1 Service de Biologie Cellulaire, Commissariat à l'Énergie Atomique/Saclay, F-91191 Gif-sur-Yvette; 2 Assistance Publique-Hôpitaux de Paris, Hôpital Sainte-Périne, F-75781 Paris cedex 16; 3 Laboratoire de Physiologie de l'Environnement, Université Claude-Bernard, Faculté de Médecine Grange-Blanche, F-69373 Lyon cedex 8, France; and 4 Department of Cell Biology, Institute of Anatomy, University of Aarhus, DK-8000 Aarhus, Denmark
![]() |
ABSTRACT |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
First published August 8, 2001;
10.1152/ajprenal.00139.2001.The mechanisms underlying the prevention
of age-related polyuria by chronic food restriction were investigated
in female WAG/Rij rats. The decreased osmolality of renal papilla
observed in senescent rats was not corrected by food restriction. A
reduced urea content in the inner medulla of senescent rats, fed ad
libitum or food-restricted, was suggested by the marked decrease in
expression of UT-A1 and UT-B1 urea transporters. Aquaporin-2 (AQP2)
downregulation in the inner medulla of senescent rats was partially
prevented by food restriction. Both AQP2 and the phosphorylated form of
AQP2 (p-AQP2), the presence of which was diffuse within the cytoplasm of collecting duct principal cells in normally fed senescent rats, were
preferentially targeted at the apical region of the cells in
food-restricted senescent animals. Plasma vasopressin (AVP) was similar
in 10- and 30-mo-old rats fed ad libitum, but was doubled in
food-restricted 30-mo-old rats. This study indicates that 1)
kidney aging is associated with a marked decrease in AQP2, UT-A1, and
UT-B1 expression in the inner medulla and a reduced papillary
osmolality; and 2) the prevention of age-related polyuria by
chronic food restriction occurs through an improved recruitment of AQP2
and p-AQP2 to the apical membrane in inner medulla principal cells,
permitted by increased plasma AVP concentration.
kidney aging; phosphorylation; urea transporters
![]() |
INTRODUCTION |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
IN MAMMALIAN KIDNEY, MOST of filtered water is reabsorbed through the water channel aquaporin-1 (AQP1) constitutively present in the apical and basolateral membranes of the proximal tubule and thin descending limb (28, 35). Final urine is concentrated along the collecting duct by water subtraction owing to the corticopapillary osmotic gradient and water permeability of principal cells, endowed by AQP2 in the apical membrane and both AQP3 and AQP4 in the basolateral membrane. AQP2 targeting to the apical membrane is tightly regulated by arginine vasopressin (AVP), which induces fusion of subapical vesicles containing AQP2 with plasma membrane (13, 27). This process occurs through AVP binding to its V2 receptor in the basolateral membrane, activation of the cAMP signaling pathway, and phosphorylation of AQP2 at serine-256 by protein kinase A (22).
Polyuria with reduced urine osmolality affects elderly humans and rodents (5, 6, 9, 42). In some strains of rats, this age-related defect has been attributed to a loss of nephrons or an impaired AVP secretion (25, 32, 40, 46, 53, 55). However, it is still observed in senescent female WAG/Rij rats that remain free of glomerulosclerosis and have a normal number of nephrons, renal blood flow, and single renal filtration rates (10-12). Plasma AVP concentration, V2-receptor expression, and intracellular cAMP content in the papilla are also maintained in these old rats (15, 30). By contrast, senescent female WAG/Rij rats exhibited a low papillary osmolality and a weak expression of AQP2 and AQP3, whereas expression of AQP1 and AQP4 was unaltered (30). This suggested a low water permeability of the inner medullary collecting tubule, a hypothesis strengthened by the diffused localization of AQP2 within the cytoplasm of papillary principal cells (30).
A reduced papillary osmolality may result from a lower accumulation of NaCl and/or urea within the medulla. Defective sodium accumulation could not be implicated univocally. As a matter of fact, sodium transport in Henle's loop of the cortical nephron was similar in female WAG/Rij rats of 10 and 30 mo of age (11). On the other hand, a reduced AVP-dependent reabsorption of NaCl was found in the medullary thick ascending limb dissected from aging mice (14). This is consistent with the diminished accumulation of cAMP in the same segment of the nephron after maximal AVP stimulation reported in mice and in female WAG/Rij rats (12, 21). Accumulation of urea in the medulla is dependent on urea transporters (UTs) (2, 16, 23, 37). UT-A1 is responsible for the AVP-stimulated urea transport, allowing the diffusion of urea from the lumen of terminal inner medullary collecting duct to papilla interstitium (38, 41). UT-A3 has been recently cloned (18, 39) and located in the apical region of terminal collecting duct (45). Its function is still unknown, but its large abundance suggests a role in renal urea handling. UT-A2 and UT-B1 are located in descending structures, respectively, the thin descending limb (29) and descending vasa recta (54). Both UT-A2 and UT-B1 are involved in the medullary recycling of urea. Therefore, it is expected that any age-related change in UT-A1, UT-A2, and UT-B1 expression would modify the corticopapillary gradient and urine osmolality.
