Modification of carboxyl of Na+ channel inhibits aldosterone action on Na+ transport

1983 ◽  
Vol 245 (6) ◽  
pp. F726-F734 ◽  
Author(s):  
J. Kipnowski ◽  
C. S. Park ◽  
D. D. Fanestil
Keyword(s):  
Amphotericin B ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Na Channel ◽  
Dependent Manner ◽  
Na Channels ◽  
Na Transport ◽  
Maximal Increase ◽  
Osmotic Water ◽  
Stimulation Of

We investigated the effect of the carboxyl-selective reagent N-ethoxycarbonyl-2-ethoxy-1,2-dihydroquinoline (EEDQ) on aldosterone stimulation of Na+ transport in the urinary bladder of the toad. Na+ transport, measured as the short-circuit current (SCC), was irreversibly inhibited by EEDQ in a dose- and time-dependent manner prior to addition of aldosterone. The greater the percentage inhibition by EEDQ (X), the smaller was the maximal increase of SCC after aldosterone (Y). This relationship gave the regression equation Y = 128.41 - 1.73X, r = -0.99 (n = 35). Evidence that the inhibition of SCC produced by EEDQ was limited to effects at the mucosal membrane was attested by the following: 1) EEDQ did not alter the stimulation by aldosterone of the osmotic water flow response to antidiuretic hormone; 2) whereas inhibition of protein synthesis by cycloheximide prevented this effect of aldosterone; 3) amphotericin B fully restored SCC previously inhibited by EEDQ to the level produced in tissues not inhibited by EEDQ; 4) comparison of the effects of amiloride vs. EEDQ pretreatment on the SCC response to aldosterone and amphotericin B revealed nearly identical characteristics; 5) in contrast, amphotericin B stimulation of SCC was limited when Na+ transport was limited by antimycin A (an inhibitor of energy production) or by ouabain. The findings fail to provide positive evidence for the hypothesis that aldosterone induces the synthesis of new Na+ channels but are consistent with hormonal activation of previously existing but nonfunctioning Na+ channels.

Metabolic requirements for anaerobic active Cl and Na transport in the bullfrog cornea

1979 ◽  
Vol 236 (5) ◽  
pp. C268-C276 ◽  
Author(s):  
P. S. Reinach ◽  
H. F. Schoen ◽  
O. A. Candia
Keyword(s):  
Energy Source ◽  
Amphotericin B ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Anaerobic Glycolysis ◽  
Anaerobic Conditions ◽  
Na Transport ◽  
Sufficient Energy ◽  
Aerobic And Anaerobic ◽  
Stimulation Of

In the bullfrog cornea, the relationships between the rates of aerobic and anaerobic glycolysis and active Cl and Na transport were studied. In NaCl Ringer (glucose-free), the short-circuit current (SCC) declined much more slowly under aerobic than under anaerobic conditions. The aerobic lactate effluxes in glucose-free and glucose-rich NaCl Ringer were 0.08 and 0.23 micromol/h.cm2, respectively. The transition to anoxia caused these values to increase significantly and was accompanied by depletion of endogenous glycogen in glucose-free Ringer. In Na2SO4 Ringer, amphotericin B (10(-5) M) stimulation of the aerobic SCC was not dependent on the presence of glucose but under anoxia, SCC stimulation required glucose. In Na2SO4 (glucose-rich) Ringer, amphotericin B stimulated the aerobic lactate efflux from 0.26 to 0.36 mumol/h.cm2 and anoxia increased it to 0.55 micromol/h.cm2. In NaCl Ringer, the addition of either 0.5 mM adenosine or 1 mM ATP with 26 mM glucose restored the anaerobic-inhibited SCC and lactate efflux of glucose-depleted corneas. The results show that the reactions of glycolysis are a sufficient energy source for supporting active Na and Cl transport.

