Aquaporin expression in the human intervertebral disc.

The nucleus pulposus (NP) of the human intervertebral disc (IVD) is a hyperosmotic tissue that is subjected to daily dynamic compressive loads. In order to survive within this environment the resident chondrocyte-like cells must be able to control their cell volume, whilst also controlling the anabolism and catabolism of their extra-cellular matrix. Recent studies have demonstrated expression of a range of bi-directional, transmembrane water and solute transporters, named aquaporins (AQPs), within chondrocytes of articular cartilage. The aim of this study was to use immunohistochemsitry to investigate the expression of aquaporins 1, 2 and 3 within the human IVD. Results demonstrated expression of both AQP-1 and -3 by cells within the NP and inner annulus fibrosus (AF), while outer AF cells lacked expression of AQP-1 and showed very low numbers of AQP-3 immunopositive cells. Cells from all regions were negative for AQP-2. Therefore this study demonstrates similarities in the phenotype of NP cells and articular chondrocytes, which may be due to similarities in tissue osmolarity and mechanobiology. The decrease in expression of AQPs from the NP to the outer AF may signify changes in cellular phenotype in response to differences in mechanbiology, osmolarity and hydration between the gelatinous NP and the fibrous AF.

Hypokalemia-induced downregulation of aquaporin-2 water channel expression in rat kidney medulla and cortex.

Prolonged hypokalemia causes vasopressin-resistant polyuria. We have recently shown that another cause of severe polyuria, chronic lithium therapy, is associated with decreased aquaporin-2 (AQP2) water channel expression (Marples, D., S. Christensen, E.I. Christensen, P.D. Ottosen, and S. Nielsen, 1995. J. Clin. Invest., 95: 1838-1845). Consequently, we studied the effect in rats of 11 days' potassium deprivation on urine production and AQP2 expression and distribution. Membrane fractions were prepared from one kidney, while the contralateral kidney was perfusion-fixed for immunocytochemistry. Immunoblotting and densitometry revealed a decrease in AQP2 levels to 27+/-3.4% of control levels (n=11, P<0.001) in inner medulla, and 34+/-15% of controls (n=5, P<0.05) in cortex. Urine production increased in parallel, from 11+/-1.4 to 30+/-4.4 ml/day (n=11, P<0.01). After return to a potassium-containing diet both urine output and AQP2 labels normalized within 7 d. Immunocytochemistry confirmed decreased AQP2 labeling in principal cells of both inner medullary and cortical collecting ducts. AQP2 labeling was predominantly associated with the apical plasma membrane and intracellular vesicles. Lithium treatment for 24 d caused a more extensive reduction of AQP2 levels, to 4+/-1% of control levels in the inner medulla and 4+/-2% in cortex, in association with severe polyuria. The similar degree of downregulation in medulla and cortex suggests that interstitial tonicity is not the major factor in the regulation of AQP2 expression. Consistent with this furosemide treatment did not alter AQP2 levels. In summary,hypokalemia, like lithium treatment, results in a decrease in AQP2 expression in rat collecting ducts, in parallel with the development of polyuria, and the degree of downregulation is consistent with the level of polyuria induced, supporting the view that there is a causative link.

Lithium-induced downregulation of aquaporin-2 water channel expression in rat kidney medulla.

Lithium, a widely used treatment for bipolar affective disorders, often causes nephrogenic diabetes insipidus. The effect of chronic lithium therapy on the expression of the vasopressin-regulated water channel Aquaporin-2 (AQP2) in rat kidney was examined. Membranes were prepared from inner medulla of one kidney from each rat, while the contralateral one was fixed for immunofluorescence and immunoelectronmicroscopy. Immunoblotting revealed that lithium treatment reduced AQP2 expression dramatically, to 31 +/- 8% after 10 d and to 4 +/- 1% after 25 d, coincident with development of severe polyuria. Immunofluorescence and immunogold quantitation confirmed the lithium-induced decrease in AQP2 expression (from 11.2 +/- 1.0 to 1.1 +/- 0.2 particles/microns 2). The downregulation was only partly reversed by return to lithium-free diet for 1 wk (40 +/- 8% of control). Furthermore, immunoblotting and immunogold quantitation revealed that 2 d of thirsting or 7 d of dDAVP treatment, in the continued presence of lithium, increased AQP2 expression by six- and threefold, respectively, coincident with increased urinary osmolality. Thirsting increased AQP2 immunolabeling mainly of vesicles, whereas dDAVP caused accumulation of AQP2 predominantly in the subapical region and plasma membrane. Thus, lithium causes marked downregulation of AQP2 expression, only partially reversed by cessation of therapy, thirsting or dDAVP treatment, consistent with clinical observations of slow recovery from lithium-induced urinary concentrating defects.

