scholarly journals Na delivery and ENaC mediate flow regulation of collecting duct endothelin-1 production

2012 ◽  
Vol 302 (10) ◽  
pp. F1325-F1330 ◽  
Author(s):  
Meghana M. Pandit ◽  
Kevin A. Strait ◽  
Toshio Matsuda ◽  
Donald E. Kohan
Keyword(s):  
Plasma Membrane ◽  
Collecting Duct ◽  
Extracellular Fluid ◽  
Na Channel ◽  
Endothelin 1 ◽  
Flow Response ◽  
Mrna Content ◽  
Static Conditions ◽  
Biologic System

Collecting duct (CD) endothelin-1 (ET-1) is an important autocrine inhibitor of Na and water transport. CD ET-1 production is stimulated by extracellular fluid volume expansion and tubule fluid flow, suggesting a mechanism coupling CD Na delivery and ET-1 synthesis. A mouse cortical CD cell line, mpkCCDc14, was subjected to static or flow conditions for 2 h at 2 dyn/cm2, followed by determination of ET-1 mRNA content. Flow with 300 mosmol/l NaCl increased ET-1 mRNA to 65% above that observed under static conditions. Increasing perfusate osmolarity to 450 mosmol/l with NaCl or Na acetate increased ET-1 mRNA to ∼184% compared with no flow, which was not observed when osmolarity was increased using mannitol or urea. Reducing Na concentration to 150 mosmol/l while maintaining total osmolarity at 300 mosmol/l with urea or mannitol decreased the flow response. Inhibition of epithelial Na channel (ENaC) with amiloride or benzamil abolished the flow response, suggesting involvement of ENaC in flow-regulated ET-1 synthesis. Aldosterone almost doubled the flow response. Since Ca2+ enhances CD ET-1 production, the involvement of plasma membrane and mitochondrial Na/Ca2+ exchangers (NCX) was assessed. SEA0400 and KB-R7943, plasma membrane NCX inhibitors, did not affect the flow response. However, CGP37157, a mitochondrial NCX inhibitor, abolished the response. In summary, the current study indicates that increased Na delivery, leading to ENaC-mediated Na entry and mitochondrial NCX activity, is involved in flow-stimulated CD ET-1 synthesis. This constitutes the first report of either ENaC or mitochondrial NCX regulation of an autocrine factor in any biologic system.

scholarly journals Flow regulation of endothelin-1 production in the inner medullary collecting duct

2015 ◽  
Vol 308 (6) ◽  
pp. F541-F552 ◽  
Author(s):  
Meghana M. Pandit ◽  
Edward W. Inscho ◽  
Shali Zhang ◽  
Tsugio Seki ◽  
Rajeev Rohatgi ◽  
...  
Keyword(s):  
Volume Expansion ◽  
Transient Receptor Potential ◽  
Collecting Duct ◽  
Extracellular Fluid ◽  
Receptor Potential ◽  
P2 Receptors ◽  
Flow Response ◽  
Mrna Content ◽  
Intracellular Ca ◽  
Transient Receptor

Collecting duct-derived endothelin (ET)-1 is an autocrine inhibitor of Na+ and water reabsorption; its deficiency causes hypertension and water retention. Extracellular fluid volume expansion increases collecting duct ET-1, thereby promoting natriuresis and diuresis; however, how this coupling between volume expansion and collecting duct ET-1 occurs is incompletely understood. One possibility is that volume expansion increases tubular fluid flow. To investigate this, cultured IMCD3 cells were subjected to static or flow conditions. Exposure to a shear stress of 2 dyn/cm2 for 2 h increased ET-1 mRNA content by ∼2.3-fold. Absence of perfusate Ca2+, chelation of intracellular Ca2+, or inhibition of Ca2+ signaling (calmodulin, Ca2+/calmodulin-dependent kinase, calcineurin, PKC, or phospholipase C) prevented the flow response. Evaluation of possible flow-activated Ca2+ entry pathways revealed no role for transient receptor potential (TRP)C3, TRPC6, and TRPV4; however, cells with TRPP2 (polycystin-2) knockdown had no ET-1 flow response. Flow increased intracellular Ca2+ was blunted in TRPP2 knockdown cells. Nonspecific blockade of P2 receptors, as well as specific inhibition of P2X7 and P2Y2 receptors, prevented the ET-1 flow response. The ET-1 flow response was not affected by inhibition of either epithelial Na+ channels or the mitochondrial Na+/Ca2+ exchanger. Taken together, these findings provide evidence that in IMCD3 cells, flow, via polycystin-2 and P2 receptors, engages Ca2+-dependent signaling pathways that stimulate ET-1 synthesis.

