- mice do not exhibit NaCl-sensitive blood pressure, perhaps as a consequence of compensatory down-regulation of aldosterone levels. Consistent with this hypothesis, clamping mineralocorticoid activity at high levels unmasks greater ENaC activity and NaCl sensitivity of blood pressure in P2Y2-/- mice. The studies indicate a key role of the apical ATP/UTP-P2Y2-receptor system in the inhibition of ENaC Po in the ASDN in response to an increase in Na+ intake, thereby contributing to NaCl homeostasis and blood pressure regulation. [ABSTRACT FROM AUTHOR]

The renal late distal convoluted tubules and connecting tubules are sites for the fine regulation of Na+ and Ca+ reabsorption. The role of these segments in Na+ and K+ homeostasis is possibly underestimated, as the tubules are technically difficult to isolate in sufficient numbers and purity for functional analysis. To overcome these difficulties, we have developed a transgenic mouse model expressing enhanced green fluorescent protein in late distal convoluted tubules and connecting tubules. Enhanced green fluorescent protein expression was driven by the promoter for the transient receptor potential subfamily VT member 5. Confocal fluorescence microscopy allowed detection of enhanced green fluorescent protein in living, isolated late distal convoluted tubules and connecting tubules and in the initial cortical collecting ducts. Enhanced green fluorescent protein expression was validated by double- and triple-fluorescence immunolabeling with specific tubule markers. Freshly isolated late distal convoluted tubules and connecting tubules increased their intracellular Ca2+ levels in response to the V2 receptor-specific agonist deamino-Cys,D-Arg8-vasopressin (2 X 10-10 M) after 1 min of superfusion. In addition, both late distal convoluted tubules and connecting tubules displayed a concentration-dependent intracellular Ca2+ response to 1±,25-dihydroxyvitamin D3 (range 10-10 to 10-8 M). This suggests that 1±,25-dihydroxyvitamin D3 can act through a nongenomic signaling pathway in these tubules. In conclusion, the transgenic mouse model, expressing enhanced green fluorescent protein, is suitable for rapid isolation of viable late distal convoluted tubules, connecting tubules, and initial cortical collecting ducts and provides an ideal tool for a more exhaustive functional characterization of these segments. [ABSTRACT FROM AUTHOR]

The amiloride-sensitive epithelial sodium channel (ENaC) has previously been shown to be involved in the maintenance of body fluid volume and in Na+ absorption across the skin and urinary bladder in amphibians. However, the function and distribution of ENaC have not been clearly described in amphibian kidney. We therefore cloned the ENaC α-subunit cDNA from kidney of the marine toad, Bufo marinus. The ENaC mRNA and protein were abundantly expressed in the kidney and in the urinary bladder and ventral pelvic skin. In an immunohistochemical study, the ENaC α-subunit protein was specifically localized to the apical membrane of the principal cells but not the intercalated cells from the late distal tubule to the collecting duct in the kidney or in the apical area of cells of urinary bladder epithelia. When toads were acclimated to dry and hyper-saline environments, the levels of ENaC mRNA expression in the kidney and urinary bladder decreased under hyper-saline acclimation, but not under dry conditions. Immunohistochemical observations indicated that the levels of ENaC protein expression were much lower in the apical area of renal distal tubules and urinary bladder epithelia of hyper-saline acclimated toad compared with controls. The present study suggests that Bufo ENaC is significantly expressed and functions during Na+ reabsorption in the apical membrane domain in the distal nephron of normal and desiccated toads. Natriuresis may be caused by decreases in ENaC expression and its trafficking to the cell surface in the distal nephron, a response to prevent excessive Na+ reabsorption in hyper-saline-acclimated toads. [ABSTRACT FROM AUTHOR]

With-no-lysine kinase-1 (WNK1) gene mutations cause familial hyperkalemic hypertension (FHHt), a Mendelian disorder of excessive renal Na+ and K+ retention. Through its catalytic activity, full-length kinase-sufficient WNK1 (L-WNK1) suppresses its paralog, WNK4, thereby upregulating thiazide-sensitive Na-Cl cotransporter (NCC) activity. The predominant renal WNK1 isoform, KS-WNK1, expressed exclusively and at high levels in distal nephron, is a shorter kinase-defective product; the function of KS-WNK1 must therefore be kinase independent. Here, we report a novel role for KS-WNK1 as a dominant-negative regulator of L-WNK1. Na+ transport studies in Xenopus laevis oocytes demonstrate that KS-WNK1 downregulates NCC activity indirectly, by inhibiting L-WNK1. KS-WNK1 also associates with L-WNK1 in protein complexes in oocytes and attenuates L-WNK1 kinase activity in vitro. These observations suggest that KS-WNK1 plays an essential role in the renal molecular switch regulating Na+ and K+ balance; they provide insight into the kidney-specific phenotype of FHHt. [ABSTRACT FROM AUTHOR]

