Roles of aquaporins in kidney revealed by transgenic mice

Transgenic mouse models of aquaporin (AQP) deletion and mutation have been instructive in elucidating the role of AQPs in renal physiology. Mice lacking AQP1 are unable to concentrate their urine because of low water permeability in the proximal tubule, thin descending limb of Henle, and outer medullary descending vasa recta, resulting in defective near-isosmolar fluid absorption in the proximal tubule and defective countercurrent multiplication. Mice lacking functional AQP2, AQP3, or AQP4 manifest various degrees of nephrogenic diabetes insipidus resulting from reduced collecting duct water permeability. Mice lacking AQP7 and AQP8 can concentrate their urine fully, although AQP7 null mice manifest an interesting defect in glycerol reabsorption. Two unexpected renal phenotypes of AQP null mice have been discovered recently, including defective proximal tubule cell migration in AQP1 deficiency, and cystic renal disease in AQP11 deficiency. AQPs thus are important in several aspects of the urinary concentrating mechanism and in functions unrelated to tubular fluid transport. The mouse phenotype data suggest the renal AQPs as targets for the development of aquaretics and potentially for therapy of cystic renal disease and acute renal injury.     

Lessons on renal physiology from transgenic mice lacking aquaporin water channels

Several aquaporin-type water channels are expressed in kidney: AQP1 in the proximal tubule, thin descending limb of Henle, and vasa recta; AQP2, AQP3, and AQP4 in the collecting duct; AQP6 in the papilla; and AQP7 in the proximal tubule. AQP2 is the vasopressin-regulated water channel that is important in hereditary and acquired diseases affecting urine-concentrating ability. It has been difficult to establish the roles of the other aquaporins in renal physiology because suitable aquaporin inhibitors are not available. One approach to the problem has been to generate and analyze transgenic knockout mice in which individual aquaporins have been selectively deleted by targeted gene disruption. Phenotype analysis of kidney and extrarenal function in knockout mice has been very informative in defining the role of aquaporins in organ physiology and addressing basic questions regarding the route of transepithelial water transport and the mechanism of near iso-osmolar fluid reabsorption. This article describes new renal physiologic insights revealed by phenotype analysis of aquaporin-knockout mice and the prospects for further basic and clinical developments.     

Renal concentrating and diluting function in deficiency of specific aquaporin genes

Aquaporins (AQP) are water-transporting proteins expressed in many fluid-transporting epithelia and endothelia. In kidney, AQP1 is expressed in plasma membranes of proximal tubule, thin descending limb of Henle and descending vasa recta, AQP2 in collecting duct luminal membrane, AQP3 and AQP4 in collecting duct basolateral membrane, AQP6 in intercalated cells, and AQP7 in the S3 segment of proximal tubule. Human mutations in AQP2 cause hereditary non-X-linked nephrogenic diabetes insipidus. Transgenic mice lacking the renal aquaporins have been useful in defining their role. Mice deficient in AQP1 are polyuric and unable to form a concentrated urine because of defective proximal tubule fluid absorption and countercurrent multiplication. Mice lacking AQP3 are markedly polyuric due to low water permeability across the cortical and outer medullary collecting duct. However, mice lacking AQP4, which is expressed mainly in inner medullary collecting duct, manifest only a mild defect in maximum urinary concentrating ability. The aquaporin null mice have normal urinary diluting ability. From many renal and extrarenal phenotype studies of aquaporin null mice, we conclude that aquaporins are important for rapid near-isosmolar transepithelial fluid absorption/secretion and for rapid vectorial water movement driven by osmotic gradients. The renal phenotype in aquaporin null mice suggests the utility of aquaporin blockers as novel aquaretic-diuretic agents.     

