The Role of Pre-H2 Domains of α- and δ-Epithelial Na+Channels in Ion Permeation, Conductance, and Amiloride Sensitivity

To test the role of epithelial Na channels in the day-to-day regulation of renal Na excretion, rats were infused via osmotic minipumps with the Na channel blocker amiloride at rates that achieved drug concentrations of 2–5 μM in the lumen of the distal nephron. Daily Na excretion rates were unchanged, although amiloride-treated animals tended to excrete more Na in the afternoon and less in the late evening than controls. When the rats were given a low-Na diet, Na excretion rates were elevated in the amiloride-treated group within 4 h and remained higher than controls for at least 48 h. Adrenalectomized animals responded similarly to the low-Na diet. In contrast, rats infused with polythiazide at rates designed to inhibit NaCl transport in the distal tubule were able to conserve Na as well as did the controls. Injection of aldosterone (2 μg/100 g body wt) decreased Na excretion in control animals after a 1-h delay. This effect was largely abolished in amiloride-treated rats. On the basis of quantitative analysis of the results, we conclude that activation of amiloride-sensitive channels by mineralocorticoids accounts for 50–80% of the immediate natriuretic response of the kidney to a reduction in Na intake. Furthermore, the channels are necessary to achieve minimal rates of Na excretion during more chronic Na deprivation.

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Role of CFTR’s PDZ1-binding domain, NBF1 and Cl− conductance in inhibition of epithelial Na+ channels in Xenopus oocytes

Biochimica et Biophysica Acta (BBA) - Biomembranes ◽

10.1016/s0005-2736(01)00396-0 ◽

2001 ◽

Vol 1515 (1) ◽

pp. 64-71 ◽

Cited By ~ 16

Author(s):

A. Boucherot ◽

R. Schreiber ◽

K. Kunzelmann

Keyword(s):

Xenopus Oocytes ◽

Binding Domain ◽

Na Channels ◽

Epithelial Na Channels ◽

Cl Conductance

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Activation of epithelial Na+ channels by protein kinase A requires actin filaments

AJP Cell Physiology ◽

10.1152/ajpcell.1993.265.1.c224 ◽

1993 ◽

Vol 265 (1) ◽

pp. C224-C233 ◽

Cited By ~ 117

Author(s):

A. G. Prat ◽

A. M. Bertorello ◽

D. A. Ausiello ◽

H. F. Cantiello

Keyword(s):

Protein Kinase ◽

Actin Filaments ◽

Actin Polymerization ◽

Na Channel ◽

Na Channels ◽

Channel Activation ◽

Epithelial Na Channels ◽

Inside Out ◽

Kinase A

We have recently demonstrated a novel role for "short" actin filaments, a distinct species of polymerized actin different from either monomeric (G-actin) or long actin filaments (F-actin), in the activation of epithelial Na+ channels. In the present study, the role of actin in the activation of apical Na+ channels by the adenosine 3',5'-cyclic monophosphate-dependent protein kinase A (PKA) was investigated by patch-clamp techniques in A6 epithelial cells. In excised inside-out patches, addition of deoxyribonuclease I, which prevents actin polymerization, inhibited Na+ channel activation mediated by PKA. Disruption of endogenous actin filament organization with cytochalasin D for at least 1 h prevented the PKA-mediated activation of Na+ channels but not activation following the addition of actin to the cytosolic side of the patch. To assess the role of PKA on actin filament organization, actin was used as a substrate for the specific phosphorylation by the PKA. Actin was phosphorylated by PKA with an equilibrium stoichiometry of 2:1 mol PO4-actin monomer. Actin was phosphorylated in its monomeric form, but only poorly once polymerized. Furthermore, phosphorylated actin reduced the rate of actin polymerization. Thus actin allowed to polymerize for at least 1 h in the presence of PKA and ATP to obtain phosphorylated actin filaments induced Na+ channel activity in excised inside-out patches, in contrast to actin polymerized either in the absence of PKA or in the presence of PKA plus a PKA inhibitor (nonphosphorylated actin filaments). This was also confirmed by using purified phosphorylated G-actin incubated in a polymerizing buffer for at least 1 h at 37 degrees C. These data suggest that the form of actin required for Na+ channel activation (i.e., "short" actin filaments) may be favored by the phosphorylation of G-actin and may thus mediate or facilitate the activation of Na+ channels by PKA.