Angiotensin II (ANG II)-infused rats exhibit increases in distal nephron renin expressed in principal cells of connecting tubules and collecting ducts. This study was performed to determine whether the augmentation of distal nephron renin involves ANG II type 1 (AT1) receptor activation. Male Sprague-Dawley rats (200-220 g) were divided into three groups: 1) sham operated (n = 8); 2) ANG II infused (80 ng/min, 13 days, n = 8); and 3) ANG II infused plus AT1 receptor blocker (ARB), olmesartan (5 mg/days, n = 8). ANG II infusion increased systolic blood pressure (BP; 178 ± 4 vs. 122 ± 1 mmHg; P < 0.001) and suppressed plasma renin activity (PRA; 0.08 ± 0.1 vs. 5.3 ± 0.8 ng ANG I·ml-1·h-1). ARB treatment prevented the increase in BP (113 ± 6 mmHg) and led to increases in PRA (15.8 ± 1.5 ng ANG I·ml-1·h-1). Renin protein levels measured in the kidney medulla, to avoid contribution from juxtaglomerular appartus cells, were higher in ANG II-infused rats [1.64 ± 0.3 vs. 1.00 ± 0.1 densitometric units (DU) compared with sham-operated rats: P < 0.05], and ARB treatment prevented this increase (1.01 ± 0.1). Similarly, renin immunoreactivity increased in medullary collecting ducts of ANG II-infused compared with sham-operated rats (2.5 ± 0.3 vs. 1.0 ± 0.2 DU; P < 0.001), which was also prevented by ARB (1.01 ± 0.06). Renin qRTPCR in ANG II-infused rats showed higher mRNA levels in the kidney medulla compared with sham-operated rats (5.5 ± 2.3 vs. 0.04 ± 0.02 ratio to GAPDH mRNA levels; P < 0.001); however, renin transcript levels were normalized in the ARB-treated rats. These data demonstrate that the augmentation of distal nephron renin in ANG II-infused hypertensive rats is AT1 receptor mediated. The augmented distal tubular renin may contribute to increased intratubular ANG II levels and distal nephron sodium reabsorption in ANG II-dependent hypertension. [ABSTRACT FROM AUTHOR]

Chromophobe renal cell carcinoma (RCC) and collecting duct carcinoma (CDC) are derived from the collecting duct epithelia, although their morphology, molecular biologic characteristics and clinical behaviors are quite different. Herein is presented a case of RCC possessing the chromophobe RCC and CDC elements occurring in a 64 year-old Japanese woman. The patient was referred to Yokohama City University Hospital with complaints of persistent back pain and fever. Radiologic examinations revealed a left renal tumor, and radical nephrectomy was performed. The patient died with multiple metastases, 8 months after the operation. The resected tumor showed an invasive growth, and its cut surface was heterogenous with hemorrhage and necrosis. Histologically, the tumor was composed of chromophobe elements with dedifferentiation, and CDC elements. The chromophobe and CDC elements had obvious histological transition. Lectin histochemistry and immunohistochemistry confirmed that this tumor was derived from the distal nephron. c-KIT, p53 and Ki67 antigen showed differential localization between the chromophobe and CDC elements, even in the transitional areas. Along with the previous reports, the present case seemed to be composite RCC derived from the collecting duct, which might present clues to elucidate carcinogenesis in the distal nephron. [ABSTRACT FROM AUTHOR]

Large, Ca2+-activated K+ channels (BK), comprised of α- and β-subunits, mediate K+ secretion during high flow rates in distal nephron segments. Because the BK-β1 subunit enhances Ca2+ sensitivity of BK in a variety of cells, we determined its role in flow-induced K+ secretion and its localization in the mammalian nephron. To determine the role of BK-β1 in the kaliuretic response to volume expansion, the rate of K+ excretion (UKV) vs. varied urinary flow rates were determined in wild-type and BK-β1 knockout mice (BK-β1-/-). When flow rate was varied by volume expansion (2 ml·h-1·25 g body wt-1) for 30 to 60 min in wild-type mice, we found that the UKV increased significantly with increasing urine flow rates (r² = 0.50, P < 0.00001, n = 31), as demonstrated previously in distal nephron of rats and rabbits. However, in BK-β1-/- mice, UKV did not vary with changing flow rates (r² = 0.15, P = 0.08, n = 20). Using immunohistochemical techniques, we found that BK-β1 was strongly expressed in the apical membrane of the murine distal nephron and that 98% of BK-β1 protein detected by histochemistry colocalized with NCX, a marker of connecting tubules (CNT). Both BK-β1 and NCX colocalized with BK-α in separate experiments. Furthermore, we confirmed BK-β1 protein expression in the apical membrane of connecting tubules in rabbits. BK-β1 RNA from rabbit CNT was sequenced and was identical to previously published rabbit muscle sequences. These data show that the BK-β1 accessory subunit is present in the CNT segment of the mammalian distal nephron and has a significant role in the kaliuretic response to increased urinary flow induced by volume expansion. [ABSTRACT FROM AUTHOR]