Chronic food restriction, which is known to delay the aging process, prevents polyuria and urinary concentrating defect affecting aging female WAG/Rij rats (43). The mechanisms underlying such prevention were presently investigated by measuring water balance, papillary osmolality, plasma AVP concentration, and both AQP and UT expression in senescent rats chronically food restricted by 30% from 10 to 30 mo compared with adult and senescent animals fed ad libitum. To further investigate the recruitment of AQP2 to the apical membrane of principal cells in the inner medullary collecting duct, and the role of phosphorylation in this trafficking, indirect immunofluorescence and electron microscopy were performed to localize the phosphorylated form of AQP2 (p-AQP2).
![]() |
MATERIALS AND METHODS |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Animals and food restriction protocol. Female WAG/Rij rats were born and raised in the animal care facility of the Commissariat à l'Énergie Atomique, Saclay, Gif-sur-Yvette, France, as previously described (30, 43). The WAG/Rij strain is an inbred Wistar rat strain that has been well characterized by Burek (7). Mean lifespan for females is close to 30 mo when they are fed ad libitum. Their body weight increases until they are 6-10 mo old and reaches a plateau corresponding to 180-200 g. From 10 to 30 mo, this body weight is unchanged or slightly increased by 20-40 g (7, 9, 43). This prevention of excess weight with age in WAG/Rij rats is, in part, related to their low spontaneous food intake. Measurement of daily food intake in metabolic cages indicates that for a similar body weight, food consumption of WAG/Rij rats is 40% lower than that of the regular Wistar rats (Iffa Credo) (data not shown). In addition, female WAG/Rij rats have an unchanged blood pressure, are free of glomerulosclerosis, and have a constant number of nephrons with age, at least until 30 mo (9). This makes the WAG/Rij rat a suitable experimental model to study the effect of food restriction on kidney aging per se (4, 43).
The animals were maintained on a 12:12-h light-dark cycle, 50% humidity, and a temperature of 21°C. From weaning to 3 mo, the animals were fed a commercial growth diet (DO3, UAR, Villemoisson, France) composed of 9% fish protein, 16% vegetable protein, and a total of 3,500 kcal/kg. At the age of 3 mo, they were switched to a diet containing 2% fish and 15% vegetable protein, 0.71% phosphate, 0.78% calcium, 0.62% potassium, 0.27% sodium, 0.22% magnesium, and a total of 2,900 kcal/kg (DO4, UAR). The ad libitum food intake of animals was close to 10 g/day, regardless of age (43). Chronic food restriction consisted of offering 7 g/day of the same diet to rats, a 30% restriction, from 10 to 30 mo of age, at which time they were killed. During this period of time, the survival curves were similar for food-restricted and ad libitum fed rats; i.e., 36% of the animals had died at 30 mo in both groups (43). A short-term 30% food restriction was performed for 1 wk in 10- and 30-mo-old rats for comparison with the long-term protocol.Metabolic cage data. Daily water and food intake, as well as urinary volume, were measured for a 24-h period after 2 days of acclimatization to metabolic cages. Urine was collected under oil to avoid evaporation, and osmolality was determined with a Roebling Automatik osmometer.
Measurement of plasma AVP, plasma osmolality, and papillary osmolality. Blood and kidneys were collected immediately after decapitation of rats. Plasma AVP concentration was measured by radioimmunoassay with antiserum K9-IV (a gift of Dr. L. C. Keil, National Aeronautics and Space Administration Ames Research Center; Ref. 19) and 125I-labeled iodo-AVP (15, 19). The minimum sensitivity of the assay was 0.25 pg/assay. For each animal, determination of plasma AVP concentration was performed in duplicate on three occasions (with independent standard curves). Plasma osmolality was measured on the same samples with a Fiske one-ten osmometer. The whole white papilla was excised from each kidney, weighed, and homogenized manually with a glass potter after addition of 200 µl of distilled water. The osmolality of the homogenates was measured with a Roebling Automatik osmometer as previously described (30). Papilla osmolality was calculated on the assumption that 80% of the wet weight is water (1).
Antibodies. UT-A1 and UT-A2 were revealed with an affinity-purified rabbit polyclonal antibody directed against the COOH-terminal 19-amino acid sequence common to rat UT-A1 and UT-A2 (Alpha Diagnostic International, San Antonio, TX). UT-B1 was detected with a rabbit polyclonal antibody raised against a synthetic peptide from the NH2-terminal sequence of rat UT-B1 (Neosystem, Strasbourg, France) (17). AQP2 and AQP3 were detected with polyclonal antibodies raised against the COOH-terminal peptide sequence of rat AQP2 and rat AQP3 (3, 33). An affinity-purified antibody against p-AQP2, AN244-pp-AP, was prepared using a peptide corresponding to amino acids 253-262 of rat AQP2 phosphorylated at serine-256, as previously described (8).
Western blot analysis.