Expression of VAMP-2-like protein in kidney collecting duct intracellular vesicles. Colocalization with Aquaporin-2 water channels.

Body water balance is controlled by vasopressin, which regulates Aquaporin-2 (AQP2) water channels in kidney collecting duct cells by vesicular trafficking between intracellular vesicles and the plasma membrane. To examine the molecular apparatus involved in vesicle trafficking and vasopressin regulation of AQP2 in collecting duct cells, we tested if targeting proteins expressed in the synaptic vesicles, namely vesicle-associated membrane proteins 1 and 2 (VAMP1 and 2), are expressed in kidney collecting duct. Immunoblotting revealed specific labeling of VAMP2 (18-kD band) but not VAMP1 in membrane fractions prepared from kidney inner medulla. Controls using preadsorbed antibody or preimmune serum were negative. Bands of identical molecular size were detected in immunoblots of brain membrane vesicles and purified synaptic vesicles. VAMP2 in kidney membranes was cleaved by tetanus toxin, revealing a tetanus toxin-sensitive VAMP homologue. Similarly, tetanus toxin cleaved VAMP2 in synaptic vesicles. In kidney inner medulla, VAMP2 was predominantly expressed in the membrane fraction enriched for intracellular vesicles, with little or no VAMP2 in the plasma membrane enriched fraction. This was confirmed by immunocytochemistry using semithin cryosections, which showed mainly vesicular labeling in collecting duct principal cells, with no labeling of intercalated cells. VAMP2 immunolabeling colocalized with AQP2 labeling in intracellular vesicles, as determined by immunoelectron microscopy after double immunolabeling of isolated vesicles. Quantitative analysis of 1,310 vesicles revealed a highly significant association of both AQP2 and VAMP2 in the same vesicles (P < 0.0001). Furthermore, the presence of AQP2 in vesicles immunoisolated with anti-VAMP2 antibodies was confirmed by immunoblotting. In conclusion, VAMP2, a component of the neuronal SNARE complex, is expressed in vesicles carrying AQP2, suggesting a role in vasopressin-regulated vesicle trafficking of AQP2 water channels.

Distribution of AQP2 and AQP3 water channels in human tissue microarrays.

The objective of this investigation was to use semi-quantitative immunohistochemistry to determine the distribution and expression levels of AQP2 and AQP3 proteins in normal human Tissue MicroArrays. Expression of the vasopressin regulated AQP2 was observed in a limited number of tissues. AQP2 was prominent in the apical and subapical plasma membranes of cortical and medullary renal collecting ducts. Surprisingly, weak AQP2 immunoreactivity was also noted in pancreatic islets, fallopian tubes and peripheral nerves. AQP2 was also localized to selected parts of the central nervous system (ependymal cell layer, subcortical white matter, hippocampus, spinal cord) and selected cells in the gastrointestinal system (antral and oxyntic gastric mucosa, small intestine and colon). These findings corroborate the restricted tissue distribution of AQP2. AQP3 was strongly expressed in many of the human tissues examined particularly in basolateral membranes of the distal nephron (medullary collecting ducts), distal colon, upper airway epithelia, transitional epithelium of the urinary bladder, tracheal, bronchial and nasopharyngeal epithelium, stratified squamous epithelial cells of the esophagus, and anus. AQP3 was moderately expressed in basolateral membranes of prostatic tubuloalveolar epithelium, pancreatic ducts, uterine endometrium, choroid plexus, articular chondrocytes, subchondral osteoblasts and synovium. Low AQP3 levels were also detected in skeletal muscle, cardiac muscle, gastric pits, seminiferous tubules, lymphoid vessels, salivary and endocrine glands, amniotic membranes, placenta and ovary. The abundance of basolateral AQP3 in epithelial tissues and its expression in many non-epithelial cells suggests that this aquaglyceroporin is a major participant in barrier hydration and water and osmolyte homeostasis in the human body.

