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Biomedical subjects

M A Knepper

Publications and source records attributed to M A Knepper.

At least 55 records · Page 3Linked to original sources
PubMed1999

Developmental expression of aquaporin 1 in the rat renal vasculature.

Aquaporin 1 (AQP-1) is a water channel protein that is constitutively expressed in renal proximal tubule and descending thin limb cells as well as in endothelial cells of the descending vasa recta. Studies in the developing rat kidney have demonstrated that AQP-1 is expressed in renal tubules before birth. However, nothing is known about the expression of AQP-1 in the renal vasculature during kidney development. The purpose of this study was to establish the distribution of AQP-1 in the renal vasculature of the developing rat kidney and follow the differentiation of the vascular system during kidney development. Kidneys from 16-, 17-, 18-, and 20-day-old fetuses and 1-, 4-, 7-, 14-, 21-, and 28-day-old pups were preserved and processed for immunohistochemical studies using a preembedding immunoperoxidase procedure. AQP-1 immunoreactivity was detected using affinity-purified rabbit polyclonal antibodies to AQP-1. AQP-1 was expressed throughout the arterial portion of the renal vasculature of the fetal and neonatal kidney from gestational age 17 days to 1 wk after birth. AQP-1 immunoreactivity gradually disappeared from the renal vasculature between 1 and 2 wk of age and remained only in the descending vasa recta. In contrast, AQP-1 immunoreactivity was not observed in lymphatic vessels until 3 wk of age and persisted in the adult kidney. AQP-1 was also expressed in a population of interstitial cells in the terminal part of the renal papilla at 3 wk of age as well as in the adult kidney. The transient expression of AQP-1 in the arterial portion of the renal vasculature in the developing rat kidney suggests that AQP-1 is important for fluid equilibrium and/or drainage in the developing kidney or, alternatively, plays a role in the regulation of growth and/or branching of the vascular tree during kidney development.

Aging↗
PubMed1999

Vasopressin regulates apical targeting of aquaporin-2 but not of UT1 urea transporter in renal collecting duct.

In the renal inner medullary collecting duct (IMCD), vasopressin regulates two key transporters, namely aquaporin-2 (AQP2) and the vasopressin-regulated urea transporter (VRUT). Both are present in intracellular vesicles as well as the apical plasma membrane. Short-term regulation of AQP2 has been demonstrated to occur by vasopressin-induced trafficking of AQP2-containing vesicles to the apical plasma membrane. Here, we have carried out studies to determine whether short-term regulation of VRUT occurs by a similar process. Cell surface labeling with NHS-LC-biotin in rat IMCD suspensions revealed that vasopressin causes a dose-dependent increase in the amount of AQP2 labeled at the cell surface, whereas VRUT labeled at the cell surface did not increase in response to vasopressin. Immunoperoxidase labeling of inner medullary thin sections from Brattleboro rats treated with 1-desamino-8-D-arginine vasopressin (DDAVP) for 20 min revealed dramatic translocation of AQP2 to the apical region of the cell, with no change in the cellular distribution of VRUT. In addition, differential centrifugation of inner medullary homogenates from Brattleboro rats treated with DDAVP for 60 min revealed a marked depletion of AQP2 from the low-density membrane fraction (enriched in intracellular vesicles) but did not alter the quantity of VRUT in this fraction. Finally, AQP2-containing vesicles immunoisolated from a low-density membrane fraction from renal inner medulla did not contain immunoreactive VRUT. Thus vasopressin-mediated regulation of AQP2, but not of VRUT, depends on regulated vesicular trafficking to the plasma membrane.

Animals↗
PubMed1999

Cyclooxygenase inhibitors increase Na-K-2Cl cotransporter abundance in thick ascending limb of Henle's loop.