Fragments of the inner stripe of the outer medulla and of the inner
medulla were dissected on ice from each kidney and were homogenized
manually with a glass potter in 500 µl of PBS, pH 7.4, containing
103 mol/l phenylmethylsulfonyl fluoride. Protein
concentration of the crude homogenates was determined by the method of
Bradford using a protein assay reagent kit (Bio-Rad, Hercules, CA).
Samples were then diluted to a final concentration of 0.5 mg/ml,
solubilized in Laemmli buffer, and heated at 65°C for 10 min.
Proteins were separated by SDS-PAGE (12% for AQPs and 10% for UTs),
and immunoblotting was performed as previously described
(30). The membranes were blocked for 30 min at room
temperature with PBS supplemented with 5% nonfat dry milk, followed by
incubation with the primary polyclonal antibody for 1 h at room
temperature. The membranes were then washed and incubated with a
peroxidase-conjugated anti-rabbit IgG polyclonal antibody (0.2 µg/ml,
Promega, Madison, WI) before visualization on Hyperfilm-ECL (Amersham)
by enhanced chemiluminescence (ECL, NEN, Boston, MA). After scanning,
semiquantitative densitometry was performed using Molecular Analyst
software (Bio-Rad). Mean density for senescent rats, fed normally or
food restricted, was expressed as a percentage of that obtained from
adult rats. Equal protein loading was verified by Coomassie blue
staining of the membranes at the end of the experiment.
Immunofluorescence. For indirect immunofluorescence, the kidneys were fixed by perfusion at a pressure of 120 mmHg through the abdominal aorta with a PBS solution containing 4% paraformaldehyde warmed to 37°C. The kidneys were sliced, postfixed overnight in PBS-4% paraformaldehyde, and washed extensively with PBS. Fixed kidney slices were infiltrated overnight in 30% sucrose and frozen in liquid nitrogen; then, 4-µm sections were collected on SuperFrost+ glass slides. Sections were preincubated in PBS containing 1% BSA (PBS/BSA) for 5 min, then incubated for 1 h at room temperature in PBS/BSA containing antibodies against AQP2 (1:300), p-AQP2 (1:50), or AQP3 (1:200), and washed for 3 × 10 min in PBS. Sections were then incubated for 45 min at room temperature in FITC-conjugated goat anti-rabbit antibodies (10 µg/ml, Promega), washed for 3 × 10 min, and mounted in 50% glycerol solution containing 2% N-propyl-gallate before observation under a fluorescence microscope (Olympus Van OX).
Electron microscopy. Kidneys were fixed by perfusion of PBS containing 4% paraformaldehyde and 0.1% glutaraldehyde, sliced, postfixed overnight in the same solution, and washed several times with PBS. Tissue pieces from the inner medulla were dehydrated in ethanol and embedded in unicryl, and 90-nm sections were then cut with an ultramicrotome and collected on electron microscopy grids. Sections were preincubated for 30 min at room temperature in 20 mM Tris buffer (pH 7.5) containing 0.1% BSA, 0.1% fish gelatin, 0.05% Tween 20, incubated in Tris buffer for 2 h with the anti-p-AQP2 antibody (1:100), and washed for 6 × 5 min with Tris buffer. Grids were then incubated in 10-nm colloidal gold-conjugated goat anti-rabbit antibodies in Tris buffer for 1 h and washed 6 × 5 min with Tris buffer. Sections were stained and observed under an electron microscope (Philips EM 400).
Statistics. Results were expressed as means ± SE, and differences were analyzed by ANOVA or Student's t-test. Statistical significance was considered at P < 0.05.
![]() |
RESULTS |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Effect of chronic food restriction on water balance and urine
osmolality.
From 10 to 30 mo, body weight significantly increased from 205 ± 5 to 245 ± 10 g in animals fed ad libitum (n = 8 in each group). In food-restricted rats, body weight progressively
decreased from 10 to 30 mo, to reach a final value of 152 ± 4 g (n = 8), that is a 50-g loss. These data are
consistent with those obtained on a larger cohort of animals, as
recently published (43). Data from metabolic cage
experiments are summarized in Table 1.
Food intake was comparable in 10- and 30-mo-old rats fed ad libitum, as
was total solute excretion. In control animals, urine flow rate
increased more than twofold from 10 to 30 mo, and urine osmolality was
reduced in proportion. The age-related loss of water was compensated for by a larger water intake in senescent rats. Chronic food
restriction prevented the age-related changes in water balance. Urinary
volume and osmolality were similar in food-restricted 30-mo-old rats and 10-mo-old normal animals. Food-restricted senescent rats had a
lower water intake than unrestricted senescent rats and even lower than
unrestricted adult rats. Total solute excretion was diminished by 30%,
the same proportion as food intake.
|
Plasma AVP, plasma osmolality, and papillary osmolality.
Plasma osmolality and AVP concentration were unchanged from 10 to 30 mo
in normally fed rats (Table 2). Long-term
food restriction doubled plasma AVP concentration and increased plasma
osmolality in senescent animals (Table 2). In animals fed ad libitum,
osmolality of whole papilla was significantly lower in 30-mo-old
compared with 10-mo-old rats (645 ± 48 and 1,084 ± 103 mosmol/kg H2O, respectively, n = 14/group). Food
restriction did not prevent this age-related decrease in papillary
osmolality (678 ± 35 mosmol/kg H2O, n = 17).
|
UT-A1, UT-A2, and UT-B1 expression.