Severe reversible azotemia from captopril therapy. Report of three cases and review of the literature.

We saw three cases of severe reversible azotemia secondary to captopril therapy in hypertension. All patients had extensive peripheral vascular disease involving the renal arteries, and two patients (patients 2 and 3) had high levels of peripheral plasma renin activity. The azotemia occurred approximately two weeks after exposure to captopril, and fever, a maculopapular pruritic cutaneous rash, and eosinophilia developed in two patients (patients 2 and 3). The cause of the azotemia in our patients is not clearly known, since renal biopsies were not performed. The most likely cause for the azotemia was volume contraction with reduction in the glomerular filtration rate, although a direct insult to the kidney could not be excluded.

Distribution of the muscarinic K+ channel proteins Kir3.1 and Kir3.4 in the ventricle, atrium, and sinoatrial node of heart.

The functionally important effects on the heart of ACh released from vagal nerves are principally mediated by the muscarinic K+ channel. The aim of this study was to determine the abundance and cellular location of the muscarinic K+ channel subunits Kir3.1 and Kir3.4 in different regions of heart. Western blotting showed a very low abundance of Kir3.1 in rat ventricle, although Kir3.1 was undetectable in guinea pig and ferret ventricle. Although immunofluorescence on tissue sections showed no labeling of Kir3.1 in rat, guinea pig, and ferret ventricle and Kir3.4 in rat ventricle, immunofluorescence on single ventricular cells from rat showed labeling in t-tubules of both Kir3.1 and Kir3.4. Kir3.1 was abundant in the atrium of the three species, as shown by Western blotting and immunofluorescence, and Kir3.4 was abundant in the atrium of rat, as shown by immunofluorescence. Immunofluorescence showed Kir3.1 expression in SA node from the three species and Kir3.4 expression in the SA node from rat. The muscarinic K+ channel is activated by ACh via the m2 muscarinic receptor and, in atrium and SA node from ferret, Kir3.1 labeling was co-localized with m2 muscarinic receptor labeling throughout the outer cell membrane.

Presence of the Kv1.5 K(+) channel in the sinoatrial node.

The aim of this study was to establish, using immunolabeling, whether the Kv1.5 K(+) channel is present in the pacemaker of the heart, the sinoatrial (SA) node. In the atrial muscle surrounding the SA node and in the SA node itself (from guinea pig and ferret), Western blotting analysis showed a major band of the expected molecular weight, approximately 64 kD. Confocal microscopy and immunofluorescence labeling showed Kv1.5 labeling clustered in atrial muscle but punctate in the SA node. In atrial muscle, Kv1.5 labeling was closely associated with labeling of Cx43 (gap junction protein) and DPI/II (desmosomal protein), whereas in SA node Kv1.5 labeling was closely associated with labeling of DPI/II but not labeling of Cx43 (absent in the SA node) or Cx45 (another gap junction protein present in the SA node). Electron microscopy and immunogold labeling showed that the Kv1.5 labeling in atrial muscle is preferentially associated with desmosomes rather than gap junctions.

Mechanism of antidiuresis caused by bendroflumethiazide in conscious rats with diabetes insipidus.