Cyclooxygenase inhibitors, such as indomethacin and diclofenac, have well-described effects to enhance renal water reabsorption and urinary concentrating ability. Concentrating ability is regulated in part at the level of the thick ascending limb of Henle's loop, where active NaCl absorption drives the countercurrent multiplication mechanism. We used semiquantitative immunoblotting to test the effects of indomethacin and diclofenac, given over a 48-h period, on the expression levels of the ion transporters responsible for active NaCl transport in the thick ascending limb. Both agents strongly increased the expression level of the apical Na-K-2Cl cotransporter in both outer medulla and cortex. Neither agent significantly altered outer medullary expression levels of other thick ascending limb proteins, namely, the type 3 Na/H exchanger (NHE-3), Tamm-Horsfall protein, or alpha1- or beta1-subunits of the Na-K-ATPase. Administration of the EP3-selective PGE(2) analog, misoprostol, to indomethacin-treated rats reversed the stimulatory effect of indomethacin on Na-K-2Cl cotransporter expression. We conclude that cyclooxygenase inhibitors enhance urinary concentrating ability in part through effects to increase Na-K-2Cl cotransporter expression in the thick ascending limb of Henle's loop. This action is most likely due to elimination of an EP3-receptor-mediated tonic inhibitory effect of PGE(2) on cAMP production.

Animals↗
PubMed1999

Decreased renal Na-K-2Cl cotransporter abundance in mice with heterozygous disruption of the G(s)alpha gene.

Transport processes along the nephron are regulated in part by hormone stimulation of adenylyl cyclases mediated by the heterotrimeric G protein G(s). To assess the role of this pathway in the regulation of Na-K-2Cl cotransporter abundance in the renal thick ascending limb (TAL), we studied mice with heterozygous disruption of the Gnas gene, which codes for the alpha-subunit of G(s). Outer medullary G(s)alpha protein abundance (as assessed by semiquantitative immunoblotting) and glucagon-stimulated cAMP production were significantly reduced in the heterozygous G(s)alpha knockout mice (GSKO) relative to their wild-type (WT) littermates. Furthermore, Na-K-2Cl cotransporter protein abundance in the outer medulla was significantly reduced (band density, 48% of WT). In addition, GSKO mice had a significantly reduced (72% of WT) urinary osmolality in response to a single injection of 1-deamino-[8-D-arginine]vasopressin (DDAVP), a vasopressin analog. In contrast, outer medullary protein expression of the type 3 Na/H exchanger (NHE-3) or Tamm-Horsfall protein did not differ between the GSKO mice and their WT littermates. However, abundance of type VI adenylyl cyclase was markedly decreased in the outer medullas of GSKO mice, suggesting a novel feed-forward regulatory mechanism. We conclude that expression of the Na-K-2Cl cotransporter of the TAL is dependent on G(s)alpha-mediated hormone stimulation, most likely due to long-term changes in cellular cAMP levels.

Adenylyl Cyclases↗
PubMed1999

Altered expression of Na transporters NHE-3, NaPi-II, Na-K-ATPase, BSC-1, and TSC in CRF rat kidneys.

In chronic renal failure (CRF), reduction in renal mass leads to an increase in the filtration rates of the remaining nephrons and increased excretion of sodium per nephron. To address the mechanisms involved in the increased sodium excretion, we determined the total kidney levels and the densities per nephron of the major renal NaCl transporters in rats with experimental CRF. Two weeks after 5/6 nephrectomy (reducing the total number of nephrons to approximately 24 +/- 8%), the rats were azotemic and displayed increased Na excretion. Semiquantitative immunoblotting revealed significant reduction in the total kidney levels of the proximal tubule Na transporters NHE-3 (48% of control), NaPi-II (13%), and Na-K-ATPase (30%). However, the densities per nephron of NHE-3, NaPi-II, and Na-K-ATPase were not significantly altered in remnant kidneys, despite the extensive hypertrophy of remaining nephrons. Immunocytochemistry confirmed the reduction in NHE-3 and Na-K-ATPase labeling densities in the proximal tubule. In contrast, there was no significant reduction in the total kidney levels of the thick ascending limb and distal convoluted tubule NaCl transporters BSC-1 and TSC, respectively. This corresponded to a 3.6 and 2.5-fold increase in densities per nephron, respectively (confirmed by immunocytochemistry). In conclusion, in this rat CRF model: 1) increased fractional sodium excretion is associated with altered expression of proximal tubule Na transporter expression (NHE-3, NaPi-II, and Na-K-ATPase), consistent with glomerulotubular imbalance in the face of increased single-nephron glomerular filtration rate; and 2) compensatory increases in BSC-1 and TSC expression per nephron occur in distal segments.

