ATP-dependent proton transport in human renal medulla

1986 ◽  
Vol 251 (2) ◽  
pp. F297-F302
Author(s):  
F. D. Diaz-Diaz ◽  
E. F. LaBelle ◽  
D. C. Eaton ◽  
T. D. DuBose
Keyword(s):  
Urinary Bladder ◽  
Proton Transport ◽  
Collecting Duct ◽  
Proton Translocation ◽  
Renal Medulla ◽  
Absorbance Changes ◽  
Medullary Collecting Duct ◽  
Proton Uptake ◽  
Hydrogen Ion Transport ◽  
Proton Translocating Atpase

An electrogenic proton-translocating ATPase (H+-ATPase) has been described in turtle urinary bladder and bovine and rat renal medulla. In the present study, a membrane fraction with ATP-dependent H+ transport activity was isolated from human renal medulla. Intravesicular acidification was assessed by acridine orange absorbance changes. Proton transport was abolished by N-ethylmaleimide but not oligomycin or vanadate, differentiating this H+-ATPase from mitochondrial F0-F1 H+-ATPase and gastric H+-K+-ATPase. In addition, vesicular proton uptake was demonstrated to be independent of sodium and potassium cotransport. Proton translocation rate increased when transmembrane potential was clamped with valinomycin supporting an electrogenic mechanism. Hydrogen ion transport was dependent on the presence of chloride or bromide, since substitution by fluoride or nitrate markedly decreased intravesicular acidification. The transport characteristics of this proton-translocating ATPase are similar to those described for turtle urinary bladder and bovine and rat renal medulla, which have been assumed to play a role in urinary acidification by the medullary collecting duct.

Endothelin receptor mRNA expression in renal medulla identified by in situ RT-PCR

1995 ◽  
Vol 269 (3) ◽  
pp. F449-F457 ◽  
Author(s):  
L. H. Chow ◽  
S. Subramanian ◽  
G. J. Nuovo ◽  
F. Miller ◽  
E. P. Nord
Keyword(s):  
Mrna Expression ◽  
Collecting Duct ◽  
Renal Medulla ◽  
Receptor Subtypes ◽  
Rt Pcr ◽  
Receptor Mrna ◽  
Pcr Products ◽  
Medullary Collecting Duct ◽  
Labeled Probe

Three subtypes of endothelin (ET) receptors have been identified by cDNA cloning, namely ET-RA, ET-RB, and ET-RC. In the current study the precise cellular distribution of the ET receptor subtypes in the renal medulla was explored by detecting the corresponding polymerase chain reaction (PCR)-amplified cDNAs by in situ reverse transcription (RT)-PCR. The PCR-amplified cDNAs were detected either by direct incorporation using digoxigenin-dUTP (dig-dUTP) as a nucleotide substrate in the PCR reaction or by in situ hybridization with the dig-dUTP-labeled probe. ET-RB mRNA was detected exclusively in the epithelial cells of the inner and outer medullary collecting duct. In contrast, ET-RA message was observed primarily in interstitial cells and pericytes of the vasae rectae in the outer and inner medulla. Southern blot analysis of PCR-amplified cDNAs reverse transcribed from extracted RNA of rat renal medulla confirmed the specificity of the RT-PCR products. ET-RC mRNA was not detected. We conclude that ET-RB is the major ET receptor found in rat renal medulla and is expressed exclusively on inner medullary collecting duct cells. The pattern of ET receptor mRNA expression described suggests different physiological actions for ET on the diverse cellular structures of the renal medulla.

Increased expression of cyclooxygenase-1 and -2 in the diabetic rat renal medulla

2003 ◽  
Vol 285 (6) ◽  
pp. F1068-F1077 ◽  
Author(s):  
Rania Nasrallah ◽  
Anne Landry ◽  
Sonia Singh ◽  
Monika Sklepowicz ◽  
Richard L. Hébert
Keyword(s):  
Collecting Duct ◽  
Renal Medulla ◽  
Immunohistochemical Analysis ◽  
Diabetic Rats ◽  
Diabetic Rat ◽  
Fourfold Increase ◽  
Protein Levels ◽  
Cyclooxygenase 1 ◽  
Streptozotocin Diabetic Rats ◽  
Medullary Collecting Duct