In renal inner medulla, the 97-kDa form of UT-A1 and UT-B1 expression
was massively reduced (by 90%) in senescent rats fed ad libitum, and
this effect was maintained in food-restricted animals (Fig.
1). The 117-kDa isoform of UT-A1 was also
underexpressed but less intensively than the 97-kDa form.
|
|
AQP1, AQP2, AQP3, AQP4, and p-AQP2 expression.
AQP2 expression was reduced with age by 90% in rats fed ad libitum and
by 75% in chronically food-restricted animals (Figs. 3A and 6). Although AQP2
expression was twofold higher in food-restricted than in unrestricted
30-mo-old rats, the difference was not statistically significant (Figs.
3B and 6). AQP3 expression was decreased by 25% in
senescent animals. This change was prevented by chronic food
restriction (Figs. 4 and 6). However, the
difference between control and restricted 30-mo-olds rats was not
statistically significant. Expression of p-AQP2 was decreased by 90%
in 30-mo-old rats fed ad libitum compared with 10-mo-old rats (Figs.
5 and 6), and by 85% in food-restricted
senescent animals. The difference in p-AQP2 expression was not
significant between food-restricted and rats fed ad libitum (Figs. 5
and 6). AQP1 and AQP4 expression, unaltered from 10 to 30 mo in
normally fed rats, was not significantly modified by food restriction
(Fig. 6).
|
|
|
|
AQP2, AQP3, and p-AQP2 cellular localization.
In adult rats, AQP2 was mostly localized in the apical region of inner
medullary collecting duct (Fig.
7A, a). In
senescent rats fed ad libitum, AQP2 distribution was faint and diffuse
within the cytoplasm of principal cells (Fig. 7A,
b). In food-restricted senescent rats, the staining was
still weak, as expected from immunoblotting data, but AQP2 was totally
addressed to the apical membrane region of principal cells (Fig.
7A, c). A staining similar to that of AQP2 was
found by indirect immunofluorescence for p-AQP2 in the three groups of
rats (Fig. 7B). Electron microscopy in adult rats showed
that p-AQP2 was present mostly in the apical membrane of principal
cells, but some labeling was also observed in subapical vesicles (Fig.
8a). Localization of p-AQP2 in
the apical membrane was reduced in senescent rats fed normally (Fig. 8b), in contrast to food-restricted senescent rats, in which
it was even intensified compared with adult rats (Fig. 8c).
|
|
![]() |
DISCUSSION |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
The present study shows that chronic food restriction prevents polyuria and urinary concentrating defect in aging rats. In contrast, acute food restriction, which reduces urinary volume in adult and senescent rats, does not significantly affect the age-related decrease in urine osmolality. The corresponding molecular mechanisms involve a preferential targeting of AQP2 and p-AQP2 to the apical membrane of principal cells in the inner medullary collecting duct, in relation to a doubling of plasma AVP. In addition, AQP3 expression is increased in food-restricted rats, but AQP2 and p-AQP2 expression levels remain low. The low papillary osmolality in senescent rats is maintained during food restriction, consistent with low expression of UT-A1 and UT-B1.
Multiple factors could be involved in defective urine concentrating ability with age. They include dilution capacity of the thick ascending limb, AVP secretion, V2 receptor signaling, and corticopapillary gradient, as well as AQP and UT expression. In the present experimental model, which remains free of glomerulosclerosis and has a constant number of nephrons, impaired plasma AVP secretion and signaling are not the primary cause of defective urinary concentration (15). An abnormal number of V2 receptors or cAMP content in inner medulla can be also excluded (21, 30). In contrast, papillary osmolality is significantly reduced with age. In rodents and mammals, NaCl and urea contribute to the papillary hypertonicity required for water conservation. The expression of Na-K-2Cl transporter in senescent rat kidney is not documented, although previous studies have already suggested a reduced AVP-dependent NaCl reabsorption with age in rodents (14, 21). Urea accumulation in the renal medulla is permitted by urea transporters (36, 47, 50). It is initiated by the diffusion of urea from the terminal collecting duct lumen to the papillary interstitium, owing to the presence of UT-A1. UT-A2 and UT-B1, present in the thin descending limbs and the descending vasa recta, respectively, trap part of the urea escaping from the papilla, by countercurrent exchange with ascending vasa recta, and return it to the inner medulla. The present study shows that defective expression of UT-A1 and UT-B1 likely contributes to the observed low corticopapillary osmotic gradient of senescent rats. In addition, it reveals that the features of changes in senescence-induced polyuria are totally different from other polyuric situations.