Animals↗
PubMed1999

Reduced abundance of aquaporins in rats with bilateral ischemia-induced acute renal failure: prevention by alpha-MSH.

We examined the effect of temporary renal ischemia (30 min or 60 min) and reperfusion (1 day or 5 days) on the expression of renal aquaporins (AQPs) and urinary concentration in rats with bilateral ischemia-induced acute renal failure (ARF). Next, we tested whether reducing ischemia/reperfusion (I/R) injury by treatment with alpha-melanocyte stimulating hormone (alpha-MSH) affects the expression of AQPs and urine output. Rats with ARF showed significant renal insufficiency, and urinary concentration was markedly impaired. In rats with mild ischemic injury (30 min), urine output increased significantly to a maximum at 48 h, and then nearly normalized within 5 days. Consistent with this, semiquantitative immunoblotting revealed that kidney AQP1 and AQP2 abundance was significantly decreased after 24 h to 30 +/- 5% and 40 +/- 11% (n = 8) of controls (n = 9), respectively (P < 0.05). Five days after ischemia, AQP2 abundance was not significantly decreased and urine output was normalized. In contrast, severe ischemic injury (60 min) resulted in a marked reduction in urine output at 24 h, despite a significant decrease in urine osmolality and solute-free water reabsorption, T(c)H(2)O. AQP1 and AQP2 abundance was markedly decreased to 51 +/- 5% and 31 +/- 9% (n = 10) of controls (n = 8) at 24 h (P < 0.05). After 5 days, the rats developed gradually severe polyuria and had very low AQP2 and AQP1 levels [11 +/- 4% and 6 +/- 2% (n = 5) of controls (n = 8), respectively; P < 0.05]. A similar reduction was observed for AQP3. The reduction in AQP expression in the proximal tubule and inner medullary collecting duct was confirmed by immunocytochemistry. Next, we found that intravenous alpha-MSH treatment of rats with ARF significantly reduced the ischemia-induced downregulation of renal AQPs and reduced the polyuria. In conclusion, the I/R injury is associated with markedly reduced expression of the collecting duct and proximal tubule AQPs, in association with an impairment of urinary concentration. Moreover, alpha-MSH treatment significantly prevented the reduction in expression of AQPs and renal functional defects. Thus decreased AQP expression is likely to contribute to the impairment in urinary concentration in the postischemic period.

Animals↗
PubMed1999

Reduced water permeability and altered ultrastructure in thin descending limb of Henle in aquaporin-1 null mice.

It has been controversial whether high water permeability in the thin descending limb of Henle (TDLH) is required for formation of a concentrated urine by the kidney. Freeze-fracture electron microscopy (FFEM) of rat TDLH has shown an exceptionally high density of intramembrane particles (IMPs), which were proposed to consist of tetramers of aquaporin-1 (AQP1) water channels. In this study, transepithelial osmotic water permeability (Pf) was measured in isolated perfused segments (0.5-1 mm) of TDLH in wild-type (+/+), AQP1 heterozygous (+/-), and AQP1 null (-/-) mice. Pf was measured at 37 degrees C using a 100 mM bath-to-lumen osmotic gradient of raffinose, and fluorescein isothiocyanate (FITC)-dextran as the luminal volume marker. Pf was (in cm/s): 0.26 +/- 0.02 ([+/+]; SE, n = 9 tubules), 0.21 +/- 0.01 ([+/-]; n = 12), and 0.031 +/- 0.007 ([-/-]; n = 6) (P < 0.02, [+/+] vs. [+/-]; P < 0.0001, [+/+] vs. [-/-]). FFEM of kidney medulla showed remarkably fewer IMPs in TDLH from (-/-) vs. (+/+) and (+/-) mice. IMP densities were (in microm-2, SD, 5-12 micrographs): 5,880 +/- 238 (+/+); 5,780 +/- 450 (+/-); and 877 +/- 420 (-/-). IMP size distribution analysis revealed mean IMP diameters of 8.4 nm ([+/+] and [+/-]) and 5.2 nm ([-/-]). These results demonstrate that AQP1 is the principal water channel in TDLH and support the view that osmotic equilibration along TDLH by water transport plays a key role in the renal countercurrent concentrating mechanism. The similar Pf and AQP1 expression in TDLH of (+/+) and (+/-) mice was an unexpected finding that probably accounts for the unimpaired urinary concentrating ability in (+/-) mice.

