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KCNJ16 encodes Kir5.1 and acts in combination with Kir4.1, encoded by KCNJ10, to form an inwardly rectifying K+ channel expressed at the basolateral membrane of epithelial cells in the distal nephron. This Kir4.1/Kir5.1 channel is critical for controlling basolateral membrane potential and K+ recycling, the latter coupled to Na-K-ATPase activity, which determines renal Na+ handling. Previous work has shown that Kcnj16−/− mice and SSKcnj16−/− rats demonstrate hypokalemic, hyperchloremic metabolic acidosis. Here, we present the first report of a patient identified to have biallelic loss-of-function variants in KCNJ16 by whole exome sequencing who presented with chronic metabolic acidosis with exacerbations triggered by minor infections.

The epithelial Na(+) channel (ENaC) constitutes the rate-limiting step for Na(+) absorption in the aldosterone-sensitive distal nephron (ASDN) comprising the late distal convoluted tubule (DCT2), connecting tubule (CNT), and collecting duct (CD). Previously, we demonstrated that ENaC activity in the DCT2/CNT transition zone is constitutively high and independent of aldosterone, in contrast to its aldosterone dependence in the late CNT/initial cortical CD (CCD). The mineralocorticoid receptor (MR) is expressed in the entire ASDN. Its activation by glucocorticoids is prevented through 11β-hydroxysteroid dehydrogenase 2 (11β-HSD2) abundantly expressed in the late but probably not early part of the ASDN. We hypothesized that ENaC function in the early part of the ASDN is aldosterone independent but may depend on MR activated by glucocorticoids due to low 11β-HSD2 abundance. To test this hypothesis, we used doxycycline-inducible nephron-specific MR-deficient [MR knockout (KO)] mice. Whole cell ENaC currents were investigated in isolated nephron fragments from the DCT2/CNT or CNT/CCD transition zones using the patch-clamp technique. ENaC activity was detectable in the CNT/CCD of control mice but absent or barely detectable in the majority of CNT/CCD preparations from MR KO mice. Importantly, ENaC currents in the DCT2/CNT were greatly reduced in MR KO mice compared with ENaC currents in the DCT2/CNT of control mice. Immunofluorescence for 11β-HSD2 was abundant in the CCD, less prominent in the CNT, and very low in the DCT2. We conclude that MR is critically important for maintaining aldosterone-independent ENaC activity in the DCT2/CNT. Aldosterone-independent MR activation is probably mediated by glucocorticoids due to low expression of 11β-HSD2. NEW & NOTEWORTHY Using a mouse model with inducible nephron-specific mineralocorticoid receptor (MR) deficiency, we demonstrated that MR is not only critical for maintaining aldosterone-dependent ENaC activity in CNT/CCD but also for aldosterone-independent ENaC activity in DCT2/CNT. Furthermore, we demonstrated that cells of this latter nephron segment express little 11β-HSD2, which probably allows glucocorticoids to stimulate MR, resulting in aldosterone-independent ENaC activity in DCT2/CNT. This site-specific ENaC regulation has physiologically relevant implications for renal sodium and potassium homeostasis.

Objective: Vasopressin increases water permeability and activity of the epithelial sodium channel ENaC in the distal nephron of the kidney, favoring increased water and sodium retention. We hypothesized that genetically mediated differences in circulating vasopressin concentrations could contribute to features of hypertension that vary between non-Hispanic blacks and whites. To test this hypothesis we measured copeptin, a fragment of pre-pro arginine vasopressin with a prolonged half-life that is more easily quantified. Methods: The Genetic Epidemiology of Responses to Antihypertensives (GERA) I study was designed to identify predictors of response to diuretic therapy in non-Hispanic black and non-Hispanic white adults with essential hypertension. Blood samples were obtained after a 4 week washout of anti-hypertensive medication while on a sodium intake of 2 mmol/kg. Copeptin was measured from thawed EDTA plasma samples using the copeptin proAVP homogeneous automated immunofluorescent assay on the BRAHMS Kryptor Compact PLUS (Thermo Fisher). Results: Copeptin was significantly higher in blacks (median: 7.4 pmol/L, IQR: 4.4-10.2 pmol/L) than in whites (median: 5.4 pmol/L, IQR: 3.6-8.0 pmol/L) (p Conclusions: Higher copeptin (vasopressin) concentrations in non-Hispanic blacks compared to whites may contribute to the distinct differences in the hypertensive phenotypes between these racial groups. The underlying mechanism(s) require further study.