In old rats, both 97- and 117-kDa glycosylated isoforms of UT-A1 (6a) are underexpressed. This pattern of changes differs from water loading, furosemide treatment, or low-protein diet, where 117-kDa is increased and the 97-kDa isoform is unchanged, especially in the tip of the inner medulla (44, 48, 49). The mechanisms underlying UT-A1 underexpression with age are still unknown. However, it can be speculated that high glucocorticoid levels reported in senescent rats would contribute to this downregulation, inasmuch as providing glucocorticoids to adrenalectomized rats results in a downregulation of terminal collecting duct urea permeability and UT-A1 protein (20, 26). The restoration of urine concentration by food restriction is not associated with normalization of UT-A1 expression.
Regulation of UT-B1 by urine concentrating activity is poorly documented. UT-B1 is also strongly downregulated in the inner medulla of senescent rats but not in the outer medulla. This change is opposite to that found in AVP-lacking Brattleboro rats that exhibited an UT-B1 mRNA abundance higher in inner medulla but lower in outer medulla than in AVP-infused Brattleboro rats (31).
UT-A2 expression in the thin descending limbs was upregulated in rats fed ad libitum but not in food-restricted senescent rats. An upregulation of UT-A2 expression was previously reported in Brattleboro rats treated with AVP (31, 52). This was mediated by an increase in medullary osmolality (24, 48) due to AVP. Unexpectely, in the present rat model, neither AVP nor medullary osmolality regulated UT-A2 abundance. In normally fed senescent rats, UT-A2 was upregulated without elevation of AVP plasma levels and with decreased inner medullary osmolality.
In food-restricted senescent rats, UT-A2 expression remained constant although both AVP levels and urine osmolality were increased. In senescent rats, urea reabsorption in inner medullary collecting ducts by UT-A1 is likely low, and medullary urea recycling cannot occur efficiently. In 30-mo-old rats, the increased UT-A2 expression may contribute to medullary urea dissipation instead of accumulation by bringing to medullary collecting ducts an amount of urea that cannot be captured by UT-A1 and that will be lost in urine. In food-restricted 30-mo-old rats, the absence of UT-A2 upregulation could minimize this adverse effect and hence contribute to the prevention of urinary concentrating defect.
In the aging kidney, in addition to a reduced papillary osmolality, a decrease in transepithelial water permeability of papillary collecting duct is suggested by AQP2 and AQP3 downregulation and AQP2 diffused localization in the cytoplasm of principal cells (30). Chronic food restriction prevents the decreased expression of AQP3 with age but not that of AQP2. However, chronic food restriction induces in senescent rats a recruitment of AQP2 to the apical membrane of principal cells. This membrane recruitment may restore papillary collecting duct water permeability, which would be lost with age. This greater transepithelial permeability is consistent with the enhanced AQP3 expression which, with AQP4, contributes to the water permeability of the basolateral membrane of principal cells.
AVP-dependent insertion of AQP2 in the luminal membrane is the main mechanism for control of water permeability in collecting duct principal cells. AVP binding to basolateral V2 receptor stimulates cAMP generation from adenylyl cyclase, which is responsible for the fusion of subapical vesicles containing AQP2 to the apical membrane of the cells. Plasma osmolality and plasma AVP are unchanged from 10 to 30 mo (15, 30), but they are significantly increased in long-term food-restricted senescent rats. This chronic higher plasma AVP level may be responsible for the marked recruitment of AQP2 to the apical membrane. Besides, without being fully restored, the expression of AQP2 is weakly increased, maybe via cAMP-dependent transcriptional regulation (51).
Long-term food restriction is associated with a 30% decreased water intake in 30-mo-old rats. This mild chronic dehydration may be responsible for the higher plasma osmolality and AVP level observed in these rats. Food restriction in senescent rats induces a decreased water consumption both in long-term and short-term protocols. Short-term reduction of water intake does not improve the age-related kidney concentrating defect. Therefore, chronic food restriction may be an adaptive process, involving a mild water restriction and an increased plasma AVP release. However, the present study cannot rule out that the improvement of urinary concentrating ability in the aging kidney by long-term food restriction may be due directly to the effect of chronic reduction on water intake.
AVP-induced phosphorylation of AQP2 at serine-256 by protein kinase A is critically involved in its localization in the apical membrane. As described before in normal rat kidney (8), p-AQP2 has been detected, in the present study, both in apical plasma membrane and in intracellular vesicles in noninduced principal cells of adult and senescent rats fed ad libitum. In chronic food restriction, p-AQP2 is also localized in subapical vesicles of collecting duct principal cells but is strongly recruited in apical membrane. The expression of p-AQP2 is not significantly modified despite the AVP increase in chronic food restriction, consistent with the unchanged p-AQP2 level in normal rats treated with [desamino-Cys1, D-Arg8]AVP (8).
In summary, chronic food restriction prevents kidney concentrating defect in female WAG/Rij rats. However, it cannot be excluded that this effect may be secondary to the long duration of the mild water intake depletion associated with food restriction. The molecular mechanisms of this urinary concentrating ability rescue in aging kidney involve an AVP-dependent recruitment of AQP2 and p-AQP2 to the plasma membrane of principal cells, independently of the expression of AQP2 and UTs or changes in the corticopapillary osmotic gradient. This favors a dual mechanism for the control of water homeostasis in aging rats: one depending on plasma AVP level and related to membrane targeting of AQP2 and p-AQP2 and another associated with changes in AQP2 and UT expression and with a low corticopapillary gradient, independent of AVP level.