1. The mechanism underlying the antidiuretic effect of thiazide diuretics in diabetes insipidus (DI) is unknown. This study addressed two specific questions: is the reduction in urine flow rate (V) related to a decrease in the delivery of fluid from the pars recta of the proximal tubules ('distal delivery'), and are there any changes in the expression and/or intracellular distribution of vasopressin stimulated water channels (AQP2) in the collecting ducts, during chronic thiazide-induced antidiuresis? 2. Nine Brattleboro rats with vasopressin-deficient DI were treated for 5 days with bendroflumethiazide (BFTZ), 9 mg kg(-1) day(-1) orally, and 9 Brattleboro rats were left untreated. BFTZ-treated DI rats showed a fall in V from approximately 200 to approximately 75 ml day(-1) and an increase in urine osmolality from approximately 130 to approximately 400 mosmol kg(-1). 3. BFTZ-induced antidiuresis was associated with a persistent loss of sodium, but not of potassium. After 5 days of treatment, clearance studies in conscious rats showed a tendency towards decreases in effective renal plasma flow (-7%), GFR (-12%) and lithium clearance (C(Li); used as marker for distal delivery) (-25%), compared with untreated controls, but none of these changes were statistically significant. There was no apparent relationship between C(Li) and V in BFTZ-treated or untreated DI rats. 4. BFTZ treatment did not change the expression of AQP2 in homogenates of cortex, outer or inner medulla from DI rats, or from normal Long Evans rats. Light and electron microscopic immunocytochemistry revealed no changes in intracellular distribution of AQP2 in principal cells from inner medullary collecting ducts of BFTZ-treated DI rats. 5. We concluded, (i) that although the antidiuretic effect of BFTZ in rats with DI is associated with a net loss of Na, the decrease in V shows no association with changes in distal delivery, as estimated by C(Li). (ii) Antidiuretic treatment with BFTZ does not alter the expression of subcellular distribution of AQP2 water channels in the collecting ducts. The mechanism underlying the chronic antidiuresis caused by thiazide diuretics in DI remains elusive.

Human articular chondrocytes, synoviocytes and synovial microvessels express aquaporin water channels; upregulation of AQP1 in rheumatoid arthritis.

Recent studies have shown that aquaporin water channels are expressed in human Meckel's cartilage. The aim of the present investigation was to determine if human articular chondrocytes and synoviocytes express aquaporin 1 (AQP1) water channels and to establish if there are any alterations in AQP1 expression in osteoarticular disorders such as osteoarthritis (OA) and rheumatoid arthritis (RA). Immunohistochemistry was employed semi-quantitatively to compare the expression of AQP1 in human chondrocytes derived from normal, OA and RA joints. PCR, cloning and sequencing confirmed the presence of AQP1 transcripts in chondrocytes. Normal human tissue microarrays including samples of kidney, choroid plexus and pancreas were used as positive controls for AQP1 expression. In most tissues AQP1 was expressed along endothelial barriers. In the kidney AQP1 was present in the glomerular capillary endothelium, proximal tubule and descending thin limbs. AQP1 was also localized to pancreatic ducts and acini and the apical membrane domain of the choroid plexus. Immunohistochemistry showed that AQP1 is expressed in synovial micro-vessels, synoviocytes and predominantly in chondrocytes located in the deep zone of articular cartilage. Image analysis of normal, OA and RA cartilage suggested that AQP1 may be upregulated in RA. This is the first report of AQP1 mRNA and protein expression in articular chondrocytes and synoviocytes. These findings suggest a potential role for AQP1 and possibly other members of the AQP gene family in the movement of extracellular matrix and metabolic water across the membranes of chondrocytes and synoviocytes for the purposes of chondrocyte volume regulation and synovial homeostasis.

Pathophysiology of aquaporin-2 in water balance disorders.

The recent identification of aquaporin water channel proteins has provided detailed information about the molecular basis for transepithelial water transport. At least five aquaporins have been identified in the kidney; they have provided detailed molecular insight into the fundamental physiology of water balance. This article focuses primarily on the physiology and pathophysiologic significance of the vasopressin-regulated water channel aquaporin-2 (AQP2) in a number of conditions where body water balance is disturbed. AQP2 is regulated by vasopressin by both short- and long-term mechanisms. Acutely, vasopressin induces exocytic insertion of AQP2 into the apical plasma membrane to increase collecting duct water reabsorption. Moreover, long-term regulation of body water balance is achieved by changes in total collecting duct levels of AQP2. Recent studies have documented that both vasopressin and vasopressin-independent regulation play important roles in this. In conditions with acquired nephrogenic diabetes insipidus (eg, lithium treatment, hypokalemia, postobstructive polyuria), AQP2 expression and targeting have been found to be markedly reduced, providing an explanation for the polyuria and the inability to concentrate urine associated with these conditions. Conversely, in conditions with water retention (eg, heart failure, pregnancy), it has been shown that AQP2 levels and plasma membrane targeting are increased. Continued analysis of aquaporins is providing detailed molecular insight into the physiology and pathophysiology of water balance disorders.