Animals↗
PubMed1999

Aldosterone-mediated regulation of ENaC alpha, beta, and gamma subunit proteins in rat kidney.

Aldosterone stimulates sodium transport in the renal collecting duct by activating the epithelial sodium channel (ENaC). To investigate the basis of this effect, we have developed a novel set of rabbit polyclonal antibodies to the 3 subunits of ENaC and have determined the abundance and distribution of ENaC subunits in the principal cells of the rat renal collecting duct. Elevated circulating aldosterone (due to either dietary NaCl restriction or aldosterone infusion) markedly increased the abundance of alphaENaC protein without increasing the abundance of the beta and gamma subunits. Thus, alphaENaC is selectively induced by aldosterone. In addition, immunofluorescence immunolocalization showed a striking redistribution in ENaC labeling to the apical region of the collecting duct principal cells. Finally, aldosterone induced a shift in molecular weight of gammaENaC from 85 kDa to 70 kDa, consistent with physiological proteolytic clipping of the extracellular loop as postulated previously. Thus, at the protein level, the response of ENaC to aldosterone stimulation is heterogenous, with both quantitative and qualitative changes that can explain observed increases in ENaC-mediated sodium transport.

Aldosterone↗
PubMed1999

Kidney aquaporin-2 expression during escape from antidiuresis is not related to plasma or tissue osmolality.

Recent results indicate that renal escape from vasopressin-induced antidiuresis is accompanied by a marked downregulation of whole kidney aquaporin-2 (AQP-2) protein and mRNA expression. However, in those studies, the escaped animals were also markedly hypo-osmolar compared to controls as a result of water loading during antidiuresis. The present studies evaluated whether systemic or local osmolality contributes to the downregulation of AQP-2 expression in this model. In the first study, two groups of 1-deamino-[8-D-arginine]-vasopressin (dDAVP)-infused rats were water-loaded; after establishment of escape, one group was then water-restricted for 4 d to reverse the escape, whereas the other group continued daily water loading. Whole kidney AQP-2 protein was measured by Western blotting, and inner medulla AQP-2 mRNA was determined by Northern blotting. Results were compared to dDAVP-infused rats fed solid chow. After 4 d of water restriction, urine volume decreased to the same level as in the rats on solid chow; however, plasma sodium concentrations and plasma osmolality remained low. Despite maintenance of significant hypo-osmolality, rats in which escape was subsequently reversed by water restriction reestablished high dDAVP-stimulated kidney levels of AQP-2 after 4 d of water restriction. In the second study, AQP-2 expression was evaluated in different regions of kidneys from water-loaded rats undergoing escape from antidiuresis. Despite markedly different interstitial osmolalities, significant downregulation of AQP-2 expression compared to dDAVP-infused control rats was seen in the inner medulla, outer medulla, and cortex. Thus, neither systemic nor interstitial osmolality appears to appreciably be correlated with downregulation of kidney AQP-2 expression during escape from antidiuresis. These results therefore suggest that additional vasopressin- and osmolality-independent factors, likely related to the effects of extracellular fluid volume expansion, also regulate kidney AQP-2 expression in rats.

Analysis of Variance↗
PubMed1999

Regulation of thick ascending limb ion transporter abundance in response to altered acid/base intake.