Chronic regulation of transepithelial Na+ transport by the rate of apical Na+ entry

1996 ◽  
Vol 270 (2) ◽  
pp. C600-C607 ◽  
Author(s):  
M. D. Rokaw ◽  
E. Sarac ◽  
E. Lechman ◽  
M. West ◽  
J. Angeski ◽  
...  
Keyword(s):  
Short Circuit ◽  
Basolateral Membrane ◽  
Short Circuit Current ◽  
Maximal Activity ◽  
Na Channels ◽  
Na Transport ◽  
Bottom Structures ◽  
Stimulation Of

In several settings in vivo, prolonged inhibition of apical Na+ entry reduces and prolonged stimulation of apical entry enhances the ability of renal epithelial cells to reabsorb Na+, an important feature of the load-dependent regulation of renal tubular Na+ transport. To model this load dependency, apical Na+ entry was inhibited or stimulated for 18 h in A6 cells and vectorial transport was measured as short-circuit current (Isc) across monolayers on filter-bottom structures. Basal amiloride-sensitive Isc represents the activity of apical Na+ channels, whereas Isc after permeabilization of the apical membrane to cations with nystatin represents maximal activity of the basolateral Na(+)-K(+)-ATPase. Chronic inhibition of apical Na+ entry by 18-h apical exposure to amiloride or replacement of apical Na+ with tetramethylammonium (TMA+), followed by washing and restoration of normal apical medium, revealed a persistent decrease in Isc that remained despite exposure to nystatin. Both basal and nystatin-stimulated Isc recovered progressively after restoration of normal apical medium. In contrast, chronic stimulation of apical Na+ entry by short circuiting the epithelium increased Isc in the absence and presence of nystatin, indicating upregulation of both apical Na+ channels and basolateral Na(+)-K(+)-ATPase. Basolateral equilibrium [3H]ouabain binding was reduced to 67 +/- 5% in TMA+ vs. control cells, whereas values in 18-h short-circuited cells increased by 42 +/- 19%. The results demonstrate that load dependency of tubular Na+ transport can be modeled in vitro and indicate that the regulation of Na(+)-K(+)-ATPase observed in these studies occurs in part by changes in the density of functional transporter proteins within the basolateral membrane.

Regulation of electrogenic Na+ transport across leech skin

1995 ◽  
Vol 268 (3) ◽  
pp. R605-R613 ◽  
Author(s):  
W. M. Weber ◽  
U. Blank ◽  
W. Clauss
Keyword(s):  
Short Circuit ◽  
Short Circuit Current ◽  
Na Channel ◽  
Monovalent Cations ◽  
Serosal Side ◽  
Na Transport ◽  
Permeability Ratio ◽  
Cyclic Monophosphate ◽  
Apical Side ◽  
Stimulation Of

The dorsal integument of the medical leech Hirudo medicinalis exhibits a marked amiloride-sensitive Na+ absorption. With 20 mM Na+ in the apical solution, the transepithelial short-circuit current (Isc) was approximately 40% higher than with 115 mM Na+, whereas the transepithelial potential (VT) with 20 mM Na+ was -35.7 +/- 4.5 and -20.6 +/- 2.6 mV with 115 mM Na+. Amiloride (100 microM) inhibition at 20 mM apical Na+ was also significantly larger than with 115 mM Na+ in the solution. Benzamil (100 microM) showed additional inhibition after amiloride. Large transient overshooting currents occurred only when 115 mM Na+ was added after some minutes of Na(+)-free apical solution. Addition of adenosine 3',5'-cyclic monophosphate (cAMP) to the serosal side in the presence of 115 mM apical Na+ nearly doubled Isc. This cAMP effect was reduced to only 20% in the presence of 20 mM Na+. Guanosine 3',5'-cyclic monophosphate (cGMP) slightly increased Isc, whereas ATP showed biphasic potency. Removal of calcium from the apical side resulted in a large stimulation of amiloride-sensitive Isc only in the presence of 115 mM Na+. When currents were activated with cAMP, a deprivation of Ca2+ modestly reduced the amiloride-sensitive Isc. The Na+ channel of leech integument was found highly selective for Na+ over other monovalent cations. The permeability ratio for Na+ over K+ was approximately 30:1; the selectivity relationship for the investigated cations was Na+ > Li+ > NH4+ > K+ approximately Cs+ approximately Rb+.