Aquaporins: roles in renal function and peritoneal dialysis.

Aquaporin (AQP) water channels are important in the function of the kidney. Constitutively expressed AQP1 in the proximal tubule and descending limb is important in normal fluid absorption and in the counter-current multiplication system. The vasopressin-regulated shuttling of AQP2 is essential in antidiuresis and the regulation of water balance. Genetic damage to AQPs, or pathological changes in expression or function, impair renal water handling. The most striking examples of this involve disruption of AQP2 function, which can result in profound nephrogenic diabetes insipidus. Aquaporin 1 is present in capillaries and venules and appears to be important in peritoneal dialysis, where it appears to represent the "ultrasmall pores" of the three-pore model. Decreased expression or function of AQP1 may be responsible for some cases of ultrafiltration failure, but further evidence will be required to establish whether this is the case.

Expression and nephron segment-specific distribution of major renal aquaporins (AQP1-4) in Equus caballus, the domestic horse.

Aquaporins (AQPs) play fundamental roles in water and osmolyte homeostasis by facilitating water and small solute movement across plasma membranes of epithelial, endothelial, and other tissues. AQP proteins are abundantly expressed in the mammalian kidney, where they have been shown to play essential roles in fluid balance and urine concentration. Thus far, the majority of studies on renal AQPs have been carried out in laboratory rodents and sheep; no data have been published on the expression of AQPs in kidneys of equines or other large mammals. The aim of this comparative study was to determine the expression and nephron segment localization of AQP1-4 in Equus caballus by immunoblotting and immunohistochemistry with custom-designed rabbit polyclonal antisera. AQP1 was found in apical and basolateral membranes of the proximal convoluted tubules and thin descending limbs of the loop of Henle. AQP2 expression was specifically detected in apical membranes of cortical, medullary, and papillary collecting ducts. AQP3 was expressed in basolateral membranes of cortical, medullary, and papillary collecting ducts. Immunohistochemistry also confirmed AQP4 expression in basolateral membranes of cells lining the distal convoluted and connecting tubules. Western blots revealed high expression of AQP1-4 in the equine kidney. These observations confirm that AQPs are expressed in the equine kidney and are found in similar nephron locations to mouse, rat, and human kidney. Equine renal AQP proteins are likely to be involved in acute and chronic regulation of body fluid composition and may be implicated in water balance disorders brought about by colic and endotoxemia.

Regulation of membrane permeability by vasopressin; activation of the water permeability pathway in toad urinary bladder by N-ethyl-maleimide.

1. Vasopressin induces a rapid increase in water permeability and stimulates net sodium transport in responsive epithelia through the mediation of cAMP. 2. In amphibian urinary bladder, the increase in water permeability is dependent on an intact cytoskeleton and is associated with the exocytotic insertion of tubular vesicles containing particle aggregates (the putative water channels) into the apical membrane of the granular epithelial cells. 3. In the toad bladder, mucosal addition of NEM, 0.1 mM, elicits a slow and irreversible increase in transepithelial water flow, whilst decreasing net sodium transport. 4. The hydrosmotic response to mucosal NEM is inhibited by cellular acidification, by pretreatment with cytoskeleton-disruptive drugs, and by agents that increase cytosolic calcium. 5. Mucosal NEM potentiates the hydrosmotic response to a submaximal, but not a maximal, dose of vasopressin. 6. Mucosal NEM, like vasopressin, induces both vesicle fusion and the appearance of particle aggregates at the granular cell apical surface. 7. NEM, unlike vasopressin, does not increase cellular cAMP content. 8. Mucosal NEM appears to increase transcellular water flow by activating cellular processes normally triggered by vasopressin, at a step beyond cAMP.

Expression of the AQP-1 water channel in normal human tissues: a semiquantitative study using tissue microarray technology.