![]() |
ACKNOWLEDGEMENTS |
---|
We thank Drs. S. Le Maout and F. Tacnet (CEA, Gif-sur-Yvette, France) for providing us with AQP2 and AQP3 antibodies and Ferring Pharmaceuticals (Malmö, Sweden) for the gift of arginine-AVP. We are grateful to J.-C. Robillard and P. Héry for outstanding animal care and to G. Augoyard for skills in performing AVP assays.
![]() |
FOOTNOTES |
---|
This work was funded by a Training and Mobility of Research ERBFMRXCT970128 from the European Union. L. Teillet was supported by a grant from the Assistance Publique-Hôpitaux de Paris, Direction de la Recherche Clinique, Paris, France.
Address for reprint requests and other correspondence: J.-M. Verbavatz, Service de Biologie Cellulaire Bat. 532, CEA/Saclay, F-91191 Gif-sur-Yvette cedex, France (E-mail: jmverbavatz{at}cea.fr).
The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
First published August 8, 2001;10.1152/ajprenal.00139.2001
Received 4 May 2001; accepted in final form 9 August 2001.
![]() |
REFERENCES |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
1.
Apostol, E,
Ecelbarger C,
Terris J,
Bradford A,
Andrews P,
and
Knepper M.
Reduced renal medullary water channel expression in puromycin aminonucleoside-induced nephrotic syndrome.
J Am Soc Nephrol
8:
15-24,
1997[Abstract].
2.
Bankir, L,
and
Trinh-Trang-Tan MM.
Urea and the kidney.
In: The Kidney (6th ed.), edited by Brenner BM,
and Rector FC.. Philadelphia, PA: Saunders, 2000, p. 637-679.
3.
Bardoux, P,
Ahloulay M,
Le Maout S,
Bankir L,
and
Trinh-Trang-Tan MM.
Aquaporin-2 and urea transporter-A1 are upregulated in rats with type I diabetes mellitus.
Diabetologia.
44:
637-645,
2001[ISI][Medline].
4.
Baylis, C,
and
Corman B.
The aging kidney: insights from experimental studies.
J Am Soc Nephrol
9:
699-709,
1998[Abstract].
5.
Beck, LH.
Changes in renal function with aging.
Clin Geriatr Med
14:
199-209,
1998[ISI][Medline].
6.
Beck, N,
and
Yu BP.
Effect of aging on urinary concentration mechanism and vasopressin-dependent cAMP in rats.
Am J Physiol Renal Fluid Electrolyte Physiol
243:
F121-F125,
1982
6a.
Bradford, AD,
Terris JM,
Ecelbarger CA,
Klein JD,
Sands JM,
Chou CL,
and
Knepper MA.
97- and 117-kDa forms of collecting duct urea transporter UT-A1 are due to different states of glysosylation.
Am J Physiol Renal Physiol
281:
F133-F143,
2001
7.
Burek, JD.
Pathology of Aging Rats. Boca Raton, FL: CRC, 1978.
8.
Christensen, BM,
Zelenina M,
Aperia A,
and
Nielsen S.
Localization and regulation of PKA-phosphorylated AQP2 in response to V(2)-receptor agonist/antagonist treatment.
Am J Physiol Renal Physiol
278:
F29-F42,
2000
9.
Corman, B,
and
Michel JB.
Glomerular filtration, renal blood flow and solute excretion in conscious aging rats.
Am J Physiol Regulatory Integrative Comp Physiol
253:
R555-R560,
1987
10.
Corman, B,
and
Roinel N.
Single-nephron filtration rate and proximal reabsorption in aging rats.
Am J Physiol Renal Fluid Electrolyte Physiol
260:
F75-F80,
1991
11.
Corman, B,
Roinel N,
and
Geelen G.
Plasma vasopressin and cortical nephron function in aging rats.
Mech Ageing Dev
62:
263-277,
1992[ISI][Medline].
12.
Davidson, Y,
Davies I,
and
Goddard C.
Renal vasopressin receptors in ageing C57BL/Icrfa mice.
J Endocrinol
115:
379-385,
1987[Abstract].
13.
Deen, PM,
Verdijk MA,
Knoers NV,
Wieringa B,
Monnens LA,
van Os CH,
and
van Oost BA.
Requirement of human renal water channel aquaporin-2 for vasopressin-dependent concentration of urine.
Science
264:
92-95,
1994[ISI][Medline].
14.
Di Stefano, A,
Wittner M,
and
Corman B.
Vasopressin-stimulation of NaCl transport in the medullary thick ascending limb of Henle's loop is decreased in aging mice.
Pflügers Arch
419:
327-331,
1991[ISI][Medline].
15.
Geelen, G,
and
Corman B.
Relationship between vasopressin and renal concentrating ability in aging rats.