Alterations in renal prostaglandins (PGs) may contribute to some of the renal manifestations in diabetes leading to nephropathy. PG production is dependent on the activity of cyclooxygenases (COX-1 AND -2) and PG synthases. Our present study investigated levels of these enzymes in streptozotocin-diabetic rats at 2, 4, 6, and 8 wk of diabetes. Immunohistochemical analysis revealed an increase in COX signal in the inner and outer medulla of diabetic rats. This was confirmed by Western blotting, showing up to a fourfold increase in both COX isoforms at 4–6 wk of diabetes. Also, Western blot analysis revealed a sixfold increase in PGE2 synthase expression in the outer medullary region of 6-wk diabetic rats but no difference in the inner medulla. In cultured rat inner medullary collecting duct (IMCD), levels of COX were increased two- to threefold in cells exposed for 4 days to 37.5 mM glucose compared with control of 17.5 mM. While no change in PGE2 synthase levels was noted, PGE2 synthesis was increased. Furthermore, levels of EP1 and EP4 mRNA were increased, as well as a twofold increase in EP4 protein levels. Future studies will determine which COX isoform is contributing to the majority of PGE2 produced in the diabetic IMCD and the significance of these findings to disturbances in IMCD function and to the progression of diabetic nephropathy.

Importance of medullary events in ammonium excretion: studies in acute respiratory and acute metabolic acidosis

1983 ◽  
Vol 61 (1) ◽  
pp. 35-42 ◽  
Author(s):  
Andre Gougoux ◽  
Patrick Vinay ◽  
Guy Lemieux ◽  
Marc Goldstein ◽  
Bobby Stinebaugh ◽  
...  
Keyword(s):  
Metabolic Acidosis ◽  
Collecting Duct ◽  
Respiratory Acidosis ◽  
Renal Medulla ◽  
Hydrogen Ion ◽  
Ammonium Excretion ◽  
Urine Ph ◽  
Ion Secretion ◽  
Medullary Collecting Duct ◽  
Acute Metabolic Acidosis

The renal medulla can play an important role in acid excretion by modulating both hydrogen ion secretion in the medullary collecting duct and the medullary [Formula: see text]. The purpose of these experiments was to characterize the intrarenal events associated with ammonium excretion in acute acidosis. Cortical events were monitored in two ways: first, the rates of glutamine extraction and ammoniagenesis were assessed by measuring arteriovenous differences and the rate of renal blood flow; second, the biochemical response of the ammoniagenesis pathway was examined by measuring glutamate and 2-oxoglutarate, key renal cortical metabolites in this pathway. There were no significant differences noted in any of these cortical parameters between acute respiratory and metabolic acidosis. Despite a comparable twofold rise in ammonium excretion in both cases, the urine pH, [Formula: see text], and the urine minus blood [Formula: see text] difference (U-B [Formula: see text]) were lower during acute hypercapnia. In these experiments, the urine [Formula: see text] was 34 mmHg (1 mmHg = 133.322 Pa) lower than that of the blood during acute respiratory acidosis while the U-B [Formula: see text] was 5 ± 3 mmHg in acute metabolic acidosis. Thus there were significant differences in medullary events during these two conditions. Although the urine pH is critical in determining ammonium excretion in certain circumstances, these results suggest that regional variations in the medullary [Formula: see text] can modify this relationship.

Enhanced expression of epithelial sodium channels in the renal medulla of Dahl S rats

2011 ◽  
Vol 89 (3) ◽  
pp. 159-168 ◽  
Author(s):  
Md. Shahrier Amin ◽  
Erona Reza ◽  
Esraa El-Shahat ◽  
Hong-Wei Wang ◽  
Frédérique Tesson ◽  
...  
Keyword(s):  
Collecting Duct ◽  
Young Male ◽  
Renal Medulla ◽  
High Salt ◽  
High Salt Diet ◽  
Salt Diet ◽  
Enhanced Expression ◽  
Mrna And Protein Expression ◽  
Medullary Collecting Duct ◽  
Apical Localization