Changes in ammonium excretion with acid/base perturbations are dependent on changes in medullary ammonium accumulation mediated by active NH4+ absorption by the medullary thick ascending limb. To investigate whether alterations in the abundance of medullary thick ascending limb ion transporters, namely the apical Na+/K+(NH4+)/2Cl- -cotransporter (BSC-1), the apical Na+/H+ -exchanger (NHE3), and the Na+/K+ -ATPase alpha1-subunit, may be responsible in part for altered medullary ammonium accumulation, semiquantitative immunoblotting studies were performed using homogenates from the inner stripe of the rat renal outer medulla. After 7 d of NH4Cl (7.2 mmol/220 g body wt per d) loading (associated with increased medullary ammonium accumulation), neither BSC-1 nor Na+/K+ -ATPase protein expression was altered, but NHE3 protein abundance was significantly increased. On the other hand, both BSC-1 and Na+/K+ -ATPase protein abundance was increased significantly in rats fed NaHCO3 (7.2 mmol/220 g body wt per d) for 7 d. Rats fed a high-NaCl diet (7.7 mEq Na+/220 g body wt per d) for 5 d also showed marked increases in both BSC-1 and Na+/K+ -ATPase expression. The expression level of NHE3 protein did not change with either NaHCO3 or high NaCl intake. None of these three transporters showed a significant difference in abundance between the groups fed equimolar (7.2 mmol/220 g body wt per d for 7 d) NaHCO3 or NaCl. It is concluded that outer medullary BSC-1 and Na+/K+ -ATPase alpha1-subunit protein abundance is increased by chronic Na+ loading but not by acid/base perturbations and that outer medullary NHE3 protein abundance is increased by chronic NH4Cl loading.

Acid-Base Equilibrium↗
PubMed1999

Decreased abundance of collecting duct aquaporins in post-ischemic renal failure in rats.

Increased urine flow is often a feature of mild to moderate acute renal failure. This study examines the possible role of dysregulation of collecting duct aquaporins as a factor in this increase. In rats, the left renal pedicle was clamped for 45 min followed by contralateral nephrectomy. Control rats were identical except that the renal pedicle was not clamped. Rats were sacrificed and the kidneys were homogenized at various time points after release of the clamp for semiquantitative immunoblotting of collecting duct aquaporins, as well as the thick ascending limb Na-K-2Cl cotransporter and the proximal tubule water channel, aquaporin-1. Urinary flow rate was significantly increased 18 h after the ischemic insult and remained increased through 72 h. Whole kidney aquaporin-2 protein abundance was 45% of controls at 18 h, 55% of controls at 36 h, and returned to normal 72 h after ischemia. Whole kidney aquaporin-3 protein abundance was 37% of controls at 18 h, 13% of controls at 36 h, and 45% of controls at 72 h. The decline in aquaporin-2 and -3 was confirmed by immunocytochemistry. Abundance of the thick ascending limb Na-K-2Cl cotransporter protein was not significantly decreased. Aquaporin-1 protein abundance was not significantly decreased at 18 h after the ischemic insult, but was significantly reduced after 36 h. Thus, the post-ischemic state is associated with decreased levels of the collecting duct aquaporins, coinciding with an increase in water excretion. It is concluded that decreased aquaporin protein abundance in collecting duct cells is a contributing factor in the increased urine flow seen in moderate post-ischernic acute renal failure.

Acute Kidney Injury↗
PubMed1999

Renal expression of aquaporins in liver cirrhosis induced by chronic common bile duct ligation in rats.

