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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.

The developmental mechanisms that orchestrate differentiation of specific nephron segments are incompletely understood, and the factors that maintain their terminal differentiation after nephrogenesis remain largely unknown. Here, the transcription factor AP-2β is shown to be required for the differentiation of distal tubule precursors into early stage distal convoluted tubules (DCTs) during nephrogenesis. In contrast, its downstream target KCTD1 is essential for terminal differentiation of early stage DCTs into mature DCTs, and impairment of their terminal differentiation owing to lack of KCTD1 leads to a severe salt-losing tubulopathy. Moreover, sustained KCTD1 activity in the adult maintains mature DCTs in this terminally differentiated state and prevents renal fibrosis by repressing β-catenin activity, whereas KCTD1 deficiency leads to severe renal fibrosis. Thus, the AP-2β/KCTD1 axis links a developmental pathway in the nephron to the induction and maintenance of terminal differentiation of DCTs that actively prevents their de-differentiation in the adult and protects against renal fibrosis.

Chromophobe renal cell carcinoma (chRCC) represents 5% of all RCC. ChRCC appears to arise from the distal nephron, in contrast to clear cell RCC that arises from the proximal nephron. ChRCC is distinctive because the tumor cells contain abundant abnormal mitochondria, and frequently have loss of one copy of multiple chromosomes, with a low rate of somatic mutations. Here, we focus on recent discoveries related to genetic and metabolic factors that may promote the progression of chRCC and summarize how these findings may relate to the prognosis and treatment of chRCC.

Glycolytic activity is increased in proliferating cells, leading to the concept that glycolysis could be a therapeutic target in cystic diseases and kidney cancer. Preclinical studies using the glucose analog 2-deoxy-d-glucose have shown promise; however, inhibiting glycolysis in humans is unlikely to be without risks. While proximal tubules are predominantly aerobic, later segments are more glycolytic. Understanding exactly where and why glycolysis is important in the physiology of the distal nephron is thus crucial in predicting potential adverse effects of glycolysis inhibitors. Live imaging techniques could play an important role in the process of characterizing cellular metabolism in the functioning kidney. The goal of this review is to briefly summarize recent findings on targeting glycolysis in proliferative kidney diseases and to highlight the necessity for future research focusing on glycolysis in the healthy kidney.

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.

More than two dozen types of potassium channels, with different biophysical and regulatory properties, are expressed in the kidney, influencing renal function in many important ways. Recently, a confluence of discoveries in areas from human genetics to physiology, cell biology, and biophysics has cast light on the special function of five different potassium channels in the distal nephron, encoded by the genes KCNJ1, KCNJ10, KCNJ16, KCNMA1, and KCNN3. Research aimed at understanding how these channels work in health and go awry in disease has transformed our understanding of potassium balance and provided new insights into mechanisms of renal sodium handling and the maintenance of blood pressure. This review focuses on recent advances in this rapidly evolving field.

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.

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.

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.

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

Within the CCD of the distal nephron of the rabbit, the BK (maxi K) channel mediates Ca2+- and/or stretch-dependent flow-induced K+secretion (FIKS) and contributes to K+adaptation in response to dietary K+loading. An unresolved question is whether BK channels in intercalated cells (ICs) and/or principal cells (PCs) in the CCD mediate these K+secretory processes. In support of a role for ICs in FIKS is the higher density of immunoreactive apical BKα (pore-forming subunit) and functional BK channel activity than detected in PCs, and an increase in IC BKα expression in response to a high-K+diet. PCs possess a single apical cilium which has been proposed to serve as a mechanosensor; direct manipulation of cilia leads to increases in cell Ca2+concentration, albeit of nonciliary origin. Immunoperfusion of isolated and fixed CCDs isolated from control K+-fed rabbits with channel subunit-specific antibodies revealed colocalization of immunodetectable BKα- and β1-subunits in cilia as well as on the apical membrane of cilia-expressing PCs. Ciliary BK channels were more easily detected in rabbits fed a low-K+vs. high-K+diet. Single-channel recordings of cilia revealed K+channels with conductance and kinetics typical of the BK channel. The observations that 1) FIKS was preserved but 2) the high-amplitude Ca2+peak elicited by flow was reduced in microperfused CCDs subject to pharmacological deciliation suggest that cilia BK channels do not contribute to K+secretion in this segment, but that cilia serve as modulators of cell signaling.

