UNIPROT:P41181 - FACTA Search

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Query: UNIPROT:P41181 (collecting duct)
5,183 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Hyponatremia is associated with inappropriately elevated vasopressin levels. A brisk natriuresis precedes the escape from this antidiuresis. Thus, the hypothesis was that the abundance of one or more of the sodium transporters of the distal tubule (a site for fine tuning of sodium balance) would be altered during vasopressin escape. Semiquantitative immunoblotting was used to examine the regulation of abundance of several sodium transporters/channels of the thick ascending limb through the collecting duct in the rat model. Osmotic minipumps to infuse dDAVP, the V2-selective vasopressin agonist (5 ng/h) for the entire experiment, were implanted in Male Sprague-Dawley rats. After 4 d, rats were divided into a control (dry AIN-76 diet/ad libitum water) or a water-loaded (gelled-agar-AIN-76 diet/ad libitum water) group. Rats were killed after 1, 2, 3, or 7 additional days. The water-loaded rats were hyponatremic (plasma Na+, 98 to 122 mmol/L) and manifested the expected early natriuresis and diuresis of vasopressin escape. Water loading (with dDAVP infusion) resulted in increased whole-kidney abundances of the thiazide-sensitive Na-Cl co-transporter, the alpha-subunit of the epithelial sodium channel (ENaC), and the 70-kD dimer of the gamma-subunit of ENaC. No changes were observed for the ss-subunit of ENaC. Similar protein changes have recently been associated with elevated aldosterone levels in rats. However, plasma aldosterone levels were significantly suppressed in this model. These data suggest that several distal sodium reabsorptive mechanisms are upregulated during vasopressin escape; this may help to attenuate the developing hyponatremia resulting from water loading when vasopressin levels are inappropriately elevated.
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PMID:Increased abundance of distal sodium transporters in rat kidney during vasopressin escape. 1115 10

The Na+-H+ exchanger NHE3 and the thiazide-sensitive Na+-Cl- cotransporter NCC are the major apical sodium transporters in the proximal convoluted tubule and the distal convoluted tubule of the kidney, respectively. We investigated the mechanism of compensation that allows maintenance of sodium balance in NHE3 knockout mice and in NCC knockout mice. We used a so-called 'targeted proteomics' approach, which profiles the entire renal tubule with regard to changes in Na+ transporter and aquaporin abundance in response to the gene deletions. Specific antibodies to the Na+ transporters and aquaporins expressed along the nephron were utilized to determine the relative abundance of each transporter. Semiquantitative immunoblotting was used which gives an estimate of the percentage change in abundance of each transporter in knockout compared with wild-type mice. In NHE3 knockout mice three changes were identified which could compensate for the loss of NHE3-mediated sodium absorption. (a) The proximal sodium-phosphate cotransporter NaPi-2 was markedly upregulated. (b) In the collecting duct, the 70 kDa form of the y-subunit of the epithelial sodium channel, ENaC, exhibited an increase in abundance. This is thought to be an aldosterone-stimulated form of y-ENaC. (c) Glomerular filtration was significantly reduced. In the NCC knockout mice, amongst all the sodium transporters expressed along the renal tubule, only the 70 kDa form of the y-subunit of the epithelial sodium channel, ENaC, exhibited an increase in abundance. In conclusion, both mouse knockout models demonstrated successful compensation for loss of the deleted transporter. More extensive adaptation occurred in the case of the NHE3 knockout, presumably because NHE3 is responsible for much more sodium absorption in normal mice than in NCC knockout mice.
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PMID:Profiling of renal tubule Na+ transporter abundances in NHE3 and NCC null mice using targeted proteomics. 1115 68