Semiquantitative immunoblotting was used to investigate the expression levels of the four major renal aquaporins, the Na-K-2Cl cotransporter of the thick ascending limb, the type 3 Na-H exchanger, and the Na-K-ATPase in kidneys from rats with cirrhosis secondary to common bile duct ligation (CBDL). These rats had significant water retention and hyponatremia. In contrast to models of cirrhosis induced by carbon tetrachloride, aquaporin-2 expression in CBDL-induced cirrhosis was decreased. Thus, these results show that in the setting of extracellular fluid volume expansion, excessive water retention with hyponatremia can occur in the absence of increases in aquaporin-2 abundance. In addition, the expression levels of the two basolateral collecting duct aquaporins (aquaporin-3 and -4) were decreased in CBDL rats relative to sham-operated control rats. Similarly, the Na-K-2Cl cotransporter of the thick ascending limb and the type 3 Na-H exchanger showed decreases in expression. In contrast, the expression levels of aquaporin-1 and the all subunit of the Na-K-ATPase were not decreased. Thus, dysregulation of multiple water channels and ion transporters may play a role in water balance abnormalities associated with CBDL-induced cirrhosis in rats.

Animals↗
PubMed1998

The thiazide-sensitive Na-Cl cotransporter is an aldosterone-induced protein.

Although the collecting duct is regarded as the primary site at which mineralocorticoids regulate renal sodium transport in the kidney, recent evidence points to the distal convoluted tubule as a possible site of mineralocorticoid action. To investigate whether mineralocorticoids regulate the expression of the thiazide-sensitive Na-Cl cotransporter (TSC), the chief apical sodium entry pathway of distal convoluted tubule cells, we prepared an affinity-purified, peptide-directed antibody to TSC. On immunoblots, the antibody recognized a prominent 165-kDa band in membrane fractions from the renal cortex but not from the renal medulla. Immunofluorescence immunocytochemistry showed TSC labeling only in distal convoluted tubule cells. Semiquantitative immunoblotting studies demonstrated a large increase in TSC expression in the renal cortex of rats on a low-NaCl diet (207 +/- 21% of control diet). Immunofluorescence localization in tissue sections confirmed the strong increase in TSC expression. Treatment of rats for 10 days with a continuous subcutaneous infusion of aldosterone also increased TSC expression (380 +/- 58% of controls). Furthermore, 7-day treatment of rats with an orally administered mineralocorticoid, fludrocortisone, increased TSC expression (656 +/- 114% of controls). We conclude that the distal convoluted tubule is an important site of action of the mineralocorticoid aldosterone, which strongly up-regulates the expression of TSC.

Aldosterone↗
PubMed1998

Defective proximal tubular fluid reabsorption in transgenic aquaporin-1 null mice.

To investigate the role of aquaporin-1 (AQP1) water channels in proximal tubule function, in vitro proximal tubule microperfusion and in vivo micropuncture measurements were done on AQP1 knockout mice. The knockout mice were generated by targeted gene disruption and found previously to be unable to concentrate their urine in response to water deprivation. Unanesthetized knockout mice consumed 2.8-fold more fluid than wild-type mice and had lower urine osmolality (505 +/- 40 vs. 1081 +/- 68 milliosmolar). Transepithelial osmotic water permeability (Pf) in isolated microperfused S2 segments of proximal tubule from AQP1 knockout [-/-] mice was 0.033 +/- 0.005 cm/s (SE, n = 6 mice, 37 degreesC), much lower than that of 0.15 +/- 0.03 cm/s (n = 8) in tubules from wild-type [+/+] mice (P < 0.01). In the presence of isosmolar luminal perfusate and bath solutions, spontaneous fluid absorption rates (nl/min/mm tubule length) were 0.31 +/- 0.12 (-/-, n = 5) and 0.64 +/- 0.15 (+/+, n = 8). As determined by free-flow micropuncture, the ratios of tubular fluid-to-plasma concentrations of an impermeant marker TF/P in end proximal tubule fluid were 1.36 +/- 0. 05 (-/-, n = 8 mice [53 tubules]) and 1.95 +/- 0.09 (+/+, n = 7 mice [40 tubules]) (P < 0.001), corresponding to 26 +/- 3% [-/-] and 48 +/- 2% [+/+] absorption of the filtered fluid load. In collections of distal tubule fluid, TF/P were 2.8 +/- 0.3 [-/-] and 4.4 +/- 0.5 [+/+], corresponding to 62 +/- 4% [-/-] and 76 +/- 3% [+/+] absorption (P < 0.02). These data indicate that AQP1 deletion in mice results in decreased transepithelial proximal tubule water permeability and defective fluid absorption. Thus, the high water permeability in proximal tubule of wild-type mice is primarily transcellular, mediated by AQP1 water channels, and required for efficient near-isosmolar fluid absorption.