Immunocytochemical localization of Na+ channels in rat kidney medulla

1989 ◽  
Vol 256 (2) ◽  
pp. F366-F369 ◽  
Author(s):  
D. Brown ◽  
E. J. Sorscher ◽  
D. A. Ausiello ◽  
D. J. Benos
Keyword(s):  
Plasma Membrane ◽  
Plasma Membranes ◽  
Collecting Duct ◽  
Rat Kidney ◽  
Apical Plasma Membrane ◽  
Na Channel ◽  
Thick Ascending Limb ◽  
Na Channels ◽  
Kidney Medulla ◽  
Principal Cells

Amiloride-sensitive Na+ channels were localized in semithin frozen sections of rat renal medullary collecting ducts, using polyclonal antibodies directed against purified bovine kidney Na+ channel protein. The apical plasma membrane of collecting duct principal cells was heavily stained by indirect immunofluorescence, whereas intercalated cells were negative. Basolateral plasma membranes of both cell types were unstained, as were subapical vesicles in the cytoplasm of these cells. In the thick ascending limb of Henle, some scattered granular fluorescence was seen in the cytoplasm and close to the apical pole of epithelial cells, suggesting the presence of antigenic sites associated with some membrane domains in these cells. No staining was detected in thin limbs of Henle, or in proximal tubules in the outer medulla. These results show that amiloride-sensitive sodium channels are located predominantly on the apical plasma membrane of medullary collecting duct principal cells, the cells that are involved in Na+ homeostasis in this region of the kidney.

scholarly journals Endothelin-1 inhibits sodium reabsorption by ETA and ETB receptors in the mouse cortical collecting duct

2013 ◽  
Vol 305 (4) ◽  
pp. F568-F573 ◽  
Author(s):  
I. Jeanette Lynch ◽  
Amanda K. Welch ◽  
Donald E. Kohan ◽  
Brian D. Cain ◽  
Charles S. Wingo
Keyword(s):  
Hormonal Regulation ◽  
Collecting Duct ◽  
Early Response ◽  
Sodium Reabsorption ◽  
Na Channel ◽  
Cortical Collecting Duct ◽  
Water Excretion ◽  
Endothelin 1 ◽  
Arterial Blood

The collecting duct (CD) is a major renal site for the hormonal regulation of Na homeostasis and is critical for systemic arterial blood pressure control. Our previous studies demonstrated that the endothelin-1 gene (edn1) is an early response gene to the action of aldosterone. Because aldosterone and endothelin-1 (ET-1) have opposing actions on Na reabsorption (JNa) in the kidney, we postulated that stimulation of ET-1 by aldosterone acts as a negative feedback mechanism, acting locally within the CD. Aldosterone is known to increase JNa in the CD, in part, by stimulating the epithelial Na channel (ENaC). In contrast, ET-1 increases Na and water excretion through its binding to receptors in the CD. To date, direct measurement of the quantitative effect of ET-1 on transepithelial JNa in the isolated in vitro microperfused mouse CD has not been determined. We observed that the CD exhibits substantial JNa in male and female mice that is regulated, in part, by a benzamil-sensitive pathway, presumably ENaC. ENaC-mediated JNa is greater in the cortical CD (CCD) than in the outer medullary CD (OMCD); however, benzamil-insensitive JNa is present in the CCD and not in the OMCD. In the presence of ET-1, ENaC-mediated JNa is significantly inhibited. Blockade of either ETA or ETB receptor restored JNa to control rates; however, only ETA receptor blockade restored a benzamil-sensitive component of JNa. We conclude 1) Na reabsorption is mediated by ENaC in the CCD and OMCD and also by an ENaC-independent mechanism in the CCD; and 2) ET-1 inhibits JNa in the CCD through both ETA and ETB receptor-mediated pathways.

scholarly journals Regulation of the epithelial Na+ channel by endothelin-1 in rat collecting duct

2008 ◽  
Vol 295 (4) ◽  
pp. F1063-F1070 ◽  
Author(s):  
Vladislav Bugaj ◽  
Oleh Pochynyuk ◽  
Elena Mironova ◽  
Alain Vandewalle ◽  
Jorge L. Medina ◽  
...  
Keyword(s):  
Tyrosine Kinases ◽  
Collecting Duct ◽  
Basolateral Membrane ◽  
Na Channel ◽  
Open Probability ◽  
Endothelin 1 ◽  
Principal Cells ◽  
Patch Clamp Electrophysiology ◽  
Src Family Tyrosine Kinases