Kidney injury is a common finding in patients with liver disease. Bile cast nephropathy (also known as cholemic nephropathy) is an overlooked cause of renal injury in patients with hyperbilirubinemia. It can occur as a result of the toxic effects of bilirubin and bile acids on the renal tubules via several mechanisms. Bile cast nephropathy has characteristic histopathological changes consisting of bilirubin cast deposition in the distal nephron along with tubular epithelial cell injury. Treatment is based on the reversal of liver injury. This review aims to describe bile cast nephropathy in terms of its clinical and morphological features and to shed light on diagnostic techniques. In addition, we present data on management of such nephropathy while reviewing all the reported cases of bile cast nephropathy.

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., Graphical Abstract, In Brief Marneros shows that distal nephron defects from KCTD1 deficiency impair urinary transcellular Mg2+ reabsorption in distal convoluted tubules (DCTs) and paracellular Ca2+ and Mg2+ reabsorption in thick ascending limbs of Henle (TALs). Mice lacking KCTD1 develop hypomagnesemia and hypocalcemia, resulting in secondary hyperparathyroidism and consequently in progressive metabolic bone disease.

Aims Hyponatraemia is strongly associated with adverse outcomes in heart failure. However, accumulating evidence suggests that chloride may play an important role in renal salt sensing and regulation of neurohormonal and sodium-conserving pathways. Our objective was to determine the prognostic importance of hypochloraemia in patients with heart failure. Methods and results Patients in the BEST trial with baseline serum chloride values were evaluated (n = 2699). Hypochloraemia was defined as a serum chloride ≤96 mmol/L and hyponatraemia as serum sodium ≤135 mmol/L. Hypochloraemia was present in 13.0% and hyponatraemia in 13.7% of the population. Chloride and sodium were only modestly correlated (r = 0.53), resulting in only 48.7% of hypochloraemic patients having concurrent hyponatraemia. Both hyponatraemia and hypochloraemia identified a population with greater disease severity; however, renal function tended to be worse and loop diuretic doses higher with hypochloraemia. In univariate analysis, lower serum sodium or serum chloride as continuous parameters were each strongly associated with mortality (P

A low-Na+, high-K+diet (LNaHK) is considered a healthier alternative to the “Western” high-Na+diet. Because the mechanism for K+secretion involves Na+reabsorptive exchange for secreted K+in the distal nephron, it is not understood how K+is eliminated with such low Na+intake. Animals on a LNaHK diet produce an alkaline load, high urinary flows, and markedly elevated plasma ANG II and aldosterone levels to maintain their K+balance. Recent studies have revealed a potential mechanism involving the actions of alkalosis, urinary flow, elevated ANG II, and aldosterone on two types of K+channels, renal outer medullary K+and large-conductance K+channels, located in principal and intercalated cells. Here, we review these recent advances.

To better understand the role of the inward-rectifying K channel Kir4.1 (KCNJ10) in the distal nephron, we initially studied a global Kir4.1 knockout mouse (gKO), which demonstrated the hypokalemia and hypomagnesemia seen in SeSAME/EAST syndrome and was associated with reduced Na/Cl cotransporter (NCC) expression. Lethality by ~3 wk, however, limits the usefulness of this model, so we developed a kidney-specific Kir4.1 “knockdown” mouse (ksKD) using a cadherin 16 promoter and Cre-loxP methodology. These mice appeared normal and survived to adulthood. Kir4.1 protein expression was decreased ~50% vs. wild-type (WT) mice by immunoblotting, and immunofluorescence showed moderately reduced Kir4.1 staining in distal convoluted tubule that was minimal or absent in connecting tubule and cortical collecting duct. Under control conditions, the ksKD mice showed metabolic alkalosis and relative hypercalcemia but were normokalemic and mildly hypermagnesemic despite decreased NCC expression. In addition, the mice had a severe urinary concentrating defect associated with hypernatremia, enlarged kidneys with tubulocystic dilations, and reduced aquaporin-3 expression. On a K/Mg-free diet for 1 wk, however, ksKD mice showed marked hypokalemia (serum K: 1.5 ± 0.1 vs. 3.0 ± 0.1 mEq/l for WT), which was associated with renal K wasting (transtubular K gradient: 11.4 ± 0.8 vs. 1.6 ± 0.4 in WT). Phosphorylated-NCC expression increased in WT but not ksKD mice on the K/Mg-free diet, suggesting that loss of NCC adaptation underlies the hypokalemia. In conclusion, even modest reduction in Kir4.1 expression results in impaired K conservation, which appears to be mediated by reduced expression of activated NCC.