Inner medullary collecting duct (IMCD) cells from salt-sensitive (S) Dahl rats transport twice as much Na+ as cells from salt-resistant (R) rats, possibly related to dysregulation of the renal epithelial sodium channel (ENaC). The effect of a high-salt diet on ENaC expression in the inner medulla of S versus R rats has not yet been studied. Young, male S and R rats were placed on a regular-salt (0.3%) or high-salt (8%) diet for 2 or 4 weeks. mRNA and protein expression of ENaC subunits were studied by real-time PCR and immunoblotting. Intracellular distribution of the subunits in the IMCD was evaluated by immunohistochemistry. On regular salt, the abundance of the mRNA of β and γENaC was higher in the medulla of S rats than R rats. This was associated with a greater protein abundance of 90 kDa γENaC and higher immunoreactivity for both α and γ ENaC. High salt did not affect mRNA abundance in either strain and decreased apical staining of βENaC in IMCD of R rats. In contrast, high salt did not affect the higher apical localization of αENaC and increased the apical membrane staining for β and γENaC in the IMCD of S rats. Expression of ENaC subunits is enhanced in the medulla of S vs. R rats on regular salt, and further increased on high salt. The persistent high expression of αENaC and increase in apical localization of β and γENaC may contribute to greater retention of sodium in S rats on a high-salt diet.

scholarly journals Inhibition of biliverdin reductase increases ANG II-dependent superoxide levels in cultured renal tubular epithelial cells

2009 ◽  
Vol 297 (5) ◽  
pp. R1546-R1553 ◽  
Author(s):  
Shelby C. Young ◽  
Megan V. Storm ◽  
Joshua S. Speed ◽  
Silvia Kelsen ◽  
Chelsea V. Tiller ◽  
...  
Keyword(s):  
Collecting Duct ◽  
Renal Medulla ◽  
Superoxide Production ◽  
Heme Oxygenase 1 ◽  
Ang Ii ◽  
Small Interfering Rnas ◽  
Biliverdin Reductase ◽  
Medullary Collecting Duct ◽  
Renal Tubular ◽  
Antioxidant Effects

Induction of heme oxygenase-1 (HO-1) in the renal medulla increases carbon monoxide and bilirubin production and decreases ANG II-mediated superoxide production. The goal of this study was to determine the importance of increases in bilirubin to the antioxidant effects of HO-1 induction in cultured mouse thick ascending loop of Henle (TALH) and inner medullary collecting duct (IMCD3) cells. Bilirubin levels were decreased by using small interfering RNAs (siRNAs) targeted to biliverdin reductase (BVR), which is the cellular enzyme responsible for the conversion of biliverdin to bilirubin. Treatment of cultured TALH or IMCD-3 cells with BVR siRNA (50 or 100 nM) resulted in an 80% decrease in the level of BVR protein and decreased cellular bilirubin levels from 46 ± 5 to 23 ± 4 nM ( n = 4). We then determined the effects of inhibition of BVR on ANG II-mediated superoxide production. Superoxide production induced by ANG II (10−9 M) significantly increased in both TALH and IMCD-3 cells. Treatment of TALH cells with BVR siRNA resulted in a significant increase in ouabain-sensitive rubidium uptake from 95 ± 6 to 122 ± 5% control ( n = 4, P < 0.05). Lastly, inhibition of BVR with siRNA did not prevent the decrease in superoxide levels observed in cells pretreated with the HO-1 inducer, hemin. We conclude that decreased levels of cellular bilirubin increase ANG II-mediated superoxide production and sodium transport; however, increases in bilirubin are not necessary for HO-1 induction to attenuate ANG II-mediated superoxide production.