We sought to assess whether the distal convoluted tubule (DCT) segment of the rabbit nephron expresses a functional epithelial sodium channel. First, the transepithelial voltage (V(te), lumen vs. bath) was measured in isolated perfused DCT segments (assessed separately in the upstream half and the downstream half of the DCT). V(te) was zero and not affected by amiloride or barium in the upstream DCT. V(te) was sometimes negative in the downstream DCT and depolarized by amiloride and hyperpolarized by barium, suggesting inclusion of connecting tubule (CNT) cells. To determine expression of epithelial sodium channel (ENaC) mRNA subunits by the upstream DCT, rabbit alpha-, beta-, and gamma-ENaC cDNA fragments were cloned and primers were selected for single-nephron RT-PCR analysis. Although alpha-ENaC was expressed by the DCT, beta- and gamma-ENaC were not detected in the DCT. In contrast, the CNT, CCD, and outer medullary collecting duct (OMCD) expressed all three subunits. Nedd4 was also not detected in the DCT but was expressed by the CNT, CCD, and OMCD. When upstream DCT fragments were grown to confluent monolayers in primary culture, the epithelia exhibited negative voltages and high transepithelial resistances and expressed mRNA for all three ENaC subunits as well as for Nedd4. The absence of a negative voltage and failure to detect transcript for beta- and gamma-ENaC and Nedd4 in the native rabbit DCT suggest that the sodium channel is not a significant pathway for sodium absorption by this segment. The phenotype conversion observed when DCT cells are grown in culture does not rule out the possibility that there may be conditions in which the DCT in the intact kidney expresses sodium channel activity. The results are consistent with the notion that DCT sodium transport is predominantly, if not exclusively, electroneutral.
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PMID:The distal convoluted tubule of rabbit kidney does not express a functional sodium channel. 1118 16

Aldosterone stimulates Na(+) reabsorption in the collecting ducts by increasing the activity of the epithelial sodium channel, ENaC. Systemic administration of aldosterone increases alpha ENaC mRNA expression in mammalian kidney, suggesting that the alpha ENaC gene is a target for aldosterone action in the distal nephron. To determine whether aldosterone increases alpha ENaC gene transcription, a portion of the alpha ENaC 5'- flanking region coupled to luciferase was transfected into MDCK-C7 cells, a collecting duct cell line with aldosterone-stimulated Na(+) transport. Both dexamethasone and aldosterone stimulated alpha ENaC-coupled reporter gene activity via the glucocorticoid receptor (GR), and this response correlated with the effect of these hormones on endogenous alpha ENaC expression. The aldosterone-stimulated alpha ENaC expression was blocked by actinomycin D, and aldosterone had no effect on alpha ENaC mRNA decay, confirming a transcriptional effect. In HT-29 cells, a GR/mineralocorticoid receptor (MR)-deficient colonic cell line with constitutive alpha ENaC expression, cotransfection with GR or MR restored aldosterone-stimulated alpha ENaC gene transcription, although aldosterone had a functional preference for MR. Analysis of deletion constructs confirmed that a single imperfect glucocorticoid response element (GRE) is necessary and sufficient to confer the aldosterone responsiveness to the alpha ENaC gene promoter in MDCK-C7 and HT-29 cells. These results confirm that alpha ENaC is an aldosterone-induced transcript in the collecting duct and delineates the molecular mechanism for this effect.
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PMID:The alpha-subunit of the epithelial sodium channel is an aldosterone-induced transcript in mammalian collecting ducts, and this transcriptional response is mediated via distinct cis-elements in the 5'-flanking region of the gene. 1126 9

The effect of L-arginine on transepithelial ion transport was examined in cultured M-1 mouse renal cortical collecting duct (CCD) cells using continuous short circuit current (Isc) measurements in HCO3-/CO2 buffered solution. Steady state Isc averaged 73.8 +/- 3.2 microA/cm2 (n = 126) and was reduced by 94 +/- 0.6% (n = 16) by the apical addition of 100 microM amiloride. This confirms that the predominant electrogenic ion transport in M-1 cells is Na+ absorption via the epithelial sodium channel (ENaC). Experiments using the cationic amino acid L-lysine (radiolabeled) as a stable arginine analogue show that the combined activity of an apical system y+ and a basal amino acid transport system y+L are responsible for most cationic amino acid transport across M-1 cells. Together they generate net absorptive cationic amino acid flux. Application of L-arginine (10 mM) either apically or basolaterally induced a transient peak increase in Isc averaging 36.6 +/- 5.4 microA/cm2 (n = 19) and 32.0 +/- 7.2 microA/cm2 (n = 8), respectively. The response was preserved in the absence of bath Cl- (n = 4), but was abolished either in the absence of apical Na+ (n = 4) or by apical addition of 100 microM amiloride (n = 6). L-lysine, which cannot serve as a precursor of NO, caused a response similar to that of L-arginine (n = 4); neither L-NMMA (100 microM; n = 3) nor L-NAME (1 mM; n = 4) (both NO-synthase inhibitors) affected the Isc response to L-arginine. The effects of arginine or lysine were replicated by alkalinization that mimicked the transient alkalinization of the bath solution upon addition of these amino acids. We conclude that in M-1 cells L-arginine stimulates Na+ absorption via a pH-dependent, but NO-independent mechanism. The observed net cationic amino acid absorption will counteract passive cationic amino acid leak into the CCD in the presence of electrogenic Na+ transport, consistent with reports of stimulated expression of Na+ and cationic amino acid transporters by aldosterone.
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PMID:L-arginine effects on Na+ transport in M-1 mouse cortical collecting duct cells--a cationic amino acid absorbing epithelium. 1131 95