Am J Physiol Regulatory Integrative Comp Physiol
262:
R826-R833,
1992
16.
Hediger, MA,
Smith CP,
You G,
Lee WS,
Kanai Y,
and
Shayakul C.
Structure, regulation and physiological roles of urea transporters.
Kidney Int
49:
1615-1623,
1996[ISI][Medline].
17.
Hu, MC,
Bankir L,
Michelet S,
Rousselet G,
and
Trinh-Trang-Tan MM.
Massive reduction of urea transporters in remnant kidney and brain of uremic rats.
Kidney Int
58:
1202-1210,
2000[ISI][Medline].
18.
Karakashian, A,
Timmer RT,
Klein JD,
Gunn RB,
Sands JM,
and
Bagnasco S.
Cloning and characterization of two new isoforms of rat kidney urea transporter: UT-A3 and UT-A4.
J Am Soc Nephrol
10:
230-237,
1999
19.
Keil, LC,
and
Severs WB.
Reduction in plasma vasopressin levels of dehydrated rats following acute stress.
Endocrinology
100:
30-38,
1977[Abstract].
20.
Klein, JD,
Price SR,
Bailey JL,
Jacobs JD,
and
Sands JM.
Glucocorticoids mediate a decrease in AVP-regulated urea transporter in diabetic rat inner medulla.
Am J Physiol Renal Physiol
273:
F949-F953,
1997[ISI][Medline].
21.
Klingler, C,
Preisser Barrault M,
Lluel P,
Horgen L,
Teillet L,
Ancellin N,
and
Corman B.
Vasopressin V2 receptor mRNA expression and cAMP accumulation in aging rat kidney.
Am J Physiol Regulatory Integrative Comp Physiol
272:
R1775-R1782,
1997
22.
Knepper, MA,
and
Inoue T.
Regulation of aquaporin-2 water channel trafficking by vasopressin.
Curr Opin Cell Biol
9:
560-564,
1997[ISI][Medline].
23.
Knepper, MA,
and
Roch-Ramel F.
Pathways of urea transport in the mammalian kidney.
Kidney Int
31:
629-633,
1987[ISI][Medline].
24.
Leroy, C,
Basset G,
Gruel G,
Ripoche P,
Trinh-Trang-Tan MM,
and
Rousselet G.
Hyperosmotic NaCl and urea synergistically regulate the expression of the UT-A2 urea transporter in vitro and in vivo.
Biochem Biophys Res Commun
271:
368-373,
2000[ISI][Medline].
25.
Miller, M.
Influence of aging on vasopressin secretion and water regulation.
In: Vasopressin, edited by Schrier., 1985, p. 249-258.
26.
Naruse, M,
Klein JD,
Ashkar ZM,
Jacobs JD,
and
Sands JM.
Glucocorticoids downregulate the vasopressin-regulated urea transporter in rat terminal inner medullary collecting ducts.
J Am Soc Nephrol
8:
517-523,
1997[Abstract].
27.
Nielsen, S,
Fror J,
and
Knepper MA.
Renal aquaporins: key roles in water balance and water balance disorders.
Curr Opin Nephrol Hypertens
7:
509-516,
1998[ISI][Medline].
28.
Nielsen, S,
Smith BL,
Christensen EI,
Knepper MA,
and
Agre P.
CHIP28 water channels are localized in constitutively water-permeable segments of the nephron.
J Cell Biol
120:
371-383,
1993[Abstract].
29.
Nielsen, S,
Terris J,
Smith CP,
Hediger MA,
Ecelbarger CA,
and
Knepper MA.
Cellular and subcellular localization of the vasopressin-regulated urea transporter in rat kidney.
Proc Natl Acad Sci USA
93:
5495-5500,
1996
30.
Preisser, L,
Teillet L,
Aliotti S,
Gobin R,
Berthonaud V,
Chevalier J,
Corman B,
and
Verbavatz JM.
Downregulation of aquaporin-2 and -3 in aging kidney is independent of V2 vasopressin receptor.
Am J Physiol Renal Physiol
279:
F144-F152,
2000
31.
Promeneur, D,
Bankir L,
Hu MC,
and
Trinh-Trang-Tan MM.
Renal tubular and vascular urea transporters: influence of antidiuretic hormone on messenger RNA expression in Brattleboro rats.
J Am Soc Nephrol
9:
1359-1366,
1998[Abstract].
32.
Ravid, R,
Fliers E,
Swaab D,
and
Zurcher C.
Changes in vasopressin and testosterone in the senescent Brown-Norway (BN/Bi) rats.
Gerontology
33:
87-98,
1987[ISI][Medline].
33.
Roudier, N,
Verbavatz JM,
Maurel C,
Ripoche P,
and
Tacnet F.
Evidence for the presence of aquaporin-3 in human red blood cells.
J Biol Chem
273:
8407-8412,
1998
35.
Sabolic, I,
Valenti G,
Verbavatz JM,
Van Hoek AN,
Verkman AS,
and
Brown D.
Localization of the CHIP28 water channel in rat kidney.
Am J Physiol Cell Physiol
263:
C1225-C1233,
1992
36.