Role of H(+)-K(+)-ATPase in pHi regulation in inner medullary collecting duct cells in culture

1996 ◽  
Vol 270 (5) ◽  
pp. F852-F861 ◽  
Author(s):  
S. Ono ◽  
J. Guntupalli ◽  
T. D. DuBose
Keyword(s):  
Primary Culture ◽  
Collecting Duct ◽  
Wistar Rat ◽  
Bafilomycin A1 ◽  
Inner Medullary Collecting Duct ◽  
Murine Cell Line ◽  
Acid Load ◽  
Collecting Duct Cells ◽  
Medullary Collecting Duct ◽  
Hydrogen Ion Transport

Studies in inner medullary collecting duct (IMCD) cells in primary culture have proposed two mechanisms for Na(+)-independent hydrogen ion transport: an H(+)-adenosinetriphosphatase (H(+)-ATPase) and an H(+)-K(+)-ATPase. In the present study, we have employed two sources of IMCD cells, cells in primary culture derived from the terminal papilla of the Munich-Wistar rat (IMCDp) and an established murine cell line (mIMCD-3), to define the predominant mechanism(s) of Na(+)-independent intracellular pH (pHi) recovery in the IMCD. In confluent monolayers of IMCDp and mIMCD-3 cells, pHi was measured using the pH-sensitive dye 2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein (BCECF) following addition and withdrawal of NH4Cl. Removal of K+ completely abolished Na(+)-independent pHi recovery in both IMCDp (delta pHi/min = 0.039 +/- 0.006 to 0.005 +/- 0.003; P < 0.001) and in mIMCD-3 (delta pHi/min = 0.055 +/- 0.009 to -0.003 +/- 0.002; P < 0.001) cells, respectively. In mIMCD-3 cells, K(+)-dependent pHi recovery was abolished by either of two specific inhibitors of the H(+)-K(+)-ATPase, Sch-28080 (5 or 10 microM) or A-80915A (10 microM). In contrast, bafilomycin A1 (2.5 and 10 nM), an inhibitor of the H(+)-ATPase, failed to attenuate K(+)-dependent pHi recovery. Moreover, sequence verified mouse gastric and colonic alpha-H(+)-K(+)-ATPase probes hybridized to total RNA from mIMCD-3 cells. Based on these findings, we conclude that Na(+)-independent pHi recovery from an acid load in both IMCDp and mIMCD-3 cells in critically dependent on extracellular K(+)-That K(+)-dependent pHi recovery was inhibited by both Sch-28080 and A-80915A but not by bafilomycin A1 suggests that the predominant mechanism by which Na(+)-independent pHi recovery is accomplished in IMCD is through the H(+)-K(+)-ATPase. Expression of both gastric and colonic alpha-H(+)-K(+)-ATPase mRNA in mIMCD-3 cells suggests that one or both of these H(+)-K(+)-ATPases may be responsible for proton secretion in the IMCD.

Glucocorticoids inhibit transcription and expression of the UT-A urea transporter gene

2002 ◽  
Vol 282 (5) ◽  
pp. F853-F858 ◽  
Author(s):  
Tao Peng ◽  
Jeff M. Sands ◽  
Serena M. Bagnasco
Keyword(s):  
Northern Blot Analysis ◽  
Collecting Duct ◽  
Renal Medulla ◽  
Site Directed Mutagenesis ◽  
Protein Abundance ◽  
Urea Transporter ◽  
Dexamethasone Treatment ◽  
Gel Shift Assay ◽  
Medullary Collecting Duct ◽  
Specific Factors

Dexamethasone treatment increases urea excretion and decreases urea permeability and urea transporter UT-A1 protein abundance in the inner medullary collecting duct (IMCD) of adrenalectomized rats. We examined the effect of dexamethasone treatment for 3 days on the abundance of several UT-A mRNA transcripts in rat renal medulla. By Northern blot analysis, a significant decrease in mRNA expression was observed in the inner medulla of dexamethasone-treated rats compared with controls for UT-A1 (71%), UT-A3 (75%), and UT-A3b (75%), but not for UT-A2. We then tested the effect of 100 nM dexamethasone on the activity of promoter I in the UT-A gene, using LLC-PK1-GR101 cells that express the glucocorticoid receptor. Dexamethasone significantly decreased the activity of rat UT-A promoter I (72%) but did not affect UT-A promoter II. Deletion analysis and site-directed mutagenesis demonstrated that sequences between −423 and −244 are important for this inhibition and that a 10-bp sequence at −363, which binds a nuclear protein in a gel shift assay, is necessary for basal promoter activity. The specific factors involved in repression of UT-A promoter I activity by glucocorticoids remain to be determined.