Epithelial sodium channel (ENaC) subunit (alpha, beta, and gamma) mRNA and protein have been localized to the principal cells of the connecting tubule (CNT), cortical collecting duct (CCD), and outer medullary collecting duct (OMCD) in rat kidney. However, the subcellular localization of ENaC subunits in the principal cells of these cells is undefined. The cellular and subcellular localization of ENaC subunits in rat kidney was therefore examined. Immunocytochemistry demonstrated the presence of all three subunits in principal cells of the CNT, CCD, OMCD, and IMCD. In cortex and outer medulla, confocal microscopy demonstrated a difference in the subcellular localization of subunits. alpha-ENaC was localized mainly in a zone in the apical domains, whereas beta- and gamma-ENaC were found throughout the cytoplasm. Immunoelectron microscopy confirmed the presence of ENaC subunits in both the apical plasma membrane and intracellular vesicles. In contrast to the labeling pattern seen in cortex, alpha-ENaC labeling in IMCD cells was distributed throughout the cytoplasm. In the urothelium covering pelvis, ureters, and bladder, immunoperoxidase and confocal microscopy revealed differences the presence of all ENaC subunits. As seen in CCD, alpha-ENaC was present in a narrow zone near the apical plasma membrane, whereas beta- and gamma-ENaC were dispersed throughout the cytoplasm. In conclusion, all three subunits of ENaC are expressed throughout the collecting duct (CD), including the IMCD as well as in the urothelium. The intracellular vesicular pool in CD principal cells suggests ENaC trafficking as a potential mechanism for the regulation of Na(+) reabsorption.
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PMID:Immunocytochemical and immunoelectron microscopic localization of alpha-, beta-, and gamma-ENaC in rat kidney. 1135 48

We have examined the respective influence of aldosterone, vasopressin and cell sodium delivery on Na+,K+-ATPase expression. The level of expression of the mRNA encoding for the alpha1- and beta1-subunits of Na+,K+-ATPase was evaluated in cortical collecting duct (CCD) cells from rats under different aldosterone status, in cells from the rat CCD cell line RCCD1 treated or not with vasopressin and in CCD cells from mice inactivated or not for the a-subunit of the epithelial sodium channel. The amount of mRNA was determined by in situ hybridization. Both aldosterone and vasopressin up-regulate transcripts encoding for the alpha1-subunit of Na+,K+-ATPase while beta1 is unaltered. Interestingly, when cell sodium entry was largely reduced (alphaENaC knock-out mice), the amount of transcripts encoding for the alpha1-subunit of Na+,K+-ATPase was significantly decreased in spite of high plasma aldosterone concentrations. No effect was observed on beta1-subunit. Altogether, these results suggest a coordinated hormonal and ionic control of Na+,K+-ATPase expression by different transcriptional pathways (steroid-receptor, cAMP-dependent and Na+dependent) in CCD cells. These regulations affect only alpha1-subunit of Na,K+-ATPase but not beta1.
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PMID:Coordinate control of Na,K-atpase mRNA expression by aldosterone, vasopressin and cell sodium delivery in the cortical collecting duct. 1135 97