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

Large, Ca2+-activated K+ channels (BK) are comprised of the pore-forming α subunit (BK-α) and one of four β subunits (β1–4), which bestows functional diversity. BK channels are localized in the distal nephron of kidneys and a variety of extrarenal, fluid-secreting epithelia. In the kidneys, flow activates BK, which are located in two cell types of the distal nephron. The BK-α/β1 resides in the connecting tubule cells, and the BK-α/β4 is localized in intercalated cells, where they mediate K+ secretion in mice on a high K+, alkaline diet.

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.

The cortical collecting duct (CCD) of the distal nephron is a major site for the final renal regulation of K homeostasis. Within this segment, principal cells (PCs) possess a single apical cilium w...

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.

The mammalian kidney excretes its metabolic acid load through the proton-transporting cells, intercalated cells, in the distal nephron and collecting duct. Fish excrete acid through external organs, gill, or skin; however, the cellular function is still controversial. In this study, molecular and electrophysiological approaches were used to identify a novel cell type secreting acid in skin of zebrafish ( Danio rerio) larvae. Among keratinocytes covering the larval surface, novel proton-secreting ionocytes, proton pump (H+-ATPase)-rich cells, were identified to generate strong outward H+flux. The present work demonstrates for the first time, with a noninvasive technique, H+-secreting cells in an intact animal model, the zebrafish, showing it to be a suitable model in which to study the functions of vertebrate transporting epithelia in vivo.

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.

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...

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 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.

Carcinoma of the collecting ducts of Bellini are rare aggressive neoplasms of distal nephron origin; it accounts for 2% of all renal cell carcinomas. Clinically, collecting duct carcinoma (CDC) is characterized by an extremely aggressive phenotype, accompanying metastatic diseases at presentation in most reported cases. Most cases of CDC have been reported with median age above 50 years. To our knowledge, this is the only second case report of CDC in a young teenager patient below 19 years with metastasis to lungs and bones at presentation.

Sgk1 is a relatively unstable kinase that regulates epithelial Na+transport in the distal nephron of the kidney. We identified a 5′ variant alternate transcript of human Sgk1 (Sgk1_v3) that is expressed in the connecting tubule and collecting duct, is regulated by aldosterone and insulin, and is predicted to encode an NH2-terminal variant Sgk1 isoform, Sgk1_i3. Sgk1_i3 contains a polybasic motif, KKR, in its NH2terminus that regulates ubiquitination and stability of the expressed protein in HEK293 cells. In Fisher rat thyroid, and mpkCCDc14cells, Sgk1_i3 had a significantly greater effect on Na+transport compared with Sgk1 and its stimulatory effect was dependent on the kinase domain. Sgk1_i3 increased the abundance of cleaved epithelial Na+channel (ENaC) subunits at the cell surface, which was inhibited by coexpression of Nedd4–2. Together, the data demonstrate that a renally expressed Sgk1 isoform, Sgk1_i3, shows improved stability, is regulated by insulin and aldosterone, and stimulates ENaC activity when heterologously expressed in collecting duct cells.

Aldosterone is thought to be the main hormone to stimulate the epithelial sodium channel (ENaC) in the aldosterone-sensitive distal nephron (ASDN) comprising the late distal convoluted tubule (DCT2), the connecting tubule (CNT) and the entire collecting duct (CD). There is immunohistochemical evidence for an axial gradient of ENaC expression along the ASDN with highest expression in the DCT2 and CNT. However, most of our knowledge about renal ENaC function stems from studies in the cortical collecting duct (CCD). Here we investigated ENaC function in the transition zone of DCT2/CNT or CNT/CCD microdissected from mice maintained on different sodium diets to vary plasma aldosterone levels. Single-channel recordings demonstrated amiloride-sensitive Na+channels in DCT2/CNT with biophysical properties typical for ENaC previously described in CNT/CCD. In animals maintained on a standard salt diet, the average ENaC-mediated whole cell current (Δ Iami) was higher in DCT2/CNT than in CNT/CCD. A low salt diet increased Δ Iamiin CNT/CCD but had little effect on Δ Iamiin DCT2/CNT. To investigate whether aldosterone is necessary for ENaC activity in the DCT2/CNT, we used aldosterone synthase knockout (AS−/−) mice that lack aldosterone. In CNT/CCD of AS−/−mice, Δ Iamiwas lower than that in wild-type (WT) animals and was not stimulated by a low salt diet. In contrast, in DCT2/CNT of AS−/−mice, Δ Iamiwas similar to that in DCT2/CNT of WT animals both on a standard and on a low salt diet. We conclude that ENaC function in the DCT2/CNT is largely independent of aldosterone which is in contrast to its known aldosterone sensitivity in CNT/CCD.