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-∆9). 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-∆9-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 stress-response kinase-1, which directly phosphorylate and activate the thiazide-sensitive Na+-Cl− cotransporter. Several groups have suggested different mechanisms by which CUL3-∆9 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-∆9-mediated FHHt. Analysis of this model suggested that CUL3-∆9 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-∆9 must contribute to the development of FHHt. In this review, we focus on our current knowledge of CUL3-∆9 action gained from in vitro and in vivo models that may help unravel this complex problem.

Long-standing experimental evidence suggests that epithelial cells in the renal tubule are able to sense osmotic and pressure gradients caused by alterations in ultrafiltrate flow by elevating intracellular Ca2+concentration. These responses are viewed as critical regulators of a variety of processes ranging from transport of water and solutes to cellular growth and differentiation. A loss in the ability to sense mechanical stimuli has been implicated in numerous pathologies associated with systemic imbalance of electrolytes and to the development of polycystic kidney disease. The molecular mechanisms conferring mechanosensitive properties to epithelial tubular cells involve activation of transient receptor potential (TRP) channels, such as TRPV4, allowing direct Ca2+influx to increase intracellular Ca2+concentration. In this review, we critically analyze the current evidence about signaling determinants of TRPV4 activation by luminal flow in the distal nephron and discuss how dysfunction of this mechanism contributes to the progression of polycystic kidney disease. We also review the physiological relevance of TRPV4-based mechanosensitivity in controlling flow-dependent K+secretion in the distal renal tubule.

Aldosterone acts through the mineralocorticoid receptor (MR) to modulate gene expression in target tissues. In the kidney, the principal action of aldosterone is to promote sodium conservation in the distal nephron and so indirectly enhance water conservation under conditions of hypotension. Over the last twenty years the rapid activation of protein kinase signalling cascades by aldosterone has been described in various tissues. This review describes the integration of rapid protein kinase D signalling responses with the non-genomic actions of aldosterone and transcriptional effects of MR activation.

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 (KLHL3S433-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 KLHL3S433-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.

β-Catenin signaling plays an important role in regulation of development as well as tissue homeostasis. Recently, we have shown that this pathway is involved in regulation of collecting duct (CD) function. Here, we examined the role of β-catenin signaling in AngII-induced hypertension. C57/BL6 mice were administered for 7 d with vehicle, AngII alone (300 ng/kg/min) or in combination with an inhibitor of β-catenin pathway, ICG-001 (50 mg/kg/d) both via an osmotic mini-pump. Radiotelemetry demonstrated that ICG-001 effectively attenuated AngII-induced hypertension (MAP on day 7: 112.5 ± 2.02 in Control group vs. 137±2.07 in AngII group vs. 119.2±2.26 mmHg in AngII/ICG-001 group, n = 7-9, p+ transport as assessed by epithelial volt-ohmmeter, which was completely blocked by ICG-001. Protein expression of α-ENaC exhibited the similar pattern of changes as ENaC activity whereas the expression of β- or γ-ENaC remained constant. ICG-001 was not associated with noticeable toxicity in vivo or in vitro. In summary, these results suggest that β-catenin signaling mediates AngII-induced hypertension at least in part through regulation of intrarenal renin response and the expression of Na+ transporters in the distal nephron.