The aldosterone-sensitive distal nephron extends from the second part of the distal convoluted tubule to the inner medullary collecting duct. As recently shown, aldosterone increases within two hours the abundance of the alpha-subunit of the epithelial sodium channel along the entire aldosterone-sensitive distal nephron, whereas it induces only in an initial portion of the aldosterone-sensitive distal nephron an apical translocation of all three epithelial sodium channel subunits. This suggests that another factor or factors determines the length of the aldosterone-sensitive distal nephron portion in which aldosterone controls epithelial sodium channel surface expression. Since the glucocorticoid-induced kinase SGK1 was identified as aldosterone-induced protein in 1999, it has been postulated to play a key regulatory role. The in-vivo localization of its induction to segment-specific cells of the aldosterone-sensitive distal nephron, and the in-vitro correlation of the amount of its hyperphosphorylated form with transepithelial sodium transport, support this hypothesis. Other recent studies unravel pathways other than those activated by aldosterone and insulin that impact on SGK1 expression and/or function, and thus shed some light onto the complex network that appears to control sodium transport. In view of the ongoing research, the question of how, and formally also whether, SGK1 acts on the epithelial sodium channel should be resolved in the near future.
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PMID:Mediators of aldosterone action in the renal tubule. 1149 63

The regulation of plasma membrane Na(+)-K(+)-ATPases (NKA) expression by aldosterone and arginin vasopressin (AVP) in the cortical collecting duct (CCD) has been examined in a new rat CCD cell line, designated as RCCD(2). This cell line has maintained many characteristics of the CCD-in particular, the expression of the mineralocorticoid receptor. Mineralocorticoid receptor is expressed at the protein level and binds (3)H-aldosterone (approximately 15 to 20 fmol/mg protein). Short-circuit current (Isc) experiments showed approximately a twofold increase in Isc associated with a decrease in transepithelial resistance when cells were treated with aldosterone concentrations as low as 10(-9) M. This effect on Isc was significant 2 h after aldosterone addition and was still present after 24 h. It was accompanied by an increase in the amount of mRNA encoding for the alpha subunit of the epithelial sodium channel (sixfold) and the alpha1 subunit of NKA (fourfold) after 24 h of hormone treatment. In addition, mRNA expression of the serum- and glucocorticoid-induced kinase (Sgk) was increased by 10(-9) M aldosterone treatment as early as 45 min after hormone addition. As had already been documented in native CCD obtained by microdissection, incubation of RCCD(2) cells for 24 h with aldosterone resulted in the constitution of a latent pool of NKA that could be rapidly recruited by AVP (15 min). NKA biotinylation experiments and preparation of membrane fractions show that this latent pool of NKA is present in the intracellular compartment of the cells and is recruited by AVP in the basolateral membrane through a translocation process. This mechanism is accompanied by dephosphorylation of the alpha(1) catalytic subunit of NKA.
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PMID:Basolateral translocation by vasopressin of the aldosterone-induced pool of latent Na-K-ATPases is accompanied by alpha1 subunit dephosphorylation: study in a new aldosterone-sensitive rat cortical collecting duct cell line. 1151 73

Vasopressin plays a role in both salt and water balance in the kidney. Classic studies, utilizing isolated perfused tubules, have revealed that vasopressin increases sodium reabsorption in the kidney thick ascending limb and the collecting duct. Furthermore, the activity of several sodium transport proteins expressed in these segments, such as the bumetanide-sensitive Na-K-2Cl cotransporter (NKCC2) and the epithelial sodium channel (ENaC), have been shown to be directly increased by vasopressin. Increased protein abundance might be one means through which sodium transporter and channel activity is enhanced. We have used immunoblotting and immunohistochemistry in order to investigate the regulation of abundance of the major sodium transporters and channels expressed along the renal tubule in response to vasopressin. Chronic (7-day) studies were performed in which vasopressin levels were elevated either endogenously by water restriction of Sprague-Dawley rats or exogenously through infusion of the vasopressin V2-receptor-selective agonist, dDAVP (1-deamino-8d-arginine-vasopressin), to Brattleboro rats. We found a significant increase in protein abundance for NKCC2 and the beta- and gamma-subunits of ENaC with either water restriction or dDAVP infusion. The alpha-subunit of Na-K-ATPase was increased by water restriction, but not by dDAVP infusion, and alpha-ENaC and the thiazide-sensitive cotransporter (NCC) were increased by dDAVP infusion but not by water restriction. Acute (60-min) in vivo exposure to dDAVP led to an increase in both beta- and gamma-ENaC abundance in kidney cortex homogenates, displaying the rapid nature of some of these changes. Overall these increases in sodium transporter and channel abundances likely contribute to both the antidiuretic and antinatriuretic actions of vasopressin.
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PMID:Regulation of the abundance of renal sodium transporters and channels by vasopressin. 1157 75

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