Animals↗
PubMed1998

Studies of renal aquaporin-2 expression during renal escape from vasopressin-induced antidiuresis.

In animal models of the syndrome of inappropriate antidiuresis (SIADH), sustained administration of vasopressin and water results in free-water retention and progressive hyponatremia for several days, which is then followed by escape from the vasopressin-induced antidiuresis. With the onset of vasopressin escape, water excretion increases despite sustained administration of vasopressin, allowing water balance to be re-established and the serum sodium to be stabilized at a steady, albeit decreased, level. Studies from our laboratories have investigated whether this escape phenomenon can be attributed to altered regulation of aquaporin water channels. After four-day pre-treatment with 1-deamino-[8-D-arginine]-vasopressin (dDAVP) by osmotic minipump, rats were divided into control (continued dDAVP) and water-loaded (continued dDAVP plus a daily oral water load) groups. A significant increase in urine volume in the water-loaded rats was observed by the second day of water loading, indicating escape from antidiuresis. The onset of escape coincided temporally with a marked decrease in renal aquaporin-2 protein (measured by semi-quantitative immunoblotting), which began at day 2 and fell to 17% of control levels by day 3. In contrast, there was no decrease in the renal expression of aquaporins 1, 3, or 4. The marked suppression of whole kidney aquaporin-2 protein was accompanied by a concomitant suppression of whole kidney aquaporin-2 mRNA levels. Immunocytochemical localization and differential centrifugation studies demonstrated that trafficking of aquaporin-2 to the plasma membrane remained intact during vasopressin escape. Additional studies have indicated that the observed down-regulation of aquaporin-2 expression also occurs in the renal cortex as well as the inner and outer medullas, and can be reversed simply by water restriction despite maintenance of hyponatremia. Our results therefore suggest that escape from vasopressin-induced antidiuresis is attributable, at least in part, to a vasopressin-independent and osmolality-independent decrease in aquaporin-2 water channel expression in the renal collecting duct. Similar mechanisms likely contribute to the phenomenon of escape from antidiuresis seen clinically in patients with SIADH as well.

Animals↗
PubMed1998

Impaired aquaporin and urea transporter expression in rats with adriamycin-induced nephrotic syndrome.

Nephrotic syndrome is associated with abnormal regulation of renal water excretion. To investigate the role of collecting duct water channels and solute transporters in this process, we have carried out semiquantitative immunoblotting of kidney tissues from rats with adriamycin-induced nephrotic syndrome. These experiments demonstrated that adriamycin-induced nephrotic syndrome is associated with marked decreases in expression of aquaporin-2, aquaporin-3, aquaporin-4, and the vasopressin-regulated urea transporter in renal inner medulla, indicative of a suppression of the capacity for water and urea absorption by the inner medullary collecting duct. In contrast, expression of the alpha(1)-subunit of the Na,K-ATPase in the inner medulla was unaltered. Light and electron microscopy of perfusion-fixed kidneys demonstrated that the collecting ducts are morphologically normal and unobstructed. Inner medullary expression of the descending limb water channel, aquaporin-1, was not significantly altered, pointing to a selective effect on the collecting duct. Aquaporin-2 and aquaporin-3 expression was also markedly diminished in the renal cortex, indicating that the effect is not limited to the inner medullary collecting duct. Differential centrifugation studies and immunocytochemistry in inner medullary thin sections demonstrated increased targeting of aquaporin-2 to the plasma membrane, consistent with the expected short-term action of vasopressin on aquaporin-2 trafficking. The extensive down-regulation of aquaporin and urea transporter expression may represent an appropriate renal response to the extracellular volume expansion observed in nephrotic syndrome, but may occur at the expense of decreased urinary concentrating and diluting capacity.

Adenylyl Cyclases↗
PubMed1998

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.

Animals↗