The ClC-K2 chloride channel has long been known to robustly express in the basolateral plasma membranes of epithelial cells lining the thick ascending limb, distal convoluted tubule, and distal nephron (including the collecting duct system; [Stauber et al., 2012][1]; [Jentsch, 2015][2]). Our

To summarize recent evidence regarding the role of distal nephron segments other than the macula densa in sensing the tubular environment and transmitting this signal to the adjacent vasculature.In addition to the classical contact site between the macula densa plaque and the afferent arteriole, there is accumulating evidence suggesting a functional association between the distal nephron and the vasculature at three distinct additional sites: at the terminal cortical thick ascending limb, at the early distal tubule and also at the connecting tubule segment. The epithelial cells around the macula densa also sense and respond to changes in tubular flow and salt content and may transmit this signal to the adjacent afferent arteriole.There are multiple sites of anatomical and functional contact between the distal nephron and the vasculature supplying the glomerulus, and these may contribute to the regulation of glomerular filtration rate and renal hemodynamics.

High-conductance (BK or maxi) K+channels were observed in cell-attached patches of the apical membrane of the isolated split-open rat connecting tubule (CNT). These channels were quite rare in cells identified visually as principal cells (PCs; 5/162 patches) but common in intercalated cells (ICs; 24/26 patches). The BK-expressing intercalated cells in the CNT and cortical collecting duct (CCD) were characterized by a low membrane potential (−36 mV) under short-circuit conditions, measured from the reversal potential of the channel currents with similar K+concentrations on both sides of the membrane. Under whole-cell clamp conditions with low intracellular Ca2+, ICs had a very low K+conductance. When cell Ca2+was increased to 200 nM, a voltage-dependent, tetraethylammonium (TEA)-sensitive outward conductance was activated with a limiting value of 90 and 140 nS/cell in the CNT and CCD, respectively. Feeding animals a high-K diet for 1 wk did not increase these currents. TEA-sensitive currents were much smaller in PCs and usually below detection limits. To examine the possibility that the ICs participate in transepithelial K+secretion, we measured Na/K pump activity as a ouabain-sensitive current. Although these currents were easily observed in PCs, averaging 79 ± 14 and 250 ± 50 pA/cell in the CCD and CNT, respectively, they were below the level of detection in the ICs. We conclude that ICs have BK channel densities that are sufficient to support renal secretion of K+if cell Ca2+is elevated. However. a pathway for K+entry into these cells has not been identified.

Oxidative stress contributes to the development and maintenance of hypertension in male rodents, but studies in females have rarely shown a reduction in blood pressure (BP) with antioxidants. Tempol, a superoxide dismutase mimetic, decreases BP in young male SHR, but fails to reduce BP in either young or old female SHR, despite the fact that females have similar or higher levels of oxidative stress markers. The reason for the sex difference in the response to tempol remains unclear. Acetazolamide inhibits carbonic anhydrase in the proximal tubule thus increasing sodium delivery to the distal nephron, and thereby should increase distal oxidative stress. Acetazolamide was used to test the hypothesis that with increased sodium delivery to the distal nephron, tempol would reduce the BP in aging female SHR. Female SHR, 20-22 mos old, were divided into three groups (n=4-6/grp): Control (C), Acetazolamide (A), and Acetazolamide+Tempol (A+T). After baseline mean arterial pressure (MAP; telemetry), rats received vehicle (C) or acetazolamide (A and A+T). On day 8, rats in C and A+T groups were given tempol (30 mg/kg) for 11 days. Baseline MAP was similar (C: 170±7; A: 182±4; A+T: 172±6 mm Hg, p=NS). Tempol had no effect on MAP in C+T, but reduced MAP in A+T group (C+T: 169±1; A: 171±1; A+T: 151±5 mm Hg; p

The electrogenic Na+-HCO3− cotransporter 2 (NBCe2) is a newly discovered protein in the distal nephron. Our understanding is minimal regarding its physiological role in renal electrolyte transport. In this mini-review, we summarize the potential function of NBCe2 in the regulation of blood pressure, acid-base, and K+ and Ca2+ transport in the distal nephron.

Dietary K+ load results in hyperkalemia, with consequent aldosterone release in order to stimulate K+ secretion in the distal nephron. The molecular mechanisms of this regulation are still not full...

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.