We used patch-clamp electrophysiology to investigate regulation of the epithelial Na+ channel (ENaC) by endothelin-1 (ET-1) in isolated, split-open rat collecting ducts. ET-1 significantly decreases ENaC open probability by about threefold within 5 min. ET-1 decreases ENaC activity through basolateral membrane ETB but not ETA receptors. In rat collecting duct, we find no role for phospholipase C or protein kinase C in the rapid response of ENaC to ET-1. ET-1, although, does activate src family tyrosine kinases and their downstream MAPK1/2 effector cascade in renal principal cells. Both src kinases and MAPK1/2 signaling are necessary for ET-1-dependent decreases in ENaC open probability in the split-open collecting duct. We conclude that ET-1 in a physiologically relevant manner rapidly suppresses ENaC activity in native, mammalian principal cells. These findings may provide a potential mechanism for the natriuresis observed in vivo in response to ET-1, as well as a potential cause for the salt-sensitive hypertension found in animals with impaired endothelin signaling.

Increased expression and apical targeting of renal ENaC subunits in puromycin aminonucleoside-induced nephrotic syndrome in rats

2004 ◽  
Vol 286 (5) ◽  
pp. F922-F935 ◽  
Author(s):  
Soo Wan Kim ◽  
Weidong Wang ◽  
Jakob Nielsen ◽  
Jeppe Praetorius ◽  
Tae-Hwan Kwon ◽  
...  
Keyword(s):  
Nephrotic Syndrome ◽  
Plasma Membrane ◽  
Sodium Excretion ◽  
Collecting Duct ◽  
Apical Plasma Membrane ◽  
Control Group ◽  
Na Channel ◽  
Sodium Retention ◽  
Puromycin Aminonucleoside ◽  
Connecting Tubule

Nephrotic syndrome is often accompanied by sodium retention and generalized edema. However, the molecular basis for the decreased renal sodium excretion remains undefined. We hypothesized that epithelial Na channel (ENaC) subunit dysregulation may be responsible for the increased sodium retention. An experimental group of rats was treated with puromycin aminonucleoside (PAN; 180 mg/kg iv), whereas the control group received only vehicle. After 7 days, PAN treatment induced significant proteinuria, hypoalbuminemia, decreased urinary sodium excretion, and extensive ascites. The protein abundance of α-ENaC and β-ENaC was increased in the inner stripe of the outer medulla (ISOM) and in the inner medulla (IM) but was not altered in the cortex. γ-ENaC abundance was increased in the cortex, ISOM, and IM. Immunoperoxidase brightfield- and laser-scanning confocal fluorescence microscopy demonstrated increased targeting of α-ENaC, β-ENaC, and γ-ENaC subunits to the apical plasma membrane in the distal convoluted tubule (DCT2), connecting tubule, and cortical and medullary collecting duct segments. Immunoelectron microscopy further revealed an increased labeling of α-ENaC in the apical plasma membrane of cortical collecting duct principal cells of PAN-treated rats, indicating enhanced apical targeting of α-ENaC subunits. In contrast, the protein abundances of Na+/H+ exchanger type 3 (NHE3), Na+-K+-2Cl- cotransporter (BSC-1), and thiazide-sensitive Na+-Cl- cotransporter (TSC) were decreased. Moreover, the abundance of the α1-subunit of the Na-K-ATPase was decreased in the cortex and ISOM, but it remained unchanged in the IM. In conclusion, the increased or sustained expression of ENaC subunits combined with increased apical targeting in the DCT2, connecting tubule, and collecting duct are likely to play a role in the sodium retention associated with PAN-induced nephrotic syndrome. The decreased abundance of NHE3, BSC-1, TSC, and Na-K-ATPase may play a compensatory role to promote sodium excretion.

scholarly journals Flow regulation of collecting duct endothelin-1 production

2011 ◽  
Vol 300 (3) ◽  
pp. F650-F656 ◽  
Author(s):  
Brianna Lyon-Roberts ◽  
Kevin A. Strait ◽  
Evan van Peursem ◽  
Wararat Kittikulsuth ◽  
Jennifer S. Pollock ◽  
...  
Keyword(s):  
Collecting Duct ◽  
Primary Cultures ◽  
Extracellular Fluid ◽  
Maximal Effect ◽  
Mrna Levels ◽  
Mrna Accumulation ◽  
Salt Loading ◽  
Endothelin 1 ◽  
Definitive Evidence ◽  
Intracellular Ca