Cholinergic modulation of apical Na+ channels in turtle colon: analysis of CDPC-induced fluctuations

1990 ◽  
Vol 259 (4) ◽  
pp. C668-C674 ◽  
Author(s):  
D. J. Wilkinson ◽  
D. C. Dawson
Keyword(s):  
Single Channel ◽  
Short Circuit ◽  
Basolateral Membrane ◽  
Short Circuit Current ◽  
Rate Coefficients ◽  
Corner Frequency ◽  
Na Channel ◽  
Fluctuation Analysis ◽  
Na Channels ◽  
Na Current

Current fluctuation analysis was used to investigate the properties of apical Na+ channels during muscarinic inhibition of active Na+ absorption. A reversible Na+ channel blocker, 6-chloro-3,5-diaminopyrazine-2-carboxamide (CDPC), was used to induce fluctuations in the short-circuit current (I(sc)). Power density spectra of the CDPC-induced fluctuations exhibited a clearly discernible Lorentzian component, characterized by a corner frequency that was linearly related to CDPC concentration between 20 and 100 microM. The on (k'on) and off (k(off)) rate coefficients for the CDPC blocking reaction were k'on = 11.1 +/- 0.8 rad.s-1.microM-1 and k(off) = 744 +/- 53 rad/s, and the microscopic inhibition constant was 67 microM (n = 11). CDPC blocking kinetics were not significantly different after inhibition of Isc by 5 microM serosal carbachol. Single-channel Na+ current (iNa) and the density of open and blocked Na+ channels (N(ob)) were estimated from the fluctuations induced by 40 microM CDPC. Under control conditions, iNa was 0.43 +/- 0.05 pA and N(ob) was 251 +/- 42 X 10(6)/cm2 (n = 10). After exposure to serosal carbachol (2-10 microM) for 60 min, Na+ current and N(ob) were reduced by approximately 50%, but iNa was not changed significantly. These results indicate that muscarinic inhibition of electrogenic Na+ absorption was associated with a reduction in the number of open Na+ channels in the apical membrane. They also suggest that this downregulation of transport involved a coordinated decrease in both apical and basolateral membrane conductances.

scholarly journals Amiloride-sensitive trypsinization of apical sodium channels. Analysis of hormonal regulation of sodium transport in toad bladder.

1983 ◽  
Vol 81 (6) ◽  
pp. 785-803 ◽  
Author(s):  
H Garty ◽  
I S Edelman
Keyword(s):  
Hormonal Regulation ◽  
Apical Membrane ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Toad Urinary Bladder ◽  
Apical Surface ◽  
Na Channels ◽  
Na Transport ◽  
Na Permeability ◽  
Trypsin Inhibition

Incubation of the mucosal surface of the toad urinary bladder with trypsin (1 mg/ml) irreversibly decreased the short-circuit current to 50% of the initial value. This decrease was accompanied by a proportionate decrease in apical Na permeability, estimated from the change in amiloride-sensitive resistance in depolarized preparations. In contrast, the paracellular resistance was unaffected by trypsinization. Amiloride, a specific blocker of the apical Na channels, prevented inactivation by trypsin. Inhibition of Na transport by substitution of mucosal Na, however, had no effect on the response to trypsin. Trypsinization of the apical membrane was also used to study regulation of Na transport by anti-diuretic hormone (ADH) and aldosterone. Prior exposure of the apical surface to trypsin did not reduce the response to ADH, which indicates that the ADH-induced Na channels were inaccessible to trypsin before addition of the hormone. On the other hand, stimulation of short-circuit current by aldosterone or pyruvate (added to substrate-depleted, aldosterone-repleted bladders) was substantially reduced by prior trypsinization of the apical surface. Thus, the increase in apical Na permeability elicited by aldosterone or substrate involves activation of Na channels that are continuously present in the apical membrane in nonconductive but trypsin-sensitive forms.

