The distribution and function of aquaporins in the kidney: resolved and unresolved questions

The membrane water channel aquaporin (AQP) family is composed of 13 isoforms in mammals, eight of which are reportedly expressed in the kidney: AQP1, 2, 3, 4, 6, 7, 8, and 11. These isoforms are differentially expressed along the renal tubules and collecting ducts. AQP1 and 7 are distributed in the proximal tubules, whereas AQP2, 3, and 4 occur in the collecting duct system. They play important roles in the reabsorption of water and some solutes across the plasma membrane. In contrast to other aquaporins found in the kidney, AQP6, 8, and 11 are localized to the cytoplasm rather than to the apical or basolateral membranes. It is therefore doubtful that these isoforms are directly involved in water or solute reabsorption. AQP6 is localized in acid-secreting type A intercalated cells of the collecting duct. AQP8 has been found in the proximal tubule but its cellular location has not yet been defined by immunohistochemistry. AQP11 seems to be localized in the endoplasmic reticulum (ER) of proximal tubule cells. Interestingly, polycystic kidneys develop in AQP11-null mice. Many vacuole-like structures are seen in proximal tubule cells in kidneys of newborn AQP11-null mice. Subsequently, cysts are generated, and most of the mice die within a month due to severe renal failure. Although ER stress and impairment of polycystin-1, the product of the gene mutated in autosomal-dominant polycystic kidney disease, are possible causes of cystogenesis in AQP11-null mice, the exact mechanism of pathogenesis and the physiological function of AQP11 are yet to be resolved.     

Water transport in renal tubules is mediated by aquaporins

Water reabsorption in mammalian renal tubules is mediated by channel-forming membrane glycoproteins termed aquaporins (AQP). So far three different kinds of AQP have been described in renal tubules. AQP CHIP is localized to the luminal and contraluminal membranes of the proximal tubule and descending thin limb cells, i.e., in tubule segments that exhibit a constitutive high permeability to water that is insensitive to vasopressin. AQP-CD is present in subapical vesicles and the luminal membrane of collecting duct principal cells. Its intracellular distribution depends on vasopressin or hydration status of the animal and, thus, may represent the vasopressin-sensitive water channel. The basolateral integral protein (BLIP) may represent the vasopressin-insensitive water channel in basolateral membrane of collecting duct principal cells. The exact localization of a recently cloned homologue, WCH3, which may be either related to BLIP or represent yet another kind of AQP, is not known. Heterogeneity of aquaporins in the renal tubule may provide a molecular basis for the treatment of certain diseases with disturbances in water homeostasis.Abbreviations AQP aquaporin - CHIP28 28 kDa channelforming integral protein - WCH-CD water channel-collecting duct - BLIP basolateral integral protein - PT proximal tubule - DTL descending thin limbs Correspondence to: I. Saboli     

Fluvastatin modulates renal water reabsorption in vivo through increased AQP2 availability at the apical plasma membrane of collecting duct cells

X-linked nephrogenic diabetes insipidus (XNDI), a severe pathological condition characterized by greatly impaired urine-concentrating ability of the kidney, is caused by inactivating mutations in the V2 vasopressin receptor (V2R) gene. The lack of functional V2Rs prevents vasopressin-induced shuttling of aquaporin-2 (AQP2) water channels to the apical plasma membrane of kidney collecting duct principal cells, thus promoting water reabsorption from urine to the interstitium. At present, no specific pharmacological therapy exists for the treatment of XNDI. We have previously reported that the cholesterol-lowering drug lovastatin increases AQP2 membrane expression in renal cells in vitro. Here we report the novel finding that fluvastatin, another member of the statins family, greatly increases kidney water reabsorption in vivo in mice in a vasopressin-independent fashion. Consistent with this observation, fluvastatin is able to increase AQP2 membrane expression in the collecting duct of treated mice. Additional in vivo and in vitro experiments indicate that these effects of fluvastatin are most likely caused by fluvastatin-dependent changes in the prenylation status of key proteins regulating AQP2 trafficking in collecting duct cells. We identified members of the Rho and Rab families of proteins as possible candidates whose reduced prenylation might result in the accumulation of AQP2 at the plasma membrane. In conclusion, these results strongly suggest that fluvastatin, or other drugs of the statin family, may prove useful in the therapy of XNDI.     