Collecting duct (CD) endothelin-1 (ET-1) is an important autocrine inhibitor of CD Na+ reabsorption. Salt loading is thought to increase CD ET-1 production; however, definitive evidence of this, as well as understanding of the mechanisms transducing this effect, is lacking. Tubule fluid flow increases in response to Na+ loading; hence, we studied flow modulation of CD ET-1 production. Three days of a high-salt diet increased mouse and rat inner medullary CD (IMCD) ET-1 mRNA expression. Acute furosemide infusion increased urinary ET-1 excretion in anesthetized rats. Primary cultures of mouse or rat IMCD detached in response to flow using a closed perfusion chamber, consequently a CD cell line (mpkCCDcl4) was examined. Flow increased ET-1 mRNA at shear stress rates exceeding 1 dyne/cm2, with the maximal effect seen between 2 and 10 dyne/cm2. Induction of ET-1 mRNA was first evident after 1 h, and most apparent after 2 h, of flow. Inhibition of calmodulin or dihydropyridine-sensitive Ca2+ channels did not alter the flow response; however, chelation of intracellular Ca2+ or removal of extracellular Ca2+ largely prevented flow-stimulated ET-1 mRNA accumulation. Downregulation of protein kinase C (PKC) using phorbol 12-myristate 13-acetate, or PKC inhibition with calphostin C, markedly reduced flow-stimulated ET-1 mRNA levels. Flow-stimulated ET-1 mRNA accumulation was abolished by inhibition of phospholipase C (PLC). Taken together, these data indicate that flow increases CD ET-1 production and this is dependent on extracellular and intracellular Ca2+, PKC, and PLC. These studies suggest a novel pathway for coupling alterations in extracellular fluid volume to CD ET-1 production and ultimately control of CD Na+ reabsorption.

scholarly journals Role for reactive oxygen species in flow-stimulated inner medullary collecting duct endothelin-1 production

2017 ◽  
Vol 313 (2) ◽  
pp. F514-F521 ◽  
Author(s):  
Will Wheatley ◽  
Donald E. Kohan
Keyword(s):  
Reactive Oxygen Species ◽  
Collecting Duct ◽  
Reactive Oxygen ◽  
Ros Production ◽  
Oxygen Species ◽  
First Report ◽  
Endothelin 1 ◽  
Inner Medullary Collecting Duct ◽  
Flow Response ◽  
Medullary Collecting Duct

Inner medullary collecting duct (IMCD)-derived endothelin-1 (ET-1) is stimulated by volume expansion, in part through augmented luminal flow, whereupon it can elicit natriuresis and diuresis. Since flow can alter nitric oxide (NO) and reactive oxygen species (ROS), both of which can affect collecting duct salt transport, we asked whether NO and/or ROS mediate flow-stimulated IMCD ET-1. Mouse IMCD3 cells were exposed to flow, and ET-1/GAPDH mRNA was assessed. A shear stress of 10 dyn/cm2 for 1 h increased ET-1 mRNA by fourfold compared with no flow (ET-1 flow response). Global NO synthase (NOS) inhibition [ NG-nitro-l-arginine methyl ester (l-NAME)] reduced the ET-1 flow response; however, pharmacological inhibition of NOS1 or NOS2, inhibition of NOS3 siRNA, inhibition of arginase inhibition, removal of media l-Arg, or inhibition of NO-dependent signaling pathways (PKG, guanylyl cyclase, or NF-κB) did not affect the ET-1 flow response. Tempol reduced the ET-1 flow response; no further inhibition occurred with l-NAME. Superoxide dismutase, but not catalase, reduced the ET-1 flow response. Inhibition of NAPDH oxidase (NOX) (apocynin), pharmacological inhibition of NOX1/4, or NOX4 siRNA reduced the ET-1 flow response. Finally, flow increased IMCD3 ROS production and this was inhibited by apocynin, NOX1/4 inhibition, and, to a small extent, by l-NAME. Taken together, these data suggest that NOX4-derived ROS in general, and possibly superoxide in particular, are involved in flow-stimulated IMCD ET-1 production. To our knowledge, this is the first report of flow-stimulated ROS production by the CD, as well as the first report of such flow-stimulated CD ROS exerting a biological effect.

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