Faculty Opinions recommendation of The Na+-dependent chloride-bicarbonate exchanger SLC4A8 mediates an electroneutral Na+ reabsorption process in the renal cortical collecting ducts of mice.

Intercalated cells (ICs) from kidney collecting ducts contain proton-transporting ATPases (H+-ATPases) whose plasma membrane expression is regulated under a variety of conditions. It has been shown that net proton secretion occurs in the distal nephron from chronically K+-depleted rats and that upregulation of tubular H+- ATPase is involved in this process. However, regulation of this protein at the level of individual cells has not so far been examined. In the present study, H+-ATPase activity was determined in individually identified ICs from control and chronically K+-depleted rats (9–14 days on a low-K+ diet) by monitoring K+- and Na+-independent H+ extrusion rates after an acute acid load. Split-open rat cortical collecting tubules were loaded with the intracellular pH (pHi) indicator 2′,7′-bis(2-carboxyethyl)-5(6)-carboxyfluorescein, and pHiwas determined by using ratiometric fluorescence imaging. The rate of pHi recovery in ICs in response to an acute acid load, a measure of plasma membrane H+-ATPase activity, was increased after K+ depletion to almost three times that of controls. Furthermore, the lag time before the start of pHirecovery after the cells were maximally acidified fell from 93.5 ± 13.7 s in controls to 24.5 ± 2.1 s in K+-depleted rats. In all ICs tested, Na+- and K+-independent pHi recovery was abolished in the presence of bafilomycin (100 nM), an inhibitor of the H+-ATPase. Analysis of the cell-to-cell variability in the rate of pHi recovery reveals a change in the distribution of membrane-bound proton pumps in the IC population of cortical collecting duct from K+-depleted rats. Immunocytochemical analysis of collecting ducts from control and K+-depleted rats showed that K+-depletion increased the number of ICs with tight apical H+ATPase staining and decreased the number of cells with diffuse or basolateral H+-ATPase staining. Taken together, these data indicate that chronic K+ depletion induces a marked increase in plasma membrane H+ATPase activity in individual ICs.

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Downregulation of renal TonEBP in hypokalemic rats

AJP Renal Physiology ◽

10.1152/ajprenal.00502.2006 ◽

2007 ◽

Vol 293 (1) ◽

pp. F408-F415 ◽

Cited By ~ 22

Author(s):

Un Sil Jeon ◽

Ki-Hwan Han ◽

Soo-Hyun Park ◽

Sang Do Lee ◽

Mee Rie Sheen ◽

...

Keyword(s):

Cultured Cells ◽

Renal Medulla ◽

Distal Tubule ◽

Cell Type ◽

Outer Medulla ◽

Sodium Transporters ◽

Enhancer Binding Protein ◽

Collecting Ducts ◽

Cell Type Specific ◽

Underlying Mechanisms

Hypokalemia causes a significant decrease in the tonicity of the renal medullary interstitium in association with reduced expression of sodium transporters in the distal tubule. We asked whether hypokalemia caused downregulation of the tonicity-responsive enhancer binding protein (TonEBP) transcriptional activator in the renal medulla due to the reduced tonicity. We found that the abundance of TonEBP decreased significantly in the outer and inner medullas of hypokalemic rats. Underlying mechanisms appeared different in the two regions because the abundance of TonEBP mRNA was lower in the outer medulla but unchanged in the inner medulla. Immunohistochemical examination of TonEBP revealed cell type-specific differences. TonEBP expression decreased dramatically in the outer and inner medullary collecting ducts, thick ascending limbs, and interstitial cells. In the descending and ascending thin limbs, TonEBP abundance decreased modestly. In the outer medulla, TonEBP shifted to the cytoplasm in the descending thin limbs. As expected, transcription of aldose reductase, a target of TonEBP, was decreased since the abundance of mRNA and protein was reduced. Downregulation of TonEBP appeared to have also contributed to reduced expression of aquaporin-2 and UT-A urea transporters in the renal medulla. In cultured cells, expression and activity of TonEBP were not affected by reduced potassium concentrations in the medium. These data support the view that medullary tonicity regulates expression and nuclear distribution of TonEBP in the renal medulla in cell type-specific manners.