Thiazide and loop diuretics induce renal K+secretion, often leading to renal K+wasting and hypokalemia. This phenomenon has been proposed to reflect an increase in delivery to and reabsorption of Na+by the distal nephron, with a resultant increase in the driving force for passive K+efflux across the apical membrane. Recent studies suggest that cellular mechanisms that lead to enhanced rates of Na+reabsorption as well as K+secretion in response to increases tubular flow rates are more complex. Increases in tubular flow rates directly enhance the activity of apical membrane Na+channels and indirectly activate a class of K+channels, referred to as maxi-K, that are functionally inactive under low flow states. This review addresses the role of biomechanical forces, generated by variations in urinary flow rate and tubular fluid volume, in the regulation of transepithelial Na+and K+transport in the distal nephron. The question of why the distal nephron has evolved to include a component of flow-dependent K+secretion is also addressed.

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.

In the kidney, the fine control of NaCl absorption takes place in the distal nephron and is controlled by aldosterone and vasopressin. This review summarizes the effects of vasopressin on Na+ transport mediated by the amiloride-sensitive epithelial sodium channel (ENaC) and the cystic fibrosis transmembrane conductance regulator (CFTR) Cl- channel in immortalized or primary cultured cortical collecting duct cells, expressing either the wild-type ENaC subunits, or mutations, or deletions of the PY domain of the beta- or gamma-ENaC subunits responsible for Liddle's syndrome, an inherited form of hypertension due to excessive salt absorption.

Potassium secretory flux ( JK) by the distal nephron is regulated by systemic and luminal factors. In the present investigation, JKwas measured with a double-barreled K+electrode during paired microperfusion of superficial segments of the rat distal nephron. We used control solutions (100 mM NaCl, pH 7.0) and experimental solutions in which Cl−had been replaced with a less permeant anion and/or pH had been increased to 8.0. JKincreased when Cl−was replaced by either acetate (∼37%), sulfate (∼32%), or bicarbonate (∼62%), and also when the pH of the control perfusate was increased (∼26%). The majority (80%) of acetate-stimulated JKwas Ba2+sensitive, but furosemide (1 mM) further reduced secretion (∼10% of total), suggesting that K+-Cl−cotransport was operative. Progressive reduction in luminal Cl−concentration from 100 to 20 to 2 mM caused increments in JKthat were abolished by inhibitors of K+-Cl−cortransport, i.e., furosemide and [(dihydroindenyl)oxy]alkanoic acid. Increasing the pH of the luminal perfusion fluid also increased JKeven in the presence of Ba2+, suggesting that this effect cannot be accounted for only by K+channel modulation of K+secretion in the distal nephron of the rat. Collectively, these data suggest a role for K+-Cl−cotransport in distal nephron K+secretion.

Renal acid-base balance may become deranged in a number of ways, some of which are the consequence of inherited disorders. Two main groups of distal renal acidopathies result either from a direct inability to secrete acid in the distal nephron (giving rise to type 1 renal tubular acidosis [RTA]) or

Mice on a low Na, high K diet (LNaHK) are able to excrete excess K and maintain K homeostasis by increasing tubular fluid flow in the distal nephron of the kidney. High tubular fluid flow increases...

The renal distal nephron [distal convoluted tubule 2 (DCT2) and connecting tubule (CNT)] is crucial for sodium and blood pressure regulation, and is tightly controlled by aldosterone. Due to diffic...

Recent studies point to direct actions of Ang II on ENaC-mediated Na+ reabsorption in the distal nephron via stimulation of AT1R that occurs independently of aldosterone-MR signaling. We used split...

The epithelial sodium channel (ENaC) is a major determinant of sodium (Na+) and water balance in the kidney. We previously reported that aldosterone (aldo) modifies expression of microRNAs (miRs) t...

It becomes recognized that Ca2+-permeable TRPV4 channel is predominantly expressed in the distal nephron and its activity is essential for [Ca2+]i elevations in response to increased tubular flow. Here, we probe whether TRPV4 function in the kidney is regulated by dietary intake of major electrolytes. We found that renal TRPV4 expression was increased by high potassium diet (5% K+) and decreased by dietary potassium restriction (0.003 % K+). Saturation of systemic mineralocorticoid status with DOCA had little effect on TRPV4 protein abundance pointing to an aldosterone-independent nature of this regulation. Increased TRPV4 expression by high potassium intake was associated with significantly augmented [Ca2+]i responses to elevated flow as was detected using Fura-2 Ca2+ imaging in freshly isolated split-opened distal nephrons. Furthermore, dietary sodium load (2% Na+) similarly increased renal TRPV4 expression and enhanced flow-induced [Ca2+]i responses in distal nephron cells. Interestingly, protein level...