Claudins play a major role in the regulation of paracellular electrolyte reabsorption in the kidney. This review describes the recent findings of the physiological function of claudins underlying the paracellular transport mechanisms for Cl(-) reabsorption in the collecting duct.There are two parallel mechanisms for transepithelial Cl(-) reabsorption in the collecting duct that utilize the Na-driven Cl-bicarbonate exchanger (NDCBE) and the claudin-based paracellular channel. Histological studies have demonstrated the renal localization of claudin-3, claudin-4, claudin-7, and claudin-8 in the collecting duct. Molecular analyses using several collecting duct cell models have come to the conclusion that claudin-4 functions as a paracellular Cl(-) channel. The channel function of claudin-4 is conferred by a charged lysine residue (K65) in its extracellular loop. Claudin-8 is required for paracellular Cl(-) permeation through its interaction with and recruitment of claudin-4 during tight junction assembly. Claudin-7 provides the basic barrier function to the collecting duct. Genetic ablation of claudin-7 in animals results in systemic dehydration owing to the loss of extracellular ions and fluid in the kidney.The paracellular pathway in the collecting duct is an important route for transepithelial Cl(-) reabsorption that determines the extracellular NaCl content and the blood pressure. In the collecting duct cells, claudin-4 and claudin-8 interact to form a paracellular Cl(-) channel, whereas claudin-7 maintains the transepithelial resistance. Different subsets of the claudin family proteins fulfill diverse aspects of the tight junction function that will be fundamental to understanding the physiology of the paracellular pathway.

PGE2 plays an important role in the regulation of fluid metabolism chiefly via antagonizing vasopressin-induced osmotic permeability in the distal nephron, but its enzymatic sources remain uncertain. The present study was undertaken to investigate the potential role of microsomal PGE synthase (mPGES)-1 in the regulation of urine concentrating ability after water deprivation (WD). Following 24-h WD, wild-type (WT) mice exhibited a significant reduction in urine volume, accompanied by a significant elevation in urine osmolality compared with control groups. In contrast, in response to WD, mPGES-1 knockout (KO) mice had much less urine volume and higher urine osmolality. Analysis of plasma volume by measurement of hematocrit and by using a nanoparticle-based method consistently demonstrated that dehydrated WT mice were volume depleted, which was significantly improved in the KO mice. WD induced a twofold increase in urinary PGE2 output in WT mice, which was completely blocked by mPGES-1 deletion. At baseline, the KO mice had a 20% increase in V2 receptor mRNA expression in the renal medulla but not the cortex compared with WT controls; the expression was unaffected by WD irrespective of the genotype. In response to WD, renal medullary aquaporin-2 (AQP2) mRNA exhibited a 60% increase in WT mice, and this increase was greater in the KO mice. Immunoblotting demonstrated increased renal medullary AQP2 protein abundance in both genotypes following WD, with a greater increase in the KO mice. Similar results were obtained by using immunohistochemistry. Paradoxically, plasma AVP response to WD seen in WT mice was absent in the KO mice. Taken together, these results suggest that mPGES-1-derived PGE2 reduces urine concentrating ability through suppression of renal medullary expression of V2 receptors and AQP2 but may enhance it by mediating the central AVP response.

The nephron segments beyond the proximal tubule are sometimes termed “the distal nephron”. In this chapter we will deal with the loop of Henle separately, and then consider the other components of the “distal nephron”.

Low molecular weight proteins are normally, essentially, entirely catabolized by the proximal tubules. Physiological and pathological aspects of the renal interactions of three of these proteins: light chains, B-2 microglobulin, and myoglobin share similarities, but are also unique in certain ways. In pathological conditions, some may produce glomerular, proximal or distal nephron and/or vascular alterations.