Network thermodynamic modeling of hormone regulation of active Na+ transport in cultured renal epithelium (A6)

1986 ◽  
Vol 250 (6) ◽  
pp. C978-C991 ◽  
Author(s):  
M. L. Fidelman ◽  
D. C. Mikulecky
Keyword(s):  
Steady State ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Open Circuit ◽  
Hormone Regulation ◽  
Na Channels ◽  
Na Transport ◽  
Renal Epithelium ◽  
Ion Concentrations ◽  
Tight Junction Permeability

A network thermodynamic model was developed to describe steady-state ion flows (Na+,K+, and Cl-) and related electrical events in a cultured renal epithelium (A6) derived from toad kidney. Three hypotheses for explaining the steady-state increases in short-circuit current (SCC) produced by aldosterone and/or insulin were examined using the model. Changing only the number of basolateral Na+-K+ pumps produced virtually no change in SCC and was ruled out. Changing only the number of apical Na+ channels could produce sufficient increases in SCC but presented problems in the pattern of changes produced in cell ion concentrations and therefore appeared unlikely. Changing both apical and basolateral parameters in a balanced, coordinated manner produced the maximal changes in SCC with the minimal changes in cell ion concentrations and appeared to be the "best" hypothesis. In addition, it was found necessary for tight junction permeability to increase as active Na+ transport increased under open-circuit conditions. Simulations, using these results, compared favorably with experimental data on the stimulatory effects of aldosterone and insulin, both separately and together, on active Na+ transport.

Effect of brefeldin A on ADH-induced transport responses of toad bladder

1994 ◽  
Vol 266 (4) ◽  
pp. C1069-C1076 ◽  
Author(s):  
K. Weng ◽  
J. B. Wade
Keyword(s):  
Water Permeability ◽  
Short Circuit ◽  
Brefeldin A ◽  
Membrane Traffic ◽  
Short Circuit Current ◽  
Toad Urinary Bladder ◽  
Membrane Biogenesis ◽  
Na Transport ◽  
Stimulation Of

We have used brefeldin A (BFA) to examine the role of membrane traffic in the short-circuit current (ISC) and water permeability responses of the toad urinary bladder. BFA treatment of 1 or 5 micrograms/ml had a complex effect on the response of the ISC to antidiuretic hormone (ADH) or forskolin stimulation. Although the responses to initial challenges by ADH were not impaired by BFA, subsequent ISC responses were progressively reduced. Similarly, while the response to an initial challenge by forskolin was modestly reduced by BFA, subsequent responses were markedly reduced. Inhibition of protein synthesis with cycloheximide (CHM) affected ISC responses similarly. Neither BFA nor CHM had an effect on water permeability responses. These observations show that although the membrane traffic responsible for the water permeability response is insensitive to inhibition by BFA or CHM, the stimulation of Na+ transport becomes increasingly sensitive to these inhibitors with successive challenges by ADH or forskolin. Although initial increases in Na+ transport utilize preexisting components, subsequent responses appear to require an intact system for membrane biogenesis.

Inhibitory effect of phorbol ester on sodium transport in frog urinary bladder

1990 ◽  
Vol 259 (3) ◽  
pp. F425-F431
Author(s):  
T. Satoh ◽  
H. Endou
Keyword(s):  
Urinary Bladder ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Inhibitory Effect ◽  
Inhibitory Influence ◽  
Dependent Manner ◽  
Camp Accumulation ◽  
Frog Urinary Bladder ◽  
Na Transport

To confirm the role of protein kinase C (PKC) on epithelial Na transport, we studied the effects of phorbol 12-myristate 13-acetate (PMA) and dioctanoylglycerol (DiC8), activators of PKC, on short-circuit current (Isc) in frog urinary bladder and further examined the influence of sphingosine, an inhibitor of PKC, on PMA- or DiC8-modulated Isc. PMA reduced basal Isc in a dose-dependent manner, and sphingosine (10 and 100 microM) partially restored PMA-reduced Isc. On the other hand, DiC8 (5 x 10(-5) M) also reduced basal Isc, and this action was completely prevented by 100 microM sphingosine. Both PMA (4 x 10(-5) M) and DiC8 inhibited vasopressin (50 mU/ml)- and forskolin (5 x 10(-5) M)-stimulated increases in Isc. PMA (4 x 10(-5) M) also inhibited 8-bromoadenosine 3',5'-cyclic monophosphate (8-BrcAMP)-stimulated increase in Isc. Furthermore, PMA (4 x 10(-5) M) and DiC8 (5 x 10(-5) M) inhibited vasopressin (50 mU/ml)-stimulated cAMP accumulation. DiC8 also inhibited forskolin-stimulated cAMP accumulation. These results indicate that PMA exerts inhibitory influence on Na transport mainly by its own potency of PKC activation. In addition, it is suggested that there is a cross talk in epithelial Na transport between PKC and cAMP-dependent pathway in frog urinary bladder.