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Response of cortical collecting ducts from remnant kidneys to arginine vasopressin

Kidney International ◽

10.1038/ki.1992.175 ◽

1992 ◽

Vol 41 (5) ◽

pp. 1150-1154 ◽

Cited By ~ 8

Author(s):

Melvin Bonilla-Felix ◽

L. Lee Hamm ◽

John Herndon ◽

V. Matti Vehaskari

Keyword(s):

Arginine Vasopressin ◽

Collecting Ducts

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Automated method for the isolation of collecting ducts

AJP Renal Physiology ◽

10.1152/ajprenal.00273.2005 ◽

2006 ◽

Vol 291 (1) ◽

pp. F236-F245 ◽

Cited By ~ 18

Author(s):

R. Lance Miller ◽

Ping Zhang ◽

Tong Chen ◽

Andreas Rohrwasser ◽

Raoul D. Nelson

Keyword(s):

Flow Cytometry ◽

Large Particle ◽

Fluorescent Protein ◽

Collecting Duct ◽

Complex Object ◽

Cortical Collecting Duct ◽

Fluorescent Intensity ◽

Automated Method ◽

Collecting Ducts ◽

Time Required

The structural and functional heterogeneity of the collecting duct present a tremendous experimental challenge requiring manual microdissection, which is time-consuming, labor intensive, and not amenable to high throughput. To overcome these limitations, we developed a novel approach combining the use of transgenic mice expressing green fluorescent protein (GFP) in the collecting duct with large-particle-based flow cytometry to isolate pure populations of tubular fragments from the whole collecting duct (CD), or inner medullary (IMCD), outer medullary (OMCD), or connecting segment/cortical collecting duct (CNT/CCD). Kidneys were enzymatically dispersed into tubular fragments and sorted based on tubular length and GFP intensity using large-particle-based flow cytometry or a complex object parametric analyzer and sorter (COPAS). A LIVE/DEAD assay demonstrates that the tubules were >90% viable. Tubules were collected as a function of fluorescent intensity and analyzed by epifluorescence and phase microscopy for count accuracy, GFP positivity, average tubule length, and time required to collect 100 tubules. Similarly, mRNA and protein from sorted tubules were analyzed for expression of tubule segment-specific genes using quantitative real-time RT-PCR and immunoblotting. The purity and yield of sorted tubules were related to sort stringency. Four to six replicates of 100 collecting ducts (9.68 ± 0.44–14.5 ± 0.66 cm or 9.2 ± 0.7 mg tubular protein) were routinely obtained from a single mouse in under 1 h. In conclusion, large-particle-based flow cytometry is fast, reproducible, and generates sufficient amounts of highly pure and viable collecting ducts from single or replicate animals for gene expression and proteomic analysis.

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Na+-sensitive elevation in blood pressure is ENaC independent in diet-induced obesity and insulin resistance

AJP Renal Physiology ◽

10.1152/ajprenal.00265.2015 ◽

2016 ◽

Vol 310 (9) ◽

pp. F812-F820 ◽

Cited By ~ 16

Author(s):

Jonathan M. Nizar ◽

Wuxing Dong ◽

Robert B. McClellan ◽

Mariana Labarca ◽

Yuehan Zhou ◽

...

Keyword(s):

Insulin Resistance ◽

Blood Pressure ◽

Metabolic Syndrome ◽

Elevated Blood Pressure ◽

Elevated Blood ◽

High Fat ◽

Electrolyte Excretion ◽

Diet Induced Obesity ◽

Collecting Ducts ◽

Fat Feeding

The majority of patients with obesity, insulin resistance, and metabolic syndrome have hypertension, but the mechanisms of hypertension are poorly understood. In these patients, impaired sodium excretion is critical for the genesis of Na+-sensitive hypertension, and prior studies have proposed a role for the epithelial Na+ channel (ENaC) in this syndrome. We characterized high fat-fed mice as a model in which to study the contribution of ENaC-mediated Na+ reabsorption in obesity and insulin resistance. High fat-fed mice demonstrated impaired Na+ excretion and elevated blood pressure, which was significantly higher on a high-Na+ diet compared with low fat-fed control mice. However, high fat-fed mice had no increase in ENaC activity as measured by Na+ transport across microperfused cortical collecting ducts, electrolyte excretion, or blood pressure. In addition, we found no difference in endogenous urinary aldosterone excretion between groups on a normal or high-Na+ diet. High fat-fed mice provide a model of metabolic syndrome, recapitulating obesity, insulin resistance, impaired natriuresis, and a Na+-sensitive elevation in blood pressure. Surprisingly, in contrast to previous studies, our data demonstrate that high fat feeding of mice impairs natriuresis and produces elevated blood pressure that is independent of ENaC activity and likely caused by increased Na+ reabsorption upstream of the aldosterone-sensitive distal nephron.