Sands, JM.
Regulation of urea transporter proteins in kidney and liver.
Mt Sinai J Med
67:
112-119,
2000[ISI][Medline].
37.
Sands, JM,
Timmer RT,
and
Gunn RB.
Urea transporters in kidney and erythrocytes.
Am J Physiol Renal Physiol
273:
F321-F339,
1997
38.
Shayakul, C,
Knepper MA,
Smith CP,
DiGiovanni SR,
and
Hediger MA.
Segmental localization of urea transporter mRNAs in rat kidney.
Am J Physiol Renal Physiol
272:
F654-F660,
1997
39.
Shayakul, C,
Tsukaguchi H,
Berger UV,
and
Hediger M.
Molecular characterization of a novel urea transporter from kidney inner medullary collecting ducts.
Am J Physiol Renal Physiol
280:
F487-F494,
2001
40.
Sladek, C,
McNeill T,
Gregg C,
Blair M,
and
Baggs R.
Vasopressin and renin response to dehydration in aged rats.
Neurobiol Aging
2:
293-302,
1981[ISI][Medline].
41.
Smith, CP,
Lee WS,
Martial S,
Knepper MA,
You G,
Sands JM,
and
Hediger MA.
Cloning and regulation of expression of the rat kidney urea transporter (rUT2).
J Clin Invest
96:
1556-1563,
1995[ISI][Medline].
42.
Swenson, K,
Sands J,
Jacobs J,
and
Sladek C.
Effect of aging on vasopressin and aquaporin responses to dehydration in Fisher 344-Brown-Norway F1 rats.
Am J Physiol Regulatory Integrative Comp Physiol
273:
R35-R40,
1997
43.
Teillet, L,
Verbeke P,
Gouraud S,
Bakala H,
Borot-Laloi C,
Heudes D,
Bruneval P,
and
Corman B.
Food restriction prevents advanced glycation end product accumulation and retards kidney aging in lean rats.
J Am Soc Nephrol
11:
1488-1497,
2000
44.
Terris, J,
Ecelbarger CA,
Sands JM,
and
Knepper MA.
Long-term regulation of renal urea transporter protein expression in rat.
J Am Soc Nephrol
9:
729-736,
1998[Abstract].
45.
Terris, JM,
Knepper MK,
and
Wade J.
UT-A3: localization and characterization of an additional urea transporter isoform in the IMCD.
Am J Physiol Renal Physiol
280:
F325-F332,
2001
46.
Terwell, D,
Ten Haff J,
Markerink M,
and
Jolles J.
Changes in plasma vasopressin concentration and plasma osmolality in relation to age and time of the day in the male Wistar rat.
Acta Endocrinol
126:
357-362,
1992[ISI][Medline].
47.
Trinh-Trang-Tan, MM,
and
Bankir L.
Integrated function of urea transporters in the mammalian kidney.
Exp Nephrol
6:
471-479,
1998[ISI][Medline].
48.
Trinh-Trang-Tan, MM,
Jegou T,
Ugnon-Café S,
Bankir L,
and
Rousselet G.
Regulation of UT-A2 protein in vivo and in vitro (Abstract).
J Am Soc Nephrol
11:
23A,
2000.
49.
Trinh-Trang-Tan, MM,
Ugnon-Cafe S,
Bankir L,
and
Rousselet G.
dDAVP promotes a shift in UT-A1 expression from terminal to initial inner medullary collecting duct (IMCD) (Abstract).
J Am Soc Nephrol
11:
23A,
2000.
50.
Tsukaguchi, H,
Shayakul C,
Berger UV,
and
Hediger MA.
Urea transporters in kidney: molecular analysis and contribution to the urinary concentrating process.
Am J Physiol Renal Physiol
275:
F319-F324,
1998
51.
Uchida, S,
Sasaki S,
Fushimi K,
and
Marumo F.
Isolation of human aquaporin-CD gene.
J Biol Chem
269:
23451-23455,
1994
52.
Wade, JB,
Lee AJ,
Liu J,
Ecelbarger CA,
Mitchell C,
Bradford AD,
Terris J,
Kim GH,
and
Knepper MA.
UT-A2: a 55-kDa urea transporter in thin descending limb whose abundance is regulated by vasopressin.
Am J Physiol Renal Physiol
278:
F52-F62,
2000
53.
Wilson, PD,
and
Dillingham MA.
Age-associated decrease in vasopressin-induced renal water transport: a role for adenylate cyclase and G protein malfunction.
Gerontology
38:
315-321,
1992[ISI][Medline].
54.
Xu, Y,
Olives B,
Bailly P,
Fischer E,
Ripoche P,
Ronco P,
Cartron JP,
and
Rondeau E.
Endothelial cells of the kidney vasa recta express the urea transporter HUT11.
Kidney Int
51:
138-146,
1997[ISI][Medline].
55.
Zbuzek, VK,
Zbuzek V,
and
Wu W.
The effect of aging on vasopressin system in Fisher 344 rats.
Exp Gerontol
18:
305-311,
1983[ISI][Medline].