Suppression of coupled Na-Cl absorption by aldosterone and dexamethasone in rat distal colon in vitro

1984 ◽  
Vol 246 (6) ◽  
pp. F785-F793 ◽  
Author(s):  
R. D. Perrone ◽  
S. L. Jenks
Keyword(s):  
Short Circuit ◽  
Short Circuit Current ◽  
Distal Colon ◽  
Rat Colon ◽  
Na Transport ◽  
Unidirectional Flux ◽  
Rat Distal Colon ◽  
Marked Suppression ◽  
Stimulation Of

Basal Na absorption in the rat colon is coupled to that of Cl in an electroneutral fashion. We previously determined that aldosterone or dexamethasone induces amiloride-sensitive mucosal-to-serosal Na flux approximately equal to the amiloride-sensitive short-circuit current in rat distal colon in vitro. However, the effect of these steroids on coupled Na-Cl absorption was not examined. For this purpose, we determined the unidirectional flux of Na and Cl in voltage-clamped distal colon segments from rats treated with aldosterone or dexamethasone. Amiloride was used as a probe for conductive Na absorption, and acetazolamide and Cl-free solutions were used as probes for coupled Na-Cl absorption. Our results indicate that the nature of colonic Na absorption is markedly changed after treatment with these steroids. In contrast to findings in the untreated rat, colonic Na absorption after treatment with aldosterone or dexamethasone was largely independent of the presence of Cl. Net Cl absorption and acetazolamide sensitivity were both greatly diminished. Thus, aldosterone and dexamethasone have multiple effects on Na transport in rat distal colon. In addition to the stimulation of conductive Na absorption by aldosterone, an effect well described in other epithelia, there is marked suppression of coupled Na-Cl absorption. Dexamethasone was less effective in suppressing Cl absorption but equally effective in stimulating conductive Na absorption. These steroid effects were greater in the terminal 1-2 cm of the rat colon.

Glucocorticosteroids increase sodium transport in middle ear epithelium

1997 ◽  
Vol 272 (1) ◽  
pp. C184-C190 ◽  
Author(s):  
P. Herman ◽  
C. T. Tan ◽  
T. van den Abbeele ◽  
B. Escoubet ◽  
G. Friedlander ◽  
...  
Keyword(s):  
Middle Ear ◽  
Short Circuit ◽  
Constitutive Expression ◽  
Short Circuit Current ◽  
Na Channel ◽  
Ribonuclease Protection Assay ◽  
Na Transport ◽  
Ribonuclease Protection ◽  
Dose Dependent ◽  
Epithelial Na Channel

The effect of glucocorticosteroids on ion transport was investigated on a middle ear cell line with the short-circuit current (Isc) technique. Dexamethasone (DXM) produced a dose- and time-dependent increase in Isc. Concentration of half-maximal stimulation was 2.68 x 10(-8) M. This effect was blunted by the glucocorticoid antagonist RU-38486 and was related to Na+ transport, as evidenced by the inhibition induced by 1) apical addition of the Na+ channel inhibitor benzamil (10(-6) M) or 2) substitution of Na+ with N-methylglucamine in the incubation medium. The increase in Na+ transport resulted from a primary modulation of apical Na+ entry, since 1) the Na(+)-K(+)-ATPase activity of cellular homogenates was not modified by corticosteroids and 2) the DXM-induced increase in the ouabain-sensitive uptake of 86Rb was blunted by benzamil. Ribonuclease protection assay revealed 1) a constitutive expression of the mRNA encoding the alpha-subunit of the epithelial Na+ channel and 2) that DXM increased the expression of this transcript. This increase was dose dependent and paralleled changes in transepithelial Na+ transport. This study suggests that a component of the beneficial effect of steroid therapy for the treatment of otitis media might be related to increased fluid clearance.

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