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Intestinal inflammations increase efflux of innate lymphoid cells from the intestinal mucosa to the mesenteric lymph nodes through lymph‐collecting ducts

Microcirculation ◽

10.1111/micc.12694 ◽

2021 ◽

Author(s):

Kazuki Horiuchi ◽

Masaaki Higashiyama ◽

Chie Kurihara ◽

Kouji Matsumura ◽

Rina Tanemoto ◽

...

Keyword(s):

Lymph Nodes ◽

Intestinal Mucosa ◽

Innate Lymphoid Cells ◽

Lymphoid Cells ◽

Mesenteric Lymph Nodes ◽

Mesenteric Lymph ◽

Collecting Ducts

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Abstract 021: Nephron Specific Deletion of the Prorenin Receptor Modulates Blood Pressure and Urinary Na+ Excretion

Hypertension ◽

10.1161/hyp.66.suppl_1.021 ◽

2015 ◽

Vol 66 (suppl_1) ◽

Author(s):

Nirupama Ramkumar ◽

Deborah Stuart ◽

Elena V Mironova ◽

Vladislav Bugay ◽

Mykola Mamenko ◽

...

Keyword(s):

Blood Pressure ◽

Structural Defects ◽

Ang Ii ◽

Open Probability ◽

Protein Levels ◽

Prorenin Receptor ◽

Collecting Ducts ◽

Dependent Pathway ◽

Early Lethality ◽

Specific Deletion

The nephron prorenin receptor (PRR) may modulate blood pressure (BP) and Na+ balance. Since previous models of PRR knockout (KO) mice had early lethality and/or structural defects, we developed an inducible nephron-wide PRR KO using the Pax8/LC1 transgenes. Disruption of nephron PRR at 1 month of age caused no renal histological abnormalities. On a normal Na+ diet, wild-type (WT) and PRR KO mice had similar BP and Na+ excretion. However, PRR KO mice had elevated PRC (KO- 377 ± 77 vs WT- 127 ± 19 ng Ang-I/ml/hr) and a 50% decrease in renal ENaC-α protein. Protein levels of NHE3, NKCC2, NCC and ENaC-β/γ were similar between the two groups. Treatment with mouse prorenin (10 nM for 30 min) increased ENaC channel number by 2-fold, but not open probability, in isolated split-open cortical collecting ducts (CCD) from WT mice; this was prevented by Akt inhibition (A6730) but unaffected by blockade of AT-1 (losartan), ERK1/2 (U0126) or p38 MAPK (SB203580). Addition of prorenin (10 nM) did not change isolated CCD [Ca2+]i as assessed by Fura-2 loading (10 min exposure with readings every 3 sec). On a low Na+ diet, PRR KO mice had increased Na+ excretion (Day 2: KO - 66 ± 11 vs WT- 42 ± 6 μmol/day; Day 6: KO - 39 ± 4 vs ET- 23 ± 4 μmol/day) however, no differences in BP were observed. PRC was elevated in PRR KO mice on a low Na+ diet (KO- 384 ± 40 vs WT-174 ± 12 ng/ Ang-I/ml/hr). PRR KO mice had an attenuated hypertensive response to Angiotensin-II (Ang-II) infusion at 600 ng/Kg/min for 2 weeks (MAP: KO - 117 ± 4 vs WT - 133 ± 4 mm Hg over the course of Ang-II infusion). Urinary Na+ excretion was elevated in Ang-II treated PRR KO mice as compared to WT mice (KO-344 ± 14 vs WT-268 ±30 μmol/day). Taken together, these data indicate that nephron PRR, likely via direct prorenin/renin stimulation of an Akt-dependent pathway, stimulates CCD ENaC activity. Absence of nephron PRR promotes Na+ wasting and reduces the hypertensive response to Ang-II.

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