水通道蛋白2、 3、 4在小鼠肾集合管发育中的表达

目的:观察水通道蛋白(AQP)-2、 3、 4在小鼠肾集合管发育中的表达,探讨其与集合管发育的关系及作用.方法:免疫组织化学技术结合体视学方法半定量检测AQP-2、 AQP-3及AQP-4的表达.结果:AQP-2从胚龄17d开始表达在集合管主细胞游离面细胞膜,到生后1d其阳性表达的游离面细胞膜面密度值逐渐增加,此后无明显变化;AQP-3、 AQP-4在胚龄14d集合管侧基底细胞膜(基底膜和侧膜)可见微弱表达,并随胚龄的增长而增加,到生后1d达最高,以后无明显变化.结论:AQP对小鼠出生前后肾水平衡具有重要的调节作用,其AQP-2的作用发生在出生后,而AQP-3、 AQP-4的作用发生在出生前.     

Direct renin inhibitor aliskiren increases AQP2 expression in renal collecting ducts and improves urinary concentration defect in NDI

AIM: The direct renin inhibitor aliskiren displays antihypertensive and antialbuminuric effects in humans and in animal models.Emerging evidence has shown that aliskiren localizes and persists in medullary collecting ducts even after treatment was discontinued.The purpose of the present study was to investigate whether aliskiren regulates renal aquaporin expression and improves urinary concentrating defect induced by lithium. METHODS: The mice were either fed with normal chow or Li Cl diet(40 mmol / kg dry food per day for first 4 days and 20 mmol / kg dry food per day for last 3 days) for seven days. Some mice were intraperitoneally injected aliskiren(50 mg/kg BW per day in saline). RESULTS: Mice injected aliskiren developed decreased urine output and increased urine osmolality when compared with controls. Aliskiren significantly increased protein abundance of AQP2 and phosphorylated-S256 AQP2 in the kidney inner medulla. Immunohistochemistry and immunofluoresence showed increased apical and intracellular labeling of AQP2 and p S256-AQP2 in collecting duct principal cells of kidneys in mice treated with aliskiren. Aliskiren treatment prevented urinary concentrating defect in lithium-treated mice,and improved the downregulation of AQP2 and p S256-AQP2 protein abundance in inner medulla of the kidney. In primary cultured rat inner medulla collecting duct cells,aliskiren dramatically increased AQP2 protein abundance which was significantly inhibited either by PKA inhibitor H89 or by adenylyl cyclase inhibitor MDL12330,indicating an involvement of the c AMP signalling pathway in mediating aliskiren-induced increased AQP2 expression. CONCLUSION: The direct renin inhibitor aliskiren upregulates AQP2 protein expression in inner medullary collecting duct principal cells and prevents lithium-induced nephrogenic diabetes insipidus( NDI) likely via PKA-c AMP pathways.     

Congenital nephrogenic diabetes insipidus: what can we learn from mouse models?

Aquaporins (AQPs) are central players in mammalian physiology, allowing efficient water transport through cellular membranes. To date, 13 different aquaporins have been identified in mammals (AQP0–AQP12). Knocking out genes in mice and identification of mutations in the human genes provided important information on the role of AQPs in normal physiology. While the physiological role of many AQPs only becomes clear when the putative function is challenged, the lack of AQP2 directly results in a disease phenotype. Aquaporin 2 is highly expressed in the principal cells of the renal collecting duct, where it shuttles between intracellular storage vesicles and the apical membrane. Upon hypernatraemia or hypovolaemia, the antidiuretic hormone vasopressin (AVP) is released from the pituitary into blood and binds to its type 2 receptor on renal principal cells. This initiates a cAMP signalling cascade resulting in the translocation of AQP2-bearing vesicles to the apical membrane. Subsequently, pro-urinary water reabsorption and urine concentration occurs. This process is reversed by a reduction in circulating AVP levels, which is obtained with the establishment of isotonicity. In humans, mutations in the AQP2 gene cause congenital nephrogenic diabetes insipidus (NDI), a disorder characterized by an inability to concentrate urine in response to vasopressin. Until the recent development of several congenital NDI mouse models, our knowledge on AQP2 regulation was primarily based on in vitro studies. This review focuses on the similarities between the in vitro and in vivo studies and discusses new insights into congenital NDI obtained from the mouse models.     

The pattern of distribution of selected ATP-sensitive P2 receptor subtypes in normal rat kidney: an immunohistological study

Using immunohistological techniques and available polyclonal antibodies, we have identified several ATP-sensitive P2 receptor subtypes in specific structures of the normal rat kidney. Of the P2 receptor subtypes examined, P2X1, P2X2 and P2Y1 receptors were found in the smooth muscle layer of intrarenal vessels. The P2Y1 receptor was also found on glomerular mesangial cells, the brush border membrane of the proximal straight tubule and on peritubular fibroblasts. In the cortex, P2Y4 receptors were found on the tubule epithelium of the proximal convoluted tubule, and P2Y2 receptors on glomerular epithelial cells (podocytes). P2X4 and P2X6 receptors were present throughout the renal tubule epithelium from the proximal tubule to the collecting duct. P2X5 receptors were expressed on medullary collecting duct cells and the apical membrane of the S3 segment of the proximal tubule. Possible functions of these receptor subtypes in normal rat kidney are discussed.     

The structure of the kidney from the freshwater teleost Carassius auratus

Summary The structure of the kidney of the crucian carp (Carassius auratus; a freshwater teleost, Cypriniformes) was studied by means of reconstruction from serial paraffin and semithin sections. In C. auratus, the Wolffian duct traverses the entire kidney. At various levels collecting ducts of different length and thickness join the Wolffian duct at right angles. Each collecting duct accepts a large number of connecting tubules, which are established by the joining of many nephrons. A regular pattern concerning the distribution of nephrons and the fusion of renal tubules is not apparent. Four segments have been distinguished in renal tubules; 1) proximal tubule, 2) distal tubule, 3) connecting tubule and 4) collecting duct. A neck and an intermediate segment are absent. The proximal tubule is established by proximal tubule cells which bear a brush border and have a conspicuous apical cytoplasmic rim containing few cell organelles, ciliated cells, mucous cells and dark cells. In the first part of the proximal tubule the brush border and the apical cytoplasmic rim of proximal tubule cells are well developed. Ciliated cells are interposed between proximal tubule cells, decreasing in number toward the end of this part. In the second part ciliated cells are absent and dark cells are numerous. In the third part the brush border and the apical cytoplasmic rim of proximal tubule cells are scarcely developed. Ciliated cells reapear and increase in number toward the distal tubule. The distal and connecting tubule are similar in epithelial structure. Connecting tubules are joined distal tubules and thus they belong to two or more nephrons. The main cells of distal and connecting tubules contain abundant mitochondria, but have no brush border. The connecting tubule becomes a collecting duct before joining the Wolffian duct. The main cells of collecting ducts are characterized by division of their cytoplasm into a dark apical half and a light basal half.     

The structural organization of the kidney of Typhlonectes compressicaudus (Amphibia, Gymnophiona)

Summary The structural organization of the kidney ofTyphlonectes compressicaudus (Amphibia, Gymnophiona) was studied by light microscopic (LM) examination of serial paraffin and semithin Epon sections. The kidney is slender and quite long and has a mesonephric segmental construction; the excretory duct (Wolffian duct), running along the lateral side of the kidney, segmentally receives the terminal trunks of the collecting duct system. The nephron has the following parts: renal corpuscle, neck segment, proximal tubule, intermediate segment, distal tubule and connecting tubule. The distal tubule is located in a ventromedial (central) zone of the kidney; all other tubular segments lie in a dorsolateral (peripheral) zone. The renal corpuscles are found at the border between these two zones.The renal corpuscle is very large; its urinary pole faces the peripheral zone. A small proportion of neck segments receive either a nephrostomal duct or a blind branch. The proximal tubule is a thick, highly convoluted tubule. The intermediate segment is ciliated and makes a few coils. The distal tubule is composed of three portions: a highly convoluted part in the central zone, subsequently an attachment site with the renal corpuscle and a short postattachment-part. The connecting tubule and the collecting duct have a heterogeneous epithelium consisting of light and dark cells. The collecting duct is distinguished by dilated intercellular spaces. The Wolffian duct has a pseudostratified epithelium.The present study correlates the course and segmentation of the renal tubule ofTyphlonectes. The tubule has three major convolutions. The first occurs in the proximal tubule in the peripheral zone; the second is established by the distal tubule and occurs in the central zone; the third is formed by the connecting tubule and is found in the peripheral zone.Research fellow of the Alexander von Humboldt foundation: home address: Department of Anatomy, Faculty of Medicine, University of Tokyo, Tokyo 113, Japan     

The control of sodium excretion.

The kidneys of a normal man filter approximately 24,000 meq sodium/day, reabsorb about 23,900, and yet can make a 1--2 meq change in 24-h urinary sodium excretion. The control of urinary sodium excretion, therefore, depends, first, on ensuring that the bulk of the sodium is reabsorbed, a function which is carried out in the proximal tubule and ascending loop of Henle. Second, it depends on adjusting the reabsorption of the small quantity of sodium which is delivered into the collecting duct so that the amount excreted in the urine is that required to maintain sodium balance. The bulk reabsorptive mechanisms can be considered as buffers to prevent large fluctuations in the amount of sodium delivered to the collecting duct, thus facilitating the fine adjustments of reabsorption which are made at this site. In conditions other than extreme salt loading or deprivation, changes in sodium reabsorption in the proximal tubule and loop of Henle probably have little, if any, effect on urinary sodium excretion. Sodium reabsorption in the proximal tubule and the collecting duct appears to be influenced by unidentified circulating substances.     

Cell biology of vasopressin-regulated aquaporin-2 trafficking

Whole-body water balance is predominantly controlled by the kidneys, which have the ability to concentrate or dilute the urine in the face of altered fluid and solute intake. Regulated water excretion is controlled by various hormones and signaling molecules, with the antidiuretic hormone arginine vasopressin (AVP) playing an essential role, predominantly via its modulatory effects on the function of the water channel aquaporin-2 (AQP2). The clinical conditions, central and nephrogenic diabetes insipidus, emphasize the importance of the AVP-AQP2 axis. In this article, we summarize the most important and recent studies on AVP-regulated trafficking of AQP2, with focus on the cellular components mediating (1) AQP2 vesicle targeting to the principal cell apical plasma membrane, (2) docking and fusion of AQP2-containing vesicles, (3) regulated removal of AQP2 from the plasma membrane, and (4) posttranslational modifications of AQP2 that control several of these processes. Insight into the molecular mechanisms responsible for regulated AQP2 trafficking is proving to be fundamental for development of novel therapies for water balance disorders.     

小鼠肾集合管发育中主细胞的形态学变化及水通道蛋白-2和-3的表达

目的 观察小鼠肾集合管发育过程中主细胞的形态学变化及水通道蛋白(AQP)-2、-3的表达,探讨AQP-2、-3与小鼠肾集合管主细胞发育的关系及作用.方法 应用透射电镜、免疫组织化学、免疫印迹技术并结合体视学方法,观察并检测胚龄16、18 d 胎鼠及生后1、3、7、14、21 d仔鼠肾集合管发育过程中主细胞的形态学变化及...     

Mouse models and the urinary concentrating mechanism in the new millennium

Our understanding of urinary concentrating and diluting mechanisms at the end of the 20th century was based largely on data from renal micropuncture studies, isolated perfused tubule studies, tissue analysis studies and anatomical studies, combined with mathematical modeling. Despite extensive data, several key questions remained to be answered. With the advent of the 21st century, a new approach, transgenic and knockout mouse technology, is providing critical new information about urinary concentrating processes. The central goal of this review is to summarize findings in transgenic and knockout mice pertinent to our understanding of the urinary concentrating mechanism, focusing chiefly on mice in which expression of specific renal transporters or receptors has been deleted. These include the major renal water channels (aquaporins), urea transporters, ion transporters and channels (NHE3, NKCC2, NCC, ENaC, ROMK, ClC-K1), G protein-coupled receptors (type 2 vasopressin receptor, prostaglandin receptors, endothelin receptors, angiotensin II receptors), and signaling molecules. These studies shed new light on several key questions concerning the urinary concentrating mechanism including: 1) elucidation of the role of water absorption from the descending limb of Henle in countercurrent multiplication, 2) an evaluation of the feasibility of the passive model of Kokko-Rector and Stephenson, 3) explication of the role of inner medullary collecting duct urea transport in water conservation, 4) an evaluation of the role of tubuloglomerular feedback in maintenance of appropriate distal delivery rates for effective regulation of urinary water excretion, and 5) elucidation of the importance of water reabsorption in the connecting tubule versus the collecting duct for maintenance of water balance.     

Close association of aquaporin-2 internalization with caveolin-1

Aquaporin 2 (AQP2) is a membrane water channel protein that traffics between the intracellular membrane compartment and the plasma membrane in a vasopressin-dependent manner in the renal collecting duct cell to control the amount of water reabsorption. We examined the relation between AQP2 internalization from the plasma membrane and caveolin-1, which is a major protein in membrane microdomain caveolae, in Mardin-Darby canine kidney cells expressing human AQP2 (MDCK-hAQP2 cells). Double-immunofluorescence microscopy showed that AQP2 is colocalized with caveolin-1 in the apical plasma membrane by stimulating the intracellular signaling cascade of vasopressin with forskolin. After washing forskolin, both AQP2 and caveolin-1 were internalized to early endosomes and then separately went back to their individual compartments, which are subapical compartments and the apical membrane, respectively.Double-immunogold electron microscopy in ultrathin cryosections confirmed the colocalization of AQP2 with caveolin-1 at caveolar structures on the apical plasma membrane of forskolin-treated cells and the colocalization within the same intracellular vesicles after washing forskolin. A co-immunoprecipitation experiment showed the close interaction between AQP2 and caveolin-1 in forskolin-treated cells and in cells after washing forskolin. These results suggest that a caveolin-1-dependent and possibly caveolar-dependent pathway is a candidate for AQP2 internalization in MDCK cells.     

Renal and hepatic expression of the ammonium transporter proteins, Rh B Glycoprotein and Rh C Glycoprotein

A family of ammonium transporter proteins was recently identified. Members of this family, Rh B Glycoprotein (RhBG) and Rh C Glycoprotein (RhCG) are expressed in the kidney and the liver, important tissues for ammonium metabolism. Immunohistochemical studies demonstrate basolateral RhBG immunoreactivity in the connecting segment (CNT) and collecting ducts, but not in the proximal tubule or the loop of Henle. Colocalization with thiazide sensitive cotransporter and carbonic anhydrase II confirms expression in the CNT, initial collecting tubule (ICT), and throughout the collecting duct. Colocalization with AE1 and pendrin demonstrates expression is greatest in A-type intercalated cells in the cortical collecting duct (CCD), outer medullary collecting duct (OMCD) and inner medullary collecting duct (IMCD), present in the CCD principal cell, and not detectable in either pendrin-positive CCD intercalated cells or in non-intercalated cells in the OMCD and IMCD. RhCG immunoreactivity has a similar axial distribution as RhBG. However, RhCG immunoreactivity is apical, and is detectable in all CCD and outer stripe of OMCD cells. The liver, a second organ involved in ammonia metabolism, also expresses both RhBG and RhCG. Basolateral RhBG immunoreactivity is present in the perivenous hepatocyte, but is not present in either the periportal or mid-zonal hepatocyte. Hepatic RhCG mRNA is expressed at lower levels than RhBG, and RhCG protein is detected in bile duct epithelium. These findings indicate that RhBG and RhCG are involved in at least two organs that transport ammonia, and that they are located in sites where they are likely to mediate important roles in ammonia transport.     

Morphological and morphometrical study of the kidney of the male tropical lizard Tropidurus torquatus

A morphological and morphometrical study of the adult male Tropidurus torquatus kidney was undertaken. The nephron is composed of the following segments: renal corpuscle, neck segment, proximal convoluted tubule, intermediate segment, and distal tubule. The nephron is continued into the collecting duct and sexual segment. A large number of ciliated cells in the intermediate segment, the presence of 2 kinds of cells in the collecting ducts and a well developed permanently retained sexual segment were recorded as special features of this organ. The components of the renal parenchyma had the following relative volumes: proximal convoluted tubule = 56.4%, intermediate segment = 5.1%, distal tubule = 13.0%, collecting duct = 5.2%, and sexual segment = 11.6%.     

Aquaporins in the digestive system

Fluid transfer such as secretion and absorption is one of the major functions of the digestive system. Aquaporins are water channel proteins providing water transfer across the cellular membrane. At least six aquaporin isoforms are expressed in the digestive system. Aquaporin-1 (AQP1) is widely distributed in endothelial cells of capillaries and small vessels as well as in the central lacteals in the small intestine. AQP1 is also present in the duct system in the pancreas, liver, and bile duct. AQP3 is mainly expressed in the epithelia of the upper digestive tract from the oral cavity to the stomach and of the lower digestive tract from the distal colon to the anus. AQP4 is present in the parietal cells of the stomach and in the intestinal epithelia. AQP5 is expressed in acinar cells of the salivary, pyloric, and duodenal glands. AQP8 is expressed in the intestinal epithelia, salivary glands, pancreas, and liver. AQP9 is present in the liver and intestinal goblet cells. Aquaporins have important roles in the digestive system, such as AQP5 in saliva secretion, as shown by the studies on AQP5-null mice. In addition, water transfer across the digestive epithelia seems to occur not only via aquaporins but also via other transporter or channel systems.     

p.R254Q mutation in the aquaporin‐2 water channel causing dominant nephrogenic diabetes insipidus is due to a lack of arginine vasopressin‐induced phosphorylation

Vasopressin regulates human water homeostasis by re‐distributing homotetrameric aquaporin‐2 (AQP2) water channels from intracellular vesicles to the apical membrane of renal principal cells, a process in which phosphorylation of AQP2 at S256 by cAMP‐dependent protein kinase A (PKA) is thought to be essential. Dominant nephrogenic diabetes insipidus (NDI), a disease in which the kidney is unable to concentrate urine in response to vasopressin, is caused by AQP2 gene mutations. Here, we investigated a reported patient case of dominant NDI caused by a novel p.R254Q mutation. Expressed in oocytes, AQP2‐p.R254Q appeared to be a functional water channel, but was impaired in its transport to the cell surface to the same degree as AQP2‐p.S256A, which mimics non‐phosphorylated AQP2. In polarized MDCK cells, AQP2‐p.R254Q was retained and was distributed similarly to that of unstimulated wt‐AQP2 or AQP2‐p.S256A. Upon co‐expression, AQP2‐p.R254Q interacted with, and retained wt‐AQP2 in intracellular vesicles. In contrast to wild‐type AQP2, forskolin did not increase AQP2‐p.R254Q phosphorylation at S256 or its translocation to the apical membrane. Mimicking constitutive phosphorylation in AQP2‐p.R254Q with the p.S256D mutation, however, rescued its apical membrane expression. These date indicate that a lack of S256 phosphorylation is the sole cause of dominant NDI here, and thereby, p.R254Q is a loss of function instead of a gain of function mutation in dominant NDI. © 2009 Wiley‐Liss, Inc.     

Histology of the kidney and urinary bladder of Siphonops annulatus (Amphibia-Gymnophiona)

The histology of the kidney and urinary bladder of Siphonops annulatus was studied by light microscopy in semithin sections of tissue embedded in hydrophilic resin. The kidney's nephron comprises the renal corpuscle, neck segment, proximal tubule, intermediate segment, distal tubule and collecting tubule. Nephrostomes are present. This structure, the neck segment, and intermediate tubules present long cilia, and probably play important roles in the propulsion of the peritoneal fluid and glomerular filtrate. The proximal tubule cells possess loosely packed microvilli and contain abundant polymorphic granules and vesicles that assume the aspect of lysosomes in different stages of intracellular digestion. The distal tubules are characterized by large, vertically disposed mitochondria assuming the aspect of ions transporting cells. The urinary bladder is lined with a transitional epithelium, whose aspect varies according to the quantity of urine.     

硒酸锌金属自显影技术检测游离锌离子在小鼠肾脏的分布

目的研究游离锌离子在小鼠肾脏的定位分布。方法应用硒酸锌金属自显影技术(ZnSeAMG)检测小鼠肾脏内的游离锌离子分布。结果游离锌离子在肾脏内分布广泛,皮质中有大量AMG反应阳性颗粒,髓质中的AMG阳性颗粒较少。其中,近曲小管、远曲小管、近直小管和远直小管上皮细胞近腔侧均分布有大量的棕黑色AMG阳性颗粒,肾小体、细段和集合管上皮细胞中AMG阳性颗粒较少。结论小鼠肾脏内含有丰富的游离锌离子,锌离子可能参与肾脏的功能。