The inward rectifying potassium channels of the ROMK family are present in the distal nephron of the kidney. These channels have two membrane spanning portions, between which lies a hydrophobic domain thought to confer the majority of the conductive properties of the channel. The N- and C-termini are both intracellular. In this paper we have examined the contribution of the N- and C-termini to the pore by examining the interaction of Cs+ with the channels. ROMK1 has an additional 19 amino acids on its N-terminus in comparison to ROMK2. The C-terminus of ROMK2 was extended by addition of a streptavidin tag (sfROMK2). Currents were measured following expression in Xenopus oocytes using two-electrode voltage clamp. ROMK1, ROMK2 and sfROMK2 exhibited concentration- and voltage-dependent block of inward currents by extracellular Cs+. The Hill coefficients were not significantly different from one. The mean Kd values at 0 mV were 100.6 ± 10.6, 63.1 ± 3.9 and 40.6 ± 9.4, respectively (p < 0.05). The electric distances (δ) were 0.94 ± 0.06, 1.0 ± 0.05 and 1.37 ± 0.06 respectively. The δ of sfROMK2 was greater than either ROMK1 or ROMK2 (p < 0.001). ROMK1, ROMK2 and sfROMK2 are sensitive to extracellular Cs+. Block was both concentration- and voltage-dependent. sfROMK2 is most Cs+-sensitive. ROMK1 contains an additional N-terminal 19 amino acids. Thus the pore properties of these two isoforms are subtly different, and influenced by the N-terminus. The lower Kd in sfROMK2 suggests that the streptavidin tag, and perhaps the C-terminus, also affect the pore.

The mechanism by which dietary K+ restriction induces distal–nephron Li+ reabsorption was investigated by administration of bendroflumethiazide (BFTZ) or vehicle in conscious Wistar rats. Changes in fractional excretion of Li+ following administration of amiloride (ΔFELi) were used as an index of distal tubular Li+ reabsorption. The results revealed an absence of distal tubular Li+ reabsorption in K+–replete rats (ΔFELi = 3.6±2.4%), in contrast to K+ restriction in which ΔFELi was 24.0±2.7%. The distal tubular Li+ reabsorption in K+–depleted rats was significantly reduced by preadministration of BFTZ (ΔFELi = 9.2±0.9%). The fractions of Li+ and Na+ reabsorbed in the amiloride–sensitive segment were different in K+–replete rats (9±6 vs. 60±6%), but similar in K+–depleted rats (61±5 vs. 73±4%). BFTZ administration to K+–depleted rats resulted in a proportional decrease in these fractions, suggesting competition between Na+ and Li+ for reabsorption in the distal–nephron segment during K+ depletion. These results are compatible with the hypothesis that during K+ depletion the reabsorption of Li+ in the distal–nephron segment is competitively inhibited by Na+.

The epithelial Na+channel (ENaC) controls the rate-limiting step in the process of transepithelial Na+reabsorption in the distal nephron, the distal colon, and the airways. Hereditary salt-losing syndromes have been ascribed to loss of function mutations in the α-, β-, or γ-ENaC subunit genes, whereas gain of function mutations (located in the COOH terminus of the β- or γ-subunit) result in hypertension due to Na+retention (Liddle’s syndrome). In mice, gene-targeting experiments have shown that, in addition to the kidney salt-wasting phenotype, ENaC was essential for lung fluid clearance in newborn mice. Disruption of the α-subunit resulted in a complete abolition of ENaC-mediated Na+transport, whereas knockout of the β- or γ-subunit had only minor effects on fluid clearance in lung. Disruption of each of the three subunits resulted in a salt-wasting syndrome similar to that observed in humans.

The renal mechanisms underlying sodium retention during liver cirrhosis have been difficult to elucidate. Kim and associates describe a biphasic pattern of regulation of the renal epithelial sodium channel in the common bile duct ligation model, shedding some light on this issue.

