scholarly journals Na+ and K+ transport at basolateral membranes of epithelial cells. III. Voltage independence of basolateral membrane Na+ efflux.

1986 ◽  
Vol 87 (3) ◽  
pp. 503-509 ◽  
Author(s):  
T C Cox ◽  
S I Helman
Keyword(s):  
Frog Skin ◽  
Basolateral Membrane ◽  
Pump Current ◽  
Voltage Sensitivity ◽  
Na Transport ◽  
Previous Suggestion ◽  
Basolateral Membranes ◽  
Transmembrane Voltage ◽  
Na Efflux ◽  
K Transport

Na+ efflux across basolateral membranes of isolated epithelia of frog skin was tested for voltage sensitivity. The intracellular Na+ transport pool was loaded with 24Na from the apical solution and the rate of isotope appearance in the basolateral solution (JNa23) was measured at timed intervals of 30 s. Basolateral membrane voltage was depolarized by either 50 mM K+, 5 mM Ba++, or 80 mM NH+4. Whereas within 30 s ouabain caused inhibition of JNa23, depolarization of Vb by 30-60 mV caused no significant change of JNa23. Thus, both pump-mediated and leak Na+ effluxes were voltage independent. Although the pumps are electrogenic, pump-mediated Na+ efflux is voltage independent, perhaps because of a nonlinear relationship between pump current and transmembrane voltage. Voltage independence of the leak Na+ efflux confirms a previous suggestion (Cox and Helman, 1983. American Journal of Physiology. 245:F312-F321) that basolateral membrane Na+ leak fluxes are electroneutral.

scholarly journals Na+ and K+ transport at basolateral membranes of epithelial cells. I. Stoichiometry of the Na,K-ATPase.

1986 ◽  
Vol 87 (3) ◽  
pp. 467-483 ◽  
Author(s):  
T C Cox ◽  
S I Helman
Keyword(s):  
Short Circuit ◽  
Basolateral Membrane ◽  
Short Circuit Current ◽  
Ringer Solution ◽  
Na Transport ◽  
Basolateral Membranes ◽  
Dependent Component ◽  
Epithelial Sheets ◽  
Apical Membranes ◽  
K Transport

The stoichiometry of pump-mediated Na/K exchange was studied in isolated epithelial sheets of frog skin. 42K influx across basolateral membranes was measured with tissues in a steady state and incubated in either beakers or in chambers. The short-circuit current provided estimates of Na+ influx at the apical membranes of the cells. 42K influx of tissues bathed in Cl- or SO4-Ringer solution averaged approximately 8 microA/cm2. Ouabain inhibited 94% of the 42K influx. Furosemide was without effect on pre-ouabain-treated tissues but inhibited a ouabain-induced and Cl--dependent component of 42K influx. After taking into account the contribution of the Na+ load to the pump by way of basolateral membrane recycling of Na+, the stoichiometry was found to increase from approximately 2 to 6 as the pump-mediated Na+ transport rate increased from 10 to 70 microA/cm2. Extrapolation of the data to low rates of Na+ transport (less than 10 microA/cm2) indicated that the stoichiometry would be in the vicinity of 3:2. As pump-mediated K+ influx saturates with increasing rates of Na+ transport, Na+ efflux cannot be obligatorily coupled to K+ influx at all rates of transepithelial Na+ transport. These results are similar to those of Mullins and Brinley (1969. Journal of General Physiology. 53:504-740) in studies of the squid axon.

scholarly journals Na+ and K+ transport at basolateral membranes of epithelial cells. II. K+ efflux and stoichiometry of the Na,K-ATPase.