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Regulation of ion transport from within ion transit pathways

The Journal of General Physiology ◽

10.1085/jgp.201912455 ◽

2019 ◽

Vol 152 (1) ◽

Cited By ~ 4

Author(s):

Donald W. Hilgemann

Keyword(s):

Ion Channels ◽

Binding Sites ◽

Regulatory Mechanisms ◽

Ion Permeation ◽

Na Channels ◽

Insulin Regulation ◽

Underlying Principle ◽

Epithelial Na Channels ◽

Na Ions ◽

Secondary Activation

All cells must control the activities of their ion channels and transporters to maintain physiologically appropriate gradients of solutes and ions. The complexity of underlying regulatory mechanisms is staggering, as exemplified by insulin regulation of transporter trafficking. Simpler strategies occur in single-cell organisms, where subsets of transporters act as solute sensors to regulate expression of their active homologues. This Viewpoint highlights still simpler mechanisms by which Na transporters use their own transport sites as sensors for regulation. The underlying principle is inherent to Na/K pumps in which aspartate phosphorylation and dephosphorylation are controlled by occupation of transport sites for Na and K, respectively. By this same principle, Na binding to transport sites can control intrinsic inactivation reactions that are in turn modified by extrinsic signaling factors. Cardiac Na/Ca exchangers (NCX1s) and Na/K pumps are the best examples. Inactivation of NCX1 occurs when cytoplasmic Na sites are fully occupied and is regulated by lipid signaling. Inactivation of cardiac Na/K pumps occurs when cytoplasmic Na-binding sites are not fully occupied, and inactivation is in turn regulated by Ca signaling. Potentially, Na/H exchangers (NHEs) and epithelial Na channels (ENaCs) are regulated similarly. Extracellular protons and cytoplasmic Na ions oppose secondary activation of NHEs by cytoplasmic protons. ENaCs undergo inactivation as cytoplasmic Na rises, and small diffusible molecules of an unidentified nature are likely involved. Multiple other ion channels have recently been shown to be regulated by transiting ions, thereby underscoring that ion permeation and channel gating need not be independent processes.

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Dual Role of Prostaglandins (PGE 2 ) in Regulation of Channel Density and Open Probability of Epithelial Na + Channels in Frog Skin ( R. pipiens )

The Journal of Membrane Biology ◽

10.1007/s002329900159 ◽

1997 ◽

Vol 155 (1) ◽

pp. 75-87 ◽

Cited By ~ 24

Author(s):

W.J. Els ◽

S.I. Helman

Keyword(s):

Frog Skin ◽

Dual Role ◽

Na Channels ◽

Open Probability ◽

Channel Density ◽

Epithelial Na Channels

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Role of epithelial Na+channels in endothelial function

Journal of Cell Science ◽

10.1242/jcs.168831 ◽

2015 ◽

Vol 129 (2) ◽

pp. 290-297 ◽

Cited By ~ 28

Author(s):

Dongqing Guo ◽

Shenghui Liang ◽

Su Wang ◽

Chengchun Tang ◽

Bin Yao ◽

...

Keyword(s):

Endothelial Function ◽

Na Channels ◽

Epithelial Na Channels

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S-adenosyl-l-homocysteine hydrolase is necessary for aldosterone-induced activity of epithelial Na+ channels

AJP Cell Physiology ◽

10.1152/ajpcell.2001.281.3.c773 ◽

2001 ◽

Vol 281 (3) ◽

pp. C773-C785 ◽

Cited By ~ 13

Author(s):

James D. Stockand ◽

Shawn Zeltwanger ◽

Hui-Fang Bao ◽

Andrea Becchetti ◽

Roger T. Worrell ◽

...

Keyword(s):

Cell Line ◽

Transition Rate ◽

Critical Role ◽

Na Channels ◽

Open Probability ◽

Epithelial Na Channels ◽

Open States ◽

Kinetics Of ◽

Expression And Activity

The A6 cell line was used to study the role of S-adenosyl-l-homocysteine hydrolase (SAHHase) in the aldosterone-induced activation of the epithelial Na+channel (ENaC). Because aldosterone increases methylation of several different molecules, and because this methylation is associated with increased Na+ reabsorption, we tested the hypothesis that aldosterone increases the expression and activity of SAHHase protein. The rationale for this work is that general methylation may be promoted by activation of SAHHase, the only enzyme known to metabolize SAH, a potent end-product inhibitor of methylation. Although aldosterone increased SAHHase activity, steroid did not affect SAHHase expression. Antisense SAHHase oligonucleotide decreased SAHHase expression and activity. Moreover, this oligonucleotide, as well as a pharmacological inhibitor of SAHHase, decreased aldosterone-induced activity of ENaC via a decrease in ENaC open probability. The kinetics of ENaC in cells treated with antisense plus aldosterone were similar to those reported previously for the channel in the absence of steroid. This is the first report showing that active SAHHase, in part, increases ENaC open probability by reducing the transition rate from open states in response to aldosterone. Thus aldosterone-induced SAHHase activity plays a critical role in shifting ENaC from a gating mode with short open and closed times to one with longer open and closed times.

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