Aquaporin water channels are a family of membrane proteins that facilitate water movement across biological membranes. Aquaporin-1 (AQP-1) has been found to be important in osmotic water movement across cell membranes of epithelial and endothelial barriers. However, the distribution of AQP-1 in many normal human tissues is still unknown. The aim of this study was to use immunohistochemistry and semiquantitative histomorphometric analysis to determine the tissue distribution and relative expression of AQP-1 in normal human tissues using tissue microarray (TMA) technology. The normal human TMAs employed in this study included cardiovascular, respiratory, gastrointestinal, hepatic and pancreatobiliary, oral, salivary, nasal, mammary, fetal, endocrine, genital tract, central and peripheral nervous systems, urinary tract, skin, cartilage, and other soft connective tissues. Immunohistochemistry and semiquantitative histomorphometric analysis confirmed the presence of AQP-1 in endothelial barriers of almost all tissues and in many epithelial barriers. AQP-1 was highly expressed in the renal cortex, choroid plexus, and pancreatic ducts. AQP-1 expression levels were surprisingly high in the anus, gallbladder, and liver; moderate expression was also detected in the hippocampus and ependymal cells of the central nervous system. This is the first report of AQP-1 protein distribution in normal human TMAs. These findings confirm the presence of AQP-1 in human endothelia and selected water-transporting epithelia and several new locations, including mammary epithelium, articular chondrocytes, synoviocytes, and synovial microvessels where AQP-1 may be involved in milk production, chondrocyte volume regulation, synovial fluid secretion, and homeostasis, respectively.

Activation of the vasopressin-sensitive water permeability pathway in the toad bladder by N-ethyl maleimide.

Vasopressin stimulates transepithelial water flow in the toad urinary bladder. We report here that N-ethyl maleimide (NEM) (0.1 mM) produces a similar increase in osmotic water flow when applied to the mucosal surface of the tissue. NEM-induced water flow is sensitive to inhibitors of hormone-induced water flow, including serosal acidification, or exposure to quinidine or cytoskeleton-disruptive drugs. NEM-induced water flow is additive with that induced by a submaximal, but not a maximal, dose of vasopressin. The response to mucosal NEM is not reversed on removal of the reagent, but established NEM-induced water flow can be inhibited by serosal acidification or quinidine. Like vasopressin, mucosal NEM induces the appearance of fusion profiles and intramembranous particle aggregates (putative water channels) in the apical plasma membrane of the granular cells, and the incidence of particle aggregates correlates with water flow. NEM does not cause an increase in intracellular cAMP. Our data suggest that NEM stimulates transepithelial water flow by irreversibly activating cellular mechanisms normally triggered by vasopressin, hence causing the insertion of water channels.

Effect of a dynein inhibitor on vasopressin action in toad urinary bladder.

1. The effect of the dynein inhibitor erythro-9-[3-(2-hydroxynonyl)] adenine (EHNA) on the osmotic water flow response to vasopressin or exogenous cAMP has been investigated in isolated toad urinary bladders. 2. Pretreatment with serosal EHNA had no effect on basal water flow, but inhibited the development and maintenance of the hydrosmotic response to vasopressin (20 mU ml-1) or 8-(4-parachlorophenylthio)-adenosine 3',5'-cyclic monophosphate (8 CPT-cAMP; 0.1 mM). 3. The inhibitory effect of EHNA on vasopressin-induced water flow was dose dependent. Inhibition occurred in the dose range in which EHNA inhibits the ATPase and motor activities of dynein in vitro. 4. EHNA also inhibited the maintenance of the high rate of water flow established by prior exposure to vasopressin. 5. The inhibitory effect of EHNA on the onset phase of the vasopressin response was attenuated after exposure of the tissue to the microtubule-disruptive drug nocodazole but was fully additive with that of cytochalasin B. 6. EHNA inhibited basal and vasopressin-stimulated transepithelial sodium transport. 7. The findings support the view that EHNA inhibits hormone-induced water flow through an action on a cytoplasmic dynein. The results are consistent with the hypothesis that dynein is involved in the microtubule-based delivery of water channel-containing vesicles to the apical membrane of the granular epithelial cells during both the onset and maintenance of the water permeability response to vasopressin.