The thick ascending limb of the loop of Henle reabsorbs 20–30% of filtered sodium chloride (NaCl) and generates the osmotic gradient necessary for water absorption in the distal nephron. It is second

There is now convincing evidence that in addition to the vacuolar-type H+-ATPase, a gastric-type H+/K+-ATPase participates in acidification by the distal nephron. To determine whether a similar pump exists in the turtle bladder, we examined the dependence of acid secretion on mucosal K+ and the effects of supposedly specific inhibitors of the gastric H+/K+-ATPase, omeprazole and SCH 28080. In CO2-stimulated bladders both drugs produced dose-dependent inhibition of electrogenic H+ secretion measured as the reverse short-circuit current (RSCC). At the highest concentrations tested, H+ secretion decreased 45 ± 16% with mucosal and 20 ± 7% with serosal omeprazole (p < 0.01). SCH 28080 at 400 µM produced essentially complete inhibition of H+ secretion with either mucosal or serosal application. When H+ secretion was purposefully inhibited by DIDS or an adverse mucosal pH gradient, SCH 28080 had no effect on RSCC. Removing mucosal K+ (measured K+ < 50 µM) with or without mucosal barium, had no effect on RSCC. The inhibition of RSCC by omeprazole was reversed by mercaptoethanol. Finally, HCO3 secretion, as measured by either RSCC or pH-stat titration, increased significantly in response to 400 µM SCH 28080. The results demonstrate that these compounds inhibit acid secretion by the turtle bladder but stimulate the secretion of base. In view of the total independence of acid secretion on potassium, it is unlikely that any of the bladder’s acid secretion is mediated by an H+/K+-ATPase. The most reasonable interpretation of the data is that omeprazole and SCH 28080, previously thought to be specific inhibitors of the H+/K+-ATPase, also inhibit the vacuolar H+-ATPase of the turtle bladder. The results also indicate that HCO3 secretion by the bladder employ a different mechanism of H+ transport than is used for acid secretion; there is no simple reversal of polarity in the acid- versus base-secreting cells.

The role of mechanical forces in the regulation of glomerulotubular balance in the proximal tubule (PT) and Ca2+signaling in the distal nephron was first recognized a decade ago, when it was proposed that the microvilli in the PT and the primary cilium in the cortical collecting duct (CCD) acted as sensors of local tubular flow. In this review, we present a summary of the theoretical models and experiments that have been conducted to elucidate the structure and function of these unique apical structures in the modulation of Na+, HCO3−, and water reabsorption in the PT and Ca2+signaling in the CCD. We also contrast the mechanotransduction mechanisms in renal epithelium with those in other cells in which fluid shear stresses have been recognized to play a key role in initiating intracellular signaling, most notably endothelial cells, hair cells in the inner ear, and bone cells. In each case, small hydrodynamic forces need to be greatly amplified before they can be sensed by the cell's intracellular cytoskeleton to enable the cell to regulate its membrane transporters or stretch-activated ion channels in maintaining homeostasis in response to changing flow conditions.

Recently, the focus of interest on the role of the renin angiotensin system in the pathophysiology of hypertension has shifted towards greater emphasis on new developments in local renin angiotensin systems in specific tissues. We have focused our recent investigations on the role of the intrarenal-intratubular RAS in hypertension. All of the components needed for angiotensin II generation are present within the various compartments in the kidney. This brief review is focused on recent evidence that inappropriate activation of renin in distal nephron segments, by acting on angiotensinogen generated in the proximal tubule cells and delivered to the distal nephron may contribute to increased distal intrarenal angiotensin II formation, sodium retention and development and progression of hypertension.

In acute interstitial nephritis (AIN), mononuclear cells invade the tubules (tubulitis). The segmental localization of tubulitis is not precisely known. To clarify this question, formalin-fixed kidney biopsy specimens from 13 patients with AIN were studied stereologically by identifying cortical tubules with segment-specific markers. The periodic acid-Schiff reaction, peanut lectin, and antibodies against Tamm-Horsfall protein and epidermal cytokeratins all applied to the same section were used to identify the proximal tubules (PTs), distal convoluted tubules, distal straight tubules, and the cortical collecting system (connecting tubules and cortical collecting ducts), respectively. Morphometrically, an estimate of the relative volume of the inflammatory cell infiltrates within each category of tubular segments was obtained. Inflammatory cells were infrequently found in PTs (1.2%) but were frequently localized in distal tubules and the cortical collecting system (7.6%). There was no difference in the amount of the inflammatory cell infiltrate within these segments. Re-examination of an electron microscopic study of AIN carried out in this laboratory revealed that mononuclear cells were rarely seen in convoluted PTs but were frequently observed in straight PTs and all segments distal to them. The observations indicate that it is the distal nephron which is primarily affected by inflammatory cell infiltration in AIN.