Chronic hypokalemia enhances expression of the H(+)-K(+)-ATPase alpha 2-subunit gene in renal medulla

1996 ◽  
Vol 271 (2) ◽  
pp. F314-F321 ◽  
Author(s):  
K. Y. Ahn ◽  
K. Y. Park ◽  
K. K. Kim ◽  
B. C. Kone
Keyword(s):  
In Situ Hybridization ◽  
Collecting Duct ◽  
Rat Kidney ◽  
Renal Medulla ◽  
Northern Analysis ◽  
Thick Ascending Limb ◽  
Subunit Gene ◽  
Enhanced Expression ◽  
Medullary Collecting Duct

Recent molecular and physiological studies suggested that at least two H(+)-K(+)-adenosinetriphosphatase (H(+)-K(+)-ATPase) isozymes are expressed in the rat kidney and that these ion pumps respond to changes in dietary potassium balance. We used Northern analysis and in situ hybridization to analyze the expression of mRNA encoding the "colonic" isoform of the H(+)-K(+)-ATPase alpha-subunit (HK alpha 2) in normal and potassium-deprived (2 wk) rats. Control rats exhibited low levels of HK alpha 2 mRNA in the cortical and medullary thick ascending limb, distal convoluted tubule, connecting segment, and the entire collecting duct. The potassium-deprived rats expressed approximately fivefold higher levels of HK alpha 2 mRNA in the outer and inner medulla compared with controls, as well as hypertrophy and increased in situ hybridization signal in the intercalated cells of the inner stripe of the outer medullary collecting duct and the proximal inner medullary collecting duct. In contrast, renal cortical expression of HK alpha 2 mRNA was low and comparable in the two groups. Our results suggest that enhanced expression of the HK alpha 2 subunit gene in the renal medulla contributes to potassium conservation during chronic hypokalemia.

cGMP mediates effects of atrial peptides on medullary collecting duct cells

1987 ◽  
Vol 252 (3) ◽  
pp. F551-F559 ◽  
Author(s):  
M. L. Zeidel ◽  
P. Silva ◽  
B. M. Brenner ◽  
J. L. Seifter
Keyword(s):  
Collecting Duct ◽  
Renal Medulla ◽  
Fold Increase ◽  
Inhibitory Effect ◽  
Cyclic Monophosphate ◽  
Transport Effects ◽  
Collecting Duct Cells ◽  
Medullary Collecting Duct ◽  
Camp Levels

Atrial natriuretic peptides (ANP) stimulate renal Na+ excretion by poorly understood mechanisms, possibly involving direct inhibition of Na+ transport in the renal medulla. We have previously shown that human ANP 4-28 (hANP) inhibits Na+ entry-dependent O2 consumption (QO2) in rabbit inner medullary collecting duct (IMCD) cells. Because ANP actions in other tissues appear to be mediated by guanosine 3',5'-cyclic monophosphate (cGMP), the present studies examined the role of cyclic nucleotides in IMCD cell responses to ANP. 8-Bromo-cGMP (8-BrcGMP) diminished QO2 by 23.5 +/- 1.2% (SE) in IMCD cells but had no effect in cells derived from outer medullary collecting duct (OMCD); dibutyryl-adenosine 3',5'-cyclic monophosphate (cAMP) was without effect in IMCD cells. The inhibitory effect of BrcGMP was not additive with ANP, amiloride, or ouabain. Amphotericin, which enhances Na+ entry into cells, prevented the inhibitory effect of 8-BrcGMP. These results indicate that 8-BrcGMP, like ANP, inhibited Na+ entry in IMCD cells. hANP stimulated a 10-fold increase in cGMP in IMCD cells without altering IMCD cAMP levels or OMCD cGMP levels. Isobutyl methylxanthine, which inhibits phosphodiesterase activity, enhanced both cGMP accumulation and inhibition of QO2 by submaximal levels (10(-9) M) of ANP. Nitroprusside raised cGMP levels in both IMCD and OMCD cells but inhibited QO2 only in IMCD cells. We conclude that cGMP mediates the transport effects of ANP in IMCD cells. Our results indicate that cGMP may play an important role in the regulation of sodium transport in renal epithelia.

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