There are few published reports of low-grade renal epithelial tumor originating from the distal nephron. However, it should not be disregarded clinically, because the actual number of patients with such tumors may be higher than expected. We investigated the immunohistochemical profile of a histologically distinct subtype of such a tumor in detail, in addition to the clinical course and imaging studies. The present study demonstrated that both glandular and spindle cell components of this tumor have a persistent characteristic of an epithelial tumor arising from the distal tubule or collecting duct. This tumor is a benign complex neoplasm that can be treated successfully with radical surgery. Beta-catenin and E-cadherin are suggested to play a crucial role in tumorigenesis and the biphasic arrangement of this neoplasm, concerning the expression of epithelial membrane antigen and carbohydrate antigen 19-9. We suggest that the term ‘distal nephron epithelioma’ is appropriate for classifying such rare but clinicopathologically distinct tumors. [ABSTRACT FROM AUTHOR]

Background: Recent studies have shown that renal expression of 25-hydroxyvitamin D3-1alpha-hydroxylase (1alpha-OHase) is not restricted to proximal tubules. To investigate the significance of this expression, we characterized the regulation of 1alpha-OHase expression and activity in a human cortical collecting duct cell line (HCD).Methods: Expression of 1alpha-OHase mRNA and protein was assessed by reverse transcription-polymerase chain reaction (RT-PCR) and Western blot analyses. Enzyme activity was quantified using 25-hydroxyvitamin D3 as the substrate; conversion to 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] and 24,25-dihydroxyvitamin D3 was then determined by thin-layer chromatography.Results: HCD cells expressed mRNA and protein for 1alpha-OHase. However, basal 1,25(OH)2D3 production was lower than that observed in proximal tubule HKC-8 cells. In both cell lines, synthesis of 1,25(OH)2D3 was increased by forskolin, parathyroid hormone, and low calcium medium. Conversely, treatment with 1,25(OH)2D3 itself decreased 1alpha-OHase activity. This effect was more pronounced in HCD cells, which also demonstrated significantly higher levels of 24-hydroxylase activity. The most striking induction of 1alpha-OHase activity was observed in the HCD cells following incubation with lipopolysaccharide, which was coincident with the expression of mRNA for both CD14 and Toll-like receptor 4.Conclusions: These results highlight the capacity for synthesis of 1,25(OH)2D3 in cells from more distal areas of the nephron. However, more sensitive feedback regulation and immune induction of 1alpha-OHase in the HCD cells suggest a more localized role for 1,25(OH)2D3 production in the distal nephron. [ABSTRACT FROM AUTHOR]

Chromophobe renal cell carcinoma (RCC), a newly established subtype of renal neoplasm, is composed of tumor cells with characteristically cloudy, weakly eosinophilic and reticular cytoplasm. The tumor should be distinguished from the common clear cell RCC, because of the unique clinicopathological and molecular biological features. The tumor does not show gender bias. Patient ages are similar to those of clear cell RCC, but might occur in the 20- to 40-year-old age group. Grossly, the tumor tends to be beige in color, which is different from the yellowish color of common RCC. Electron microscopy and immunohistochemistry indicate the intercalated cell of the collecting duct as the cellular origin. Cytogenetic study shows non-random multiple chromosome loss, with mitochondrial DNA rearrangement. Alteration of the von Hippel–Lindau (VHL) gene, a cancer suppressor gene relating with clear cell RCC, has not yet been observed. In order to adopt the most appropriate treatment, including gene therapy, recognition and correct pathological diagnosis of chromophobe RCC are extremely important. [ABSTRACT FROM AUTHOR]

Kinins are vasoactive paracrine peptides which participate in a wide range of functions, including the regulation of local organ blood flow, systemic blood pressure, transepithelial water and electrolyte transport, cellular growth, capillary permeability and inflammatory response, and pain. The recent introduction of specific bradykinin receptor subtype antagonists has greatly advanced our understanding of the role of the kallikrein-kinin system (KKS) in various physiological and disease states. However, a major gap remains in our knowledge of the role of kinins in early development. In this review, evidence is presented that the developing nephron expresses both tissue kallikrein and kininogen, and that the genes encoding the components of the KKS are subject to considerable developmental regulation. The activity of the intrarenal kinin-generating system is lowest in the developing kidney and increases with age. Completion of nephrogenesis is characterized by a marked surge in intrarenal kallikrein synthesis and gene transcription. Maturation is associated with redistribution of intrarenal kallikrein and its messenger RNA from the inner to outer cortical nephrons following the centrifugal pattern of nephron development. Challenges for the future include delineation of the direct role of kinins in the maturation of renal functions and elucidation of the molecular mechanisms underlying the developmental expression of the KKS. [ABSTRACT FROM AUTHOR]

Copyright of Klinische Wochenschrift is the property of Springer Nature and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)