All of the components needed for angiotensin II generation are present within the renal tubular network of the kidney. This brief review is focused on recent evidence demonstrating that inappropriate activation of renin in distal nephron segments by acting on angiotensinogen generated in the proximal tubule cells and delivered to the distal nephron may contribute to increased distal intrarenal angiotensin II formation, sodium retention, and development and progression of hypertension. Moreover, mechanisms regulating collecting duct renin during Ang II-dependent hypertension are discussed.

Summary Three‐to‐five days of water restriction in 5‐week‐old chickens produced mild dilatation of tubules in distal nephron segments and epithelial necrosis. Most dilated tubules were empty or contained proteinaceous casts. The histopathological changes were more frequent in the left kidney. Combined feed and water restriction decreased the incidence and severity of kidney changes. No histopathological changes were identified in kidneys of feed‐restricted chickens. The nephrosis produced by water restriction has features consistent with both prerenal and intrarenal pathophysiological mechanisms. Tubular transport of organic anions was increased by feed and water restriction.

Morphology of nephrotoxic injury _________________________________________ 65 Pathophysiology of cell injury _____________________________________________ 67 Disruption of energy production __________________________________________ 67 Mitochondrial dysfunction in acute renal failure ____________________________ 67 Defects in energy generation 67 Structural abnormalities 68 Endonuclease activation is the final common pathway of cell death __________ 68 Caspases and cell death __________________________________________________ 68 In vitro evidence of caspase activation in cytotoxicity 69 In vivo evidence of caspase activation in cytotoxicity 70 The Bcl-2 and the mitochondrial permeability transition: role in cell death _____ 70 The renal stress response determines whether cells survive or not ____________ 71 Molecular aspects 71 Signal transduction pathways in the stress response 71 Renal stress engages the cell cycle and is a determinant of cytotoxicity ________ 72 Summary _______________________________________________________________ 72 References ______________________________________________________________ 73

This review covers aspects of Na and K transport along the three major regions of the collecting duct: the cortical collecting duct (CCD), the outer medullary collecting duct (OMCD), and the inner medullary collecting duct (IMCD). The basic cell models that have been developed over the past several years are included with an attempt to integrate some of the newer observations on specific transport processes and their regulation. Heterogeneity along the collecting duct is emphasized as are species differences insofar as they are known. The normal mechanisms of Na and K transport are presented in the context of their long-term regulation by steroid hormones. The enormous increase in the understanding of the subject over the past several years precludes a detailed review of collecting duct function and its hormonal regulation. Mechanisms of Na transport regulation in epithelial models of the collecting duct are mentioned to the extent that they provide illustrative mechanisms. The lessons and principles derived from such model systems form a critical portion of the integrated understanding of distal nephron and collecting duct Na transport.

Vectorial Na+absorption across the aldosterone-sensitive distal nephron plays a key role in the regulation of extracellular fluid volume and blood pressure. Within this nephron segment, Na+diffuses from the urinary fluid into principal cells through an apical, amiloride-sensitive, epithelial Na+channel (ENaC), which is considered to be the rate-limiting step for Na+absorption. We have reported that increases in tubular flow rate in microperfused rabbit cortical collecting ducts (CCDs) lead to increases in net Na+absorption and that increases in laminar shear stress activate ENaC expressed in oocytes by increasing channel open probability. We therefore examined whether flow stimulates net Na+absorption ( JNa) in CCDs by increasing channel open probability or by increasing the number of channels at the apical membrane. Both baseline and flow-stimulated JNain CCDs were mediated by ENaC, as JNawas inhibited by benzamil. Flow-dependent increases in JNawere observed following treatment of tubules with reagents that altered membrane trafficking by disrupting microtubules (colchicine) or Golgi (brefeldin A). Furthermore, reducing luminal Ca2+concentration ([Ca2+]) or chelating intracellular [Ca2+] with BAPTA did not prevent the flow-dependent increase in JNa. Extracellular trypsin has been shown to activate ENaC by increasing channel open probability, and we observed that trypsin significantly enhanced JNawhen tubules were perfused at a slow flow rate. However, trypsin did not further enhance JNain CCDs perfused at fast flow rates. Similarly, the shear-induced increase in benzamil-sensitive JNain oocytes expressing protease resistance ENaC mutants was similar to that of controls. Our results suggest the rise in JNaaccompanying increases in luminal flow rates reflects an increase in channel open probability.