Immunohistochemical localization of aquaporin 10 in the apical membranes of the human ileum: a potential pathway for luminal water and small solute absorption.

A new member of the aquaporin family (AQP10) has recently been identified in the human small intestine by molecular cloning and in situ hybridization. Ribonuclease protection assay and northern blotting have demonstrated that AQP10 is expressed in the human duodenum and jejunum. However, the subcellular distribution of the AQP10 protein and its plasma membrane polarization have not yet been established. The objective of this study was to determine the distribution of the AQP10 protein in the human ileum by immunohistochemistry and western blotting using a polyclonal antibody raised against a unique 17-amino acid peptide derived from the human AQP10 sequence. The distribution of the AQP1 and AQP3 proteins was also studied by immunohistochemical staining using affinity-purified polyclonal antibodies. Results revealed that the AQP10 protein is preferentially targeted to the apical membrane domain of absorptive intestinal epithelial cells, whereas AQP3 is located in the basolateral membrane of the cells and AQP1 expression is restricted to the mucosal microvascular endothelia. The presence of AQP10 in the apical membrane of intestinal villi suggests that this protein may represent an entry pathway for water and small solutes from the lumen across to the mucosal side.

Long-term regulation of aquaporins in the kidney.

The discovery of the aquaporin family of water channels has greatly improved our understanding of how water crosses epithelial cells, particularly in the kidney. The study of the mechanisms involved in the regulation of collecting duct water permeability, in particular, has advanced very rapidly since the identification and characterization of aquaporin-2 (AQP2) in 1993. One of the more surprising findings has been the dramatic long-term changes that are seen in the abundance of this protein, as well as the recognition that these changes represent a way of modulating the acute antidiuretic effects of vasopressin. Furthermore, such changes seem to be of etiological and pathological significance in a number of clinical disorders of water balance. This review focuses on the various conditions in which AQP2 expression is altered (either increased or decreased) and on what this can tell us about the signals and mechanisms controlling these changes. Ultimately, this may be of great value in the clinical management of water balance disorders. Evidence is also now beginning to emerge that there are similar changes in the expression of other renal aquaporins, which had previously been thought to provide an essentially constitutive water permeability pathway, suggesting that they too should be considered as regulatory factors in the control of body water balance.

Bilateral ureteral obstruction downregulates expression of vasopressin-sensitive AQP-2 water channel in rat kidney.

Polyuria after release of bilateral ureteral obstruction (BUO) is frequently seen in patients with urological disorders. In this study, we examined the effect of BUO and release of BUO on the expression of the vasopressin-regulated water channel aquaporin-2 (AQP-2) in rat kidney. Ureters were obstructed for 24 h in all experiments, and BUO was either not released or released for 24 or 48 h or 7 days. Each group of experimental rats were matched with sham-operated controls. One kidney was used for membrane fractionation and immunoblotting, whereas the contralateral was fixed for immunocytochemistry. Immunoblotting demonstrated a significant reduction in AQP-2 expression in inner medullar during 24 h of BUO to 26 +/- 8% (P < 0.001). Release of BUO was associated with immediate onset of a predominant osmotic-dependent polyuria. Forty-eight hours after release of BUO, the reduction in AQP-2 expression persisted (19 +/- 8%, P < 0.001), concurrent with a marked nonosmotic postobstructive polyuria, as determined by a significant reduction in free-water clearance (-50 +/- 7 vs. -85 +/- 10 microliters.min-1.kg-1, P < 0.05). Immunofluorescence and immunoelectron microscopy confirmed the reduced levels of AQP-2 in collecting duct principal cells. Seven days after release, the renal excretion of water and electrolytes had almost normalized. However, the downregulation of AQP-2 was not partly reversed (49 +/- 14%, P < 0.001), and, consistent with this, the urinary concentrating capacity was significantly reduced 7 days after release to a 18-h period of thirst. This strongly suggests that the persistent downregulation of AQP-2 is the cause of the slow recovery in concentration capacity. In conclusion, BUO and release of BUO were associated with a marked reduction in expression of AQP-2, coincident with the development and maintenance of postobstructive polyuria. Thus reduced AQP-2 levels may represent an important factor in the slow recovery from postobstructive diuresis.