1986 ◽  
Vol 87 (3) ◽  
pp. 485-502 ◽  
Author(s):  
T C Cox ◽  
S I Helman
Keyword(s):  
Short Circuit ◽  
Short Circuit Current ◽  
Pump Current ◽  
Ringer Solution ◽  
K Channel ◽  
Na Transport ◽  
Transepithelial Na Transport ◽  
Dependent Component ◽  
Na Efflux ◽  
K Transport

Changes of 42K efflux (J23K) caused by ouabain and/or furosemide were measured in isolated epithelia of frog skin. From the kinetics of 42K influx (J32K) studied first over 8-9 h, K+ appeared to be distributed into readily and poorly exchangeable cellular pools of K+. The readily exchangeable pool of K+ was increased by amiloride and decreased by ouabain and/or K+-free extracellular Ringer solution. 42K efflux studies were carried out with tissues shortcircuited in chambers. Ouabain caused an immediate (less than 1 min) increase of the 42K efflux to approximately 174% of control in tissues incubated either in SO4-Ringer solution or in Cl-Ringer solution containing furosemide. Whereas furosemide had no effect on J23K in control tissues bathed in Cl-rich or Cl-free solutions, ouabain induced a furosemide-inhibitable and time-dependent increase of a neutral Cl-dependent component of the J23K. Electroconductive K+ transport occurred via a single-filing K+ channel with an n' of 2.9 K+ efflux before ouabain, normalized to post-ouabain (+/- furosemide) values of short-circuit current, averaged 8-10 microA/cm2. In agreement with the conclusions of the preceding article, the macroscopic stoichiometry of ouabain-inhibitable Na+/K+ exchange by the pump was variable, ranging between 1.7 and 7.2. With increasing rates of transepithelial Na+ transport, pump-mediated K+ influx saturated, whereas Na+ efflux continued to increase with increases of pump current. In the usual range of transepithelial Na+ transport, regulation of Na+ transport occurs via changes of pump-mediated Na+ efflux, with no obligatory coupling to pump-mediated K+ influx.

Dependence of intracellular Na+ concentration on apical and basolateral membrane Na+ influx in frog skin

1985 ◽  
Vol 249 (5) ◽  
pp. F662-F671
Author(s):  
J. S. Stoddard ◽  
S. I. Helman
Keyword(s):  
Epithelial Cells ◽  
Frog Skin ◽  
Apical Membrane ◽  
Basolateral Membrane ◽  
Na Transport ◽  
Maximal Inhibition ◽  
Basolateral Membranes ◽  
Isotopic Method ◽  
Rate Of Increase ◽  
Kinetics Of

An isotopic method was developed to measure the intracellular Na+ content of the transepithelial Na+ transport pool of frog skin. Isolated epithelia (no corium) were labeled with 24Na either asymmetrically, from apical (Aa) or basolateral (Ab) solutions, or symmetrically (Aab). Transport pool Na+ could be identified from the kinetics of washout of 24Na carried out in the presence of 1 mM ouabain, 100 microM amiloride, and 1 mM furosemide that served to trap cold Na+ and 24Na within the transport pool. In control epithelia, Aab averaged 64.1 neq/cm2 (13.9 mM), and maximal inhibition of apical membrane Na+ entry with 100 microM amiloride caused Aab to decrease to 24.3 neq/cm2 (5.3 mM). Ouabain caused Aab to increase markedly to 303 neq/cm2 in 30 min, whereas amiloride inhibition of apical membrane Na+ entry reduced markedly the rate of increase of Aab caused by ouabain (7.3 neq X cm-2 X min-1 in control and 1.7 neq X cm-2 X min-1 in the presence of amiloride). These data, in part, confirmed the existence of an important basolateral membrane permeability to Na+ that was measured in separate studies of the bidirectional 24Na fluxes at the basolateral membranes of the cells. Both sets of data were supportive of the idea that a significant Na+ recycling exists at the basolateral membranes of the cells that contributes to the Na+ load on the pump and Na+ recycling participates in the regulation of the Na+ concentration of the Na+ transport pool of these epithelial cells.