Role of aquaporin water channels in kidney function studied using transgenic mice

Several aquaporin (AQP) water transporting proteins are expressed in mammalian kidney: AQP1 in plasma membranes of proximal tubule, thin descending limb of Henle, and descending vasa recta; AQP2 in collecting duct luminal membrane; AQP3 and AQP4 in collecting duct basolateral membrane; AQP6 in intercalated cells; and AQP7 in the S3 segment of proximal tubule. To define the role of aquaporins in renal physiology, we have generated and characterized transgenic null mice deficient in AQP1, AQP3, and AQP4, individually and in combinations, as well as AQP2 mutant mice, in which the T126M mutation causing human nephrogenic diabetes insipidus was introduced. AQP1-deficient mice are polyuric and unable to concentrate their urine in response to water deprivation or vasopressin administration. AQP1 deletion greatly reduces osmotic water permeability in proximal tubule, thin descending limb of Henle, and descending vasa recta, resulting in defective proximal tubule fluid absorption and medullary countercurrent exchange. Mice lacking AQP3 have low basolateral membrane water permeability in cortical collecting duct and excrete large quantities of dilute urine. Mice lacking AQP4 have low water permeability in inner medullary collecting duct, but manifest only a mild defect in maximum urinary concentrating ability. These data, taken together with phenotype analyses of brain, lung, and gastrointestinal organs, support the paradigm that aquaporins facilitate rapid near-isosmolar transepithelial fluid absorption/secretion, as well as rapid vectorial water movement driven by osmotic gradients. The renal phenotype data in aquaporin knockout mice suggests the utility of aquaporin blockers as novel aquaretic-diuretic agents. Received: March 19, 2001 / Accepted: March 22, 2001     

Urea and renal function in the 21st century: insights from knockout mice

Since the turn of the 21st century, gene knockout mice have been created for all major urea transporters that are expressed in the kidney: the collecting duct urea transporters UT-A1 and UT-A3, the descending thin limb isoform UT-A2, and the descending vasa recta isoform UT-B. This article discusses the new insights that the results from studies in these mice have produced in the understanding of the role of urea in the urinary concentrating mechanism and kidney function. Following is a summary of the major findings: (1) Urea accumulation in the inner medullary interstitium depends on rapid transport of urea from the inner medullary collecting duct (IMCD) lumen via UT-A1 and/or UT-A3; (2) as proposed by Robert Berliner and colleagues in the 1950s, the role of IMCD urea transporters in water conservation is to prevent a urea-induced osmotic diuresis; (3) the absence of IMCD urea transport does not prevent the concentration of NaCl in the inner medulla, contrary to what would be predicted from the passive countercurrent multiplier mechanism in the form proposed by Kokko and Rector and Stephenson; (4) deletion of UT-B (vasa recta isoform) has a much greater effect on urinary concentration than deletion of UT-A2 (descending limb isoform), suggesting that the recycling of urea between the vasa recta and the renal tubules quantitatively is less important than classic countercurrent exchange; and (5) urea reabsorption from the IMCD and the process of urea recycling are not important elements of the mechanism of protein-induced increases in GFR. In addition, the clinical relevance of these studies is discussed, and it is suggested that inhibitors that specifically target collecting duct urea transporters have the potential for clinical use as potassium-sparing diuretics that function by creation of urea-dependent osmotic diuresis.     

Urea transporters and renal function: lessons from knockout mice

PURPOSE OF REVIEW: Gene knockout mice have been created for the collecting duct urea transporters UT-A1 and UT-A3, the descending thin-limb urea transporter UT-A2 and the descending vasa recta isoform, UT-B. In this brief review, the new insights in our understanding of the role of urea in the urinary concentrating mechanism and kidney function resulting from studies in these mice are discussed. RECENT FINDINGS: The major findings in studies on urea transporter knockout mice are as follows: rapid transport of urea from the inner medulla collecting duct lumen via UT-A1 or UT-A3 is essential for urea accumulation in the inner medullary interstitium; inner medulla collecting duct urea transporters are essential in water conservation by preventing urea-induced osmotic diuresis; an absence of inner medulla collecting duct urea transport does not prevent the concentration of sodium chloride in the inner medulla interstitium; deletion of the vasa recta isoform UT-B has a much greater effect on urinary concentration than deleting the descending limb isoform UT-A2. SUMMARY: Multiple urea transport mechanisms within the kidney are essential for producing maximally concentrated urine.     

Discovery of aquaporins: a breakthrough in research on renal water transport

Several membranes of the kidney are highly water permeable, thereby enabling this organ to retain large quantities of water. Recently, the molecular identification of water channels responsible for this high water permeability has finally been accomplished. At present, four distinct renal water channels have been identified, all members of the family of major intrinsic proteins. Aquaporin 1 (AQP1), aquaporin 2 (AQP2) and the mercury-insensitive water channel (MIWC) are water-selective channel proteins, whereas the fourth, referred to as aquaporin 3 (AQP3), permits transport of urea and glycerol as well. Furthermore, a putative renal water channel (WCH3) has been found. AQP1 is expressed in apical and basolateral membranes of proximal tubules and descending limbs of Henle, AQP2 predominantly in apical membranes of principal and inner medullary collecting duct cells and AQP3 in basolateral membranes of kidney collecting duct cells. MIWC is expressed in the inner medulla of the kidney and has been suggested to be localised in the vasa recta. The human genes encoding AQP1 and AQP2 have been cloned, permitting deduction of their amino acid sequence, prediction of their two-dimensional structure by hydropathy analysis, speculations on their way of functioning and DNA analysis in patients with diseases possibly caused by mutant aquaporins. Mutations in the AQP1 gene were recently detected in clinically normal individuals, a finding which contradicts the presumed vital importance of this protein. Mutations in the AQP2 gene were shown to cause autosomal recessive nephrogenic diabetes insipidus. The renal unresponsiveness to arginine vasopressin, which characterises this disease, is in accordance with the assumption that AQP2 is the effector protein of the renal vasopressin pathway. The influence of amino acid substitutions on the functioning of AQP1 and 2 was demonstrated by in vitro expression studies in oocytes of the toadXenopus laevis. Future research on renal water transport will focus on the search for other aquaporins, structure-function relationship of aquaporins, the development of aquaporin inhibitors and their possible use as diuretics, and further elucidation of the renal vasopressin pathway.     

Aquaporin Water Channels in the Mammary Gland: From Physiology to Pathophysiology and Neoplasia

Aquaporins are membrane proteins that play fundamental roles in water and small solute transport across epithelial and endothelial barriers. Recent studies suggest that several aquaporin proteins are present in the mammary gland. Immunohistochemical techniques have confirmed the presence of aquaporin 1 (AQP1) and AQP3 water channels in rat, mouse, bovine and human mammary glands. Studies suggest that in addition to AQP1 and AQP3 AQP4, AQP5 and AQP7 proteins are expressed in different locations in the mammary gland. Aquaporins play key roles in tumor biology and are involved in cell growth, migration and formation of ascites via increased water permeability of micro-vessels. Emerging evidence suggests that expression of these proteins is altered in mammary tumors and in breast cancer cell lines although it is not yet clear whether this is a cause or a consequence of neoplastic development. This review analyzes the expression and potential functional roles of aquaporin water channels in the mammary gland. The physiological mechanisms involved in the transport of water and small solutes across mammary endothelial and epithelial barriers are discussed in the context of milk production and lactation. This paper also reviews papers from the recent cancer literature that implicate aquaporins in mammary neoplasia.     

水通道蛋白在多囊肾病小鼠肾囊泡上皮细胞的表达与调控

目的 探讨水通道蛋白(AQP)在多囊肾病囊泡上皮细胞的表达和调控。 方法 采用免疫荧光染色和Western印迹法分别检测不同亚型的水通道蛋白AQP1、AQP2、AQP3和AQP4在小鼠常染色体隐性遗传病jck多囊肾小鼠肾脏的表达定位和表达调控。 结果 8周龄jck纯合子小鼠的肾脏占体质量的百分率约是同窝野生型小鼠肾脏的4倍,肾脏组织出现多发、大小不一囊泡,囊泡上皮细胞呈扁平状,肾脏间质可见纤维化。jck小鼠血尿素水平为（42.6±6.7） mmol/L,约是野生型小鼠血尿素水平[（8.4±1.9） mmol/L]的5倍（P < 0.01）。免疫荧光定位分析结果表明AQP1 在近曲小管上皮细胞顶膜和基底膜表达,也表达于髓袢降支细段和直小血管降支,肾囊泡上皮细胞未见AQP1表达。AQP2在集合管和肾囊泡上皮细胞顶膜表达,AQP3和AQP4在集合管和囊泡上皮细胞基底膜表达。Western印迹分析结果表明,jck肾脏AQP2、AQP3和AQP4蛋白表达水平与野生型肾脏相似,但AQP1在jck肾脏的表达水平显著低于其在野生型肾脏的表达水平（P < 0.01）。 结论 jck多囊肾小鼠肾囊泡上皮表达AQP2、AQP3和AQP4,提示肾囊泡来源于肾集合管,水通道蛋白可能在肾囊泡生长过程中起重要作用。     

Aquaporin-1 facilitates epithelial cell migration in kidney proximal tubule

Aquaporin-1 (AQP1) is the principal water-transporting protein in cell plasma membranes in kidney proximal tubule, where it facilitates transepithelial water transport. Here, a novel role for AQP1 in kidney involving the migration of proximal tubule cells is reported. Migration was compared in primary cultures of proximal tubule cells from wild-type and AQP1 null mice. Cell cultures from AQP1 null mice were indistinguishable from those of wild-type mice in their appearance, growth/proliferation, and adhesiveness, although, as expected, they had reduced plasma membrane water permeability. Migration of AQP1-deficient cells was reduced by >50% compared with wild-type cells, as measured in a Boyden chamber in the presence of a chemotactic stimulus. Comparable slowing of migration of AQP1-deficient cells was also found in an in vitro scratch assay of wound healing, with reduced appearance of lamella-like membrane protrusions at the cell leading edge. Adenoviral-mediated expression of AQP1 in the AQP1-deficient cells, which increased their water permeability to that of wild-type cells, corrected their migration defect. The potential relevance of these in vitro findings to the intact kidney was tested in an in vivo model of acute tubular injury caused by 30 min of renal artery occlusion. At 3 to 5 d after ischemia-reperfusion, kidneys in AQP1 null mice showed remarkably greater tubular injury and cellular actin disorganization than kidneys in wild-type mice. These results provide evidence for the involvement of AQP1 in migration of proximal tubule cells and possibly in the response of the proximal tubule to injury.     

Anti-B7-1 blocks mononuclear cell adherence in vasa recta after ischemia

BACKGROUND: Blocking the costimulatory pathway by CTLA-4 Ig, reactive with both B7-1 and B7-2 costimulatory molecules, protects the kidney during acute ischemia/reperfusion injury. This study investigated whether and how B7-1 and/or B7-2 proteins are involved in renal ischemia/reperfusion injury (IRI). METHODS: Uninephrectomized rats were submitted to warm renal ischemia (30 min) and received control monoclonal antibody (mAb; 17E3), anti-B7-1 (3H5), anti-B7-2 (24F), a combination of anti-B7-1/B7-2, or CTLA-4 Ig. Renal function, morphology, and the kinetics of inflammatory cells were studied for a ten-day period. Binding sites of the injected antibodies were detected by secondary staining with anti-mouse Ab. RESULTS: Compared with controls, acute renal failure (ARF) in the anti-B7-1 group was attenuated both functionally and morphologically. Anti-B7-1/B7-2 and CTLA-4 Ig also were protective in IRI. ARF was not altered by anti-B7-2 treatment. Two hours after reperfusion, B7-1 was expressed along the endothelial cells of the ascending vasa recta. Expression of B7-1 increased over time during the first 24 hours and decreased thereafter. Two hours after reperfusion, adherence/accumulation of T cells and monocytes/macrophages was found in the vasa recta of the ischemic kidney. Anti-B7-1-treated animals had fewer T cells and monocytes/macrophages in the vasa recta compared with controls. Leukocyte accumulation in these vessels after anti-B7-2 treatment was not different from IRI controls. CONCLUSION: These observations strongly support the key role of the B7-1 protein in the protection of renal IRI through inhibition of T cell and monocyte adherence at the level of the ascending vasa recta.     

T cells as mediators in renal ischemia/reperfusion injury

Inflammation has been established to contribute substantially to the pathogenesis of ischemia/reperfusion (I/R) with a central role for particular cells, adhesion molecules, and cytokines. Until recently, most of the research trying to unravel the pathogenesis of I/R injury has been focused on the role of neutrophils. However, recent studies have brought evidence that T cells and macrophages are also important leukocyte mediators of renal and extrarenal (liver) I/R injury. In vivo depletion of CD4+ cells but not CD8+ cells in wild-type mice was protective in I/R of the kidney. A marked preservation of liver function was also found after I/R in T-cell deficient athymic mice. Blocking the b130/CD28 costimulatory pathway by CTLA-4 Ig (recombinant fusion protein) ameliorated renal dysfunction and decreased mononuclear cell infiltration in I/R of the kidney. b130-1 expression was found limited to the membrane of the endothelial cells of the ascending vasa recta, resulting in trapping of CD28-expressing CD4 T cells. This trapping of leukocytes results in the upstream congestion in the ascending arterial vasa recta, generating the since more than 150 years described medullary vascular congestion of the kidney soon after ischemic injury. It seems worthwhile to study a combination therapy using anti-inflammatory/anti-adhesion molecules in the early phase of I/R.     

Physiological importance of aquaporins: lessons from knockout mice

The phenotype analysis of transgenic mice deficient in specific aquaporin water channels has provided new insights into the role of aquaporins in organ physiology. AQP1-deficient mice are polyuric and are unable to concentrate their urine in response to water deprivation or vasopressin administration. AQP1 deletion reduces osmotic water permeability in the proximal tubule, thin descending limb of Henle and vasa recta, resulting in defective proximal tubule fluid absorption and medullary countercurrent exchange. Mice lacking AQP3, a basolateral membrane water channel expressed mainly in the cortical collecting duct, are remarkably polyuric but are able to generate a partly concentrated urine after water deprivation. In contrast, mice lacking AQP4, a water channel expressed mainly in the inner medullary collecting duct, manifest only a mild defect in maximum urinary concentrating ability. These data, together with phenotype analyses of the brain, lung, salivary gland, and gastrointestinal organs, support the paradigm that aquaporins can facilitate near-isosmolar transepithelial fluid absorption/secretion as well as rapid vectorial water movement driven by osmotic gradients. The phenotype data obtained from aquaporin knockout mice suggest the utility of aquaporin blockers as novel diuretic agents.     

P-selectin-dependent macrophage migration into the tubulointerstitium in unilateral ureteral obstruction

BACKGROUND: Interstitial infiltration of macrophages (M?) is one of the main causal factors for the tubulointerstitial injury. However, precise mechanisms of M? infiltration into tubulointerstitium have not been fully explored. The purposes of this study were to assess the role of selectins in the acute infiltration of M? in rats with unilateral ureteral obstruction (UUO) and to evaluate the role of vasa recta, that is, whether they facilitate massive influx of M? into the interstitium by functioning as specialized vessels. METHODS: To evaluate the role of selectins in M? infiltration into tubulointerstitium, the expression of selectins and L-selectin ligands was examined by immunohistochemistry and immunoelectron microscopy. The functional role of P-selectin in vasa recta was studied by Stamper-Woodruff assay, in vivo p-M? migration assay and in vivo blocking experiments with the monoclonal antibody (mAb) ARP2-4. RESULTS: Selective expression of P-selectin was detected in vasa recta as early as one hour after UUO, and the expression increased thereafter for 96 hours. In contrast, endothelial expression of L-selectin ligands and E-selectin were not detectable. In the Stamper-Woodruff assay on kidney sections of rats with UUO, the adhesion of isolated rat peritoneal M? (p-M?) to vasa recta was significantly inhibited by the mAb ARP2-4 (P-selectin blocker; P < 0.01), but not by mAb ARE-5 (E-selectin blocker) or rLECIg (rat L-selectin chimeric protein). In the in vivo transfer experiments with fluorescein-labeled p-M? into rats 48 hours after UUO, labeled p-M? had accumulated around vasa recta at three minutes and had infiltrated predominantly into the outer medulla at 180 minutes. The number of labeled p-M? was reduced when the rats were pretreated with ARP2-4 (P < 0.01). Finally, ARP2-4 (10 mg/kg), injected 15 minutes before UUO, reduced the number of infiltrated M? (P < 0.01). CONCLUSION: The results suggest that vasa recta, which express P-selectin, contribute to massive infiltration of M? into the interstitium by functioning as specialized post-capillary venules.     

Divergent regulation of circulating and intrarenal renin-angiotensin systems in response to long-term blockade

BACKGROUND/AIMS: Long-term treatment with angiotensin-converting enzyme (ACE) inhibitors or angiotensin (Ang) II type I (AT(1)) receptor blockers can improve kidney function and attenuate the progressive decline in kidney function associated with age. In this study in Wistar rats medicated for 22 months, we determined the effects of enalapril (10 mg/kg/day) and losartan (30 mg/kg/day) treatment, in comparison with vehicle (tap water), on renal AngII receptor density and circulating and urinary components of the renin-angiotensin system (RAS). METHODS: Kidney sections were incubated with [(125)I-sarcosine(1)-threonine(8)]AngII (0.6 nM) for Ang receptor density, and Ang peptides were determined using radioimmunoassays. RESULTS: Receptor density was approximately 50% higher in vasa recta, glomeruli, and tubulointerstitium in enalapril-treated rats and lower in vasa recta and glomeruli in losartan-treated relative to vehicle-treated rats. Losartan and enalapril treatment elevated plasma levels of AngI and Ang-(1-7) while AngII increased only in losartan-treated rats. In contrast, both treatments were associated with a reduction in urinary excretion of all three Ang peptides as compared with control rats. CONCLUSION: The reduction in urinary Ang peptides with losartan and enalapril treatment suggests that blockade of intrarenal AngII may be an important mechanism underlying the renoprotection seen with such treatments.     

Toll-like receptor 4 regulates early endothelial activation during ischemic acute kidney injury

Ischemic acute kidney injury (AKI) triggers an inflammatory response which exacerbates injury that requires increased expression of endothelial adhesion molecules. To study this further, we used in situ hybridization, immunohistology, and isolated endothelial cells, and found increased Toll-like receptor 4 (TLR4) expression on endothelial cells of the vasa rectae of the inner stripe of the outer medulla of the kidney 4?h after reperfusion. This increase was probably due to reactive oxygen species, known to be generated early during ischemic AKI, because the addition of hydrogen peroxide increased TLR4 expression in MS1 microvascular endothelial cells in vitro. Endothelial TLR4 may regulate adhesion molecule (CD54 and CD62E) expression as they were increased on endothelia of wild-type but not TLR4 knockout mice in vivo. Further, the addition of high-mobility group protein B1, a TLR4 ligand released by injured cells, increased adhesion molecule expression on endothelia isolated from wild-type but not TLR4 knockout mice. TLR4 was localized to proximal tubules in the cortex and outer medulla after 24?h of reperfusion. Thus, at least two different cell types express TLR4, each of which contributes to renal injury by temporally different mechanisms during ischemic AKI.     

Aquaporin 2 promotes cell migration and epithelial morphogenesis

The aquaporin 2 (AQP2) water channel, expressed in kidney collecting ducts, contributes critically to water homeostasis in mammals. Animals lacking or having significantly reduced levels of AQP2, however, have not only urinary concentrating abnormalities but also renal tubular defects that lead to neonatal mortality from renal failure. Here, we show that AQP2 is not only a water channel but also an integrin-binding membrane protein that promotes cell migration and epithelial morphogenesis. AQP2 expression modulates the trafficking and internalization of integrin β1, facilitating its turnover at focal adhesions. In vitro, disturbing the interaction between AQP2 and integrin β1 by mutating the RGD motif led to reduced endocytosis, retention of integrin β1 at the cell surface, and defective cell migration and tubulogenesis. Similarly, in vivo, AQP2-null mice exhibited significant retention of integrin β1 at the basolateral membrane and had tubular abnormalities. In summary, these data suggest that the water channel AQP2 interacts with integrins to promote renal epithelial cell migration, contributing to the structural and functional integrity of the mammalian kidney.     

Dissecting the roles of aquaporins in renal pathophysiology using transgenic mice

Transgenic mice lacking renal aquaporins (AQPs), or containing mutated AQPs, have been useful in confirming anticipated AQP functions in renal physiology and in discovering new functions. Mice lacking AQPs 1-4 manifest defects in urinary concentrating ability to different extents. Mechanistic studies have confirmed the involvement of AQP1 in near-isosmolar fluid absorption in the proximal tubule, and in countercurrent multiplication and exchange mechanisms that produce medullary hypertonicity in the antidiuretic kidney. Deletion of AQPs 2-4 impairs urinary concentrating ability by reduction of transcellular water permeability in the collecting duct. Recently created transgenic mouse models of nephrogenic diabetes insipidus produced by AQP2 gene mutation offer exciting possibilities to test new drug therapies. Several unanticipated AQP functions in kidney have been discovered recently that are unrelated to their role in transcellular water transport. There is evidence for involvement of AQP1 in kidney cell migration after renal injury, of AQP7 in renal glycerol clearance, of AQP11 in prevention of renal cystic disease, and possibly of AQP3 in regulation of collecting duct cell proliferation. Future work in renal AQPs will focus on mechanisms responsible for these non-fluid-transporting functions, and on the development of small-molecule AQP inhibitors for use as aquaretic-type diuretics.