We studied the characteristics of the basal and antidiuretic hormone (arginine vasotocin, AVT)-activated whole cell currents of an aldosterone-treated distal nephron cell line (A6) at two different cytosolic Ca2+ concentrations ([Ca2+] c , 2 and 30 nm). A6 cells were cultured on a permeable support filter for 10 ∼ 14 days in media with supplemental aldosterone (1 μm). At 30 nm [Ca2+] c , basal conductances mainly consisted of Cl− conductances, which were sensitive to 5-nitro-2-(3-phenylpropylamino)-benzoate. Reduction of [Ca2+] c to 2 nm abolished the basal Cl− conductance. AVT evoked Cl− conductances at 2 as well as 30 nm [Ca2+] c . In addition to Cl− conductances, AVT induced benzamil-insensitive nonselective cation (NSC) conductances. This action on NSC conductances was observed at 30 nm [Ca2+] c but not at 2 nm [Ca2+] c . Thus, cytosolic Ca2+ regulates NSC and Cl− conductances in a distal nephron cell line (A6) in response to AVT. Keeping [Ca2+] c at an adequate level seems likely to be an important requirement for AVT regulation of ion conductances in aldosterone-treated A6 cells.

Recent developments have led to renewed interest in the mechanisms that mediate the changes in sodium excretion in response to changes in arterial pressure, a phenomenon termed pressure-natriuresis. Pressure-natriuresis occurs in the absence of changes in filtered load and thus, the changes in sodium excretion are caused by changes in net tubular sodium reabsorption. Both proximal and distal nephron segments have been implicated as the sites of altered reabsorption. The specific mechanism responsible for pressure-natriuresis remains unresolved. One proposal suggests that changes in renal interstitial fluid pressure which may be due, in part, to a less efficient autoregulation of the renal medullary circulation than of the cortex, alter sodium reabsorption. Alternatively, recent studies indicate that increases in arterial pressure increase endothelial nitric oxide formation which inhibits sodium reabsorption via direct effects on the tubules as well as hemodynamically mediated effects.

The MDCK cell has proven to be a useful model cell line for the study of properties and regulation of renal epithelial ion channels. Patch clamp studies disclosed the existence of several K+ channels and of a Cl- channel, and their regulation by hormones, cell volume, trace elements and drugs. Most hormones affect K+ channels at least in part by increasing cytosolic Ca2+. However, indirect evidence points to additional mechanisms contributing to K+ channel activation. Cell swelling activates both K+ channels and unselective anion channels. ICln, a protein cloned from MDCK cells, is either a Cl- channel or a regulator of thereof. ICln is up-regulated by cellular acidification and is crucial for rapid regulatory cell volume decrease.

The inwardly rectifying, ATP-sensitive K+ channel (ROMK) was localized by in situ hybridization in the rat kidney. Tissue in situ hybridization revealed that transcripts encoding the ROMK channel were expressed predominantly in cortical and outer medullary nephron segments. The localization of ROMK mRNA to specific nephron segments was assessed by hybridization of isolated nephron segments with an ROMK-specific probe (single segment in situ hybridization). ROMK mRNA was present in cortical and medullary thick ascending limb, distal tubule, and cortical and outer medullary collecting ducts, but not in proximal tubule. A weak hybridization was observed with inner medullary collecting ducts. To confirm these results, serial cryosections were alternatively stained by hybridization histochemistry for ROMK mRNA or by immunocytochemistry using antibodies specific for S1, S2, or S3 proximal tubular segments. Tubular cells that displayed immunoreactivity with the proximal tubular segment-specific antibodies showed little, if any, ROMK message. In addition, using an in situ hybridization and immunocytochemistry double-labeling technique, ROMK transcripts and vitamin D-dependent calcium-binding protein were shown to colocalize to the distal tubule (distal convoluted tubule and connecting tubule). The overall nephron localization of ROMK mRNA shown in these studies is consistent with the possibility that this novel channel may represent the low-conductance ATP-sensitive K+ channel that has been identified in apical membranes of thick limb and collecting duct segments and is believed to participate in K+ secretion.