Effects of ouabain and furosemide on basolateral membrane Na efflux of frog skin

1983 ◽  
Vol 245 (3) ◽  
pp. F312-F321
Author(s):  
T. C. Cox ◽  
S. I. Helman
Keyword(s):  
Frog Skin ◽  
Basolateral Membrane ◽  
Ringer Solution ◽  
Na Transport ◽  
Consistent Effect ◽  
Unstirred Layers ◽  
Substantial Inhibition ◽  
Na Efflux ◽  
Kinetics Of ◽  
Single Exponential

The kinetics of Na tracer fluxes were reinvestigated in isolated epithelia of frog skin in which the unstirred layers of the corium were removed. The rate of appearance of 22Na in the basolateral solution (JNa13) conformed to a single exponential for tracer buildup with a mean t1/2 of 1.9 min. The Na transport pool labeled isotopically from the apical solution was about 40 neq/cm2 for epithelia bathed in either a chloride or sulfate Ringer solution. Basolateral-to-apical solution unidirectional Na flux (JNa31) was low, averaging 0.7 microA/cm2 and remained low for 40 min of ouabain treatment of the epithelia. In agreement with electrophysiological data, ouabain at 10(-4) M caused an acute inhibition (less than 1 min) of the Na efflux at the basolateral membrane (JNa23), falling to 40.6 and 26.7% of control for epithelia bathed in Cl and SO4 Ringer, respectively. Although furosemide exerted little or no consistent effect on the Na flux of control epithelia, this drug in ouabain-poisoned epithelia caused a substantial inhibition of a neutral, chloride-dependent, ouabain-insensitive Na efflux. It is suggested that ouabain "induces" a neutral mechanism of Na transport at the basolateral membrane of the cells that is chloride dependent and furosemide sensitive.

Origin of transport inhibition after omission of serosal sodium

1987 ◽  
Vol 252 (6) ◽  
pp. C623-C629 ◽  
Author(s):  
W. Nagel
Keyword(s):  
Intracellular Recording ◽  
Basolateral Membrane ◽  
Membrane Resistance ◽  
Na Transport ◽  
Transcellular Transport ◽  
Basolateral Membranes ◽  
Transport Inhibition ◽  
Intracellular Ca ◽  
K Permeability

The omission of sodium from the serosal incubation fluid in isolated frog skins inhibits transcellular Na transport. By the use of intracellular recording with microelectrodes, it has been demonstrated that this inhibition is associated with an increase of the basolateral membrane resistance, resulting in a depolarization of the short-circuited cells. This depolarization in turn accounts for the reduction of Na entry across the apical border. The resistance changes across the outer (apical) border are small in magnitude and unrelated to the inhibition of transcellular transport. The origin of the increase in basolateral membrane resistance, presumably due to decrease of K permeability, is unclear. These data do not support the hypothesis that intracellular Ca regulates the resistance of the apical and basolateral membranes.

Basolateral membrane chloride transport in isolated epithelia of frog skin

1985 ◽  
Vol 249 (3) ◽  
pp. C318-C329 ◽  
Author(s):  
J. S. Stoddard ◽  
E. Jakobsson ◽  
S. I. Helman
Keyword(s):  
Frog Skin ◽  
Specific Activity ◽  
Chloride Transport ◽  
Basolateral Membrane ◽  
Ringer Solution ◽  
Disulfonic Acid ◽  
Shunt Resistance ◽  
Basolateral Membranes ◽  
Cl Transport ◽  
Apical Membranes

Isotopic methodology was used to characterize Cl- transport in isolated epithelia of frog skin (northern Rana pipiens) bathed in Cl--rich Ringer solution and short-circuited. Cl- content of epithelia measured when loaded to 36Cl specific activity equilibrium averaged 139.6 neq/mg dry wt. The kinetics of 36Cl efflux was biexponential and consistent with binding or compartmentalization of approximately 30% of tissue Cl- within the intracellular pool. Because efflux of 36Cl to the apical solution was immeasurable, it was concluded that apical membranes were virtually impermeable to Cl- and that basolateral membranes were highly permeable to Cl- with a mean unidirectional Cl- efflux of 21.7 microA/cm2. Both furosemide (1 mM) and 4-acetamido-4'-isothiocyanostilbene-2,2'-disulfonic acid (5 X 10(-4) M) inhibited markedly the basolateral membrane chloride fluxes within seconds, as measured in chamber experiments. As inhibition of Cl- flux occurred in the absence of a change of the electrical parameters of apical and basolateral membranes, the mechanisms of Cl- transport appeared to be electroneutral and, for the most part at least, not coupled to the fluxes of Na+ and K+. Transepithelial Cl- fluxes averaged near 1 microA/cm2, proceeding via transport routes in parallel to the cells of the stratified epithelium. No correlation existed between the "shunt" resistance measured in the presence of 100 microM amiloride (greater than 1,000 omega X cm2) and the partial conductance to Cl-.

Analysis of K+ transport by rabbit CCD: conductive pathways and K(+)-K+ exchange by Na(+)-K+ pump

1992 ◽  
Vol 262 (1) ◽  
pp. F86-F97 ◽  
Author(s):  
T. Nonaka ◽  
D. H. Warden ◽  
J. B. Stokes
Keyword(s):  
Apical Membrane ◽  
Collecting Duct ◽  
Basolateral Membrane ◽  
Cortical Collecting Duct ◽  
Na Transport ◽  
Epithelial Na Channels ◽  
Electrophysiological Measurements ◽  
K Secretion ◽  
Cellular Pathways ◽  
K Transport

We studied the cellular pathways of K+ transport by the rabbit cortical collecting duct that was stimulated to absorb Na+ and to secrete K+. The vast majority of K+ secretion (into the lumen) was inhibited by benzamil, a blocker of epithelial Na+ channels. The residual K+ secretion was completely inhibited by ouabain. Thus all active K+ secretion was dependent on Na+ transport by the Na(+)-K+ pump. The passive pathways of K+ transport were further examined using tracer and electrophysiological measurements. K+ transfer across the apical membrane was predominantly or exclusively conductive; the apical K+ conductance was 31 mS/cm2. The basolateral membrane contained two pathways for K+ tracer translocation. The (barium-sensitive) conductive pathway accounted for a relatively small (12-20%) portion of the tracer permeation. A larger pathway appeared to be via K(+)-K+ exchange on the Na(+)-K+ pump. The magnitude of the Ba2(+)-sensitive (basolateral) K+ conductance predicted a substantially larger tracer flux than was actually measured. The best explanation for this difference is the presence of single-file diffusion through K+ channels on the apical and basolateral membranes. An analysis of the electrically silent K+ transport from lumen to bath suggests that the Na(+)-K+ pump can vary the ratio of its Na(+)-K+ and K(+)-K+ modes of operation. When the tubule is actively transporting Na+ and K+, the Na(+)-K+/K(+)-K+ turnover ratio is greater than 7. When Na+ transport is limited by inhibiting Na+ entry across the apical membrane, the ratio falls to less than 1. A major factor determining this ratio is probably the availability of Na+ to the cytoplasmic side of the pump.

Interactions of TPA and insulin on Na+ transport across frog skin

1989 ◽  
Vol 256 (3) ◽  
pp. C569-C578 ◽  
Author(s):  
M. M. Civan ◽  
K. Peterson-Yantorno ◽  
K. George ◽  
T. G. O'Brien
Keyword(s):  
Phorbol Ester ◽  
Frog Skin ◽  
Specific Binding ◽  
Short Circuit ◽  
Basolateral Membrane ◽  
Membrane Conductance ◽  
Na Transport ◽  
Early Stimulation ◽  
Na Permeability ◽  
Stimulation Of

The phorbol ester 12-O-tetradecanoylphorbol 13-acetate (TPA) activates protein kinase C (PKC) and produces an early stimulation of Na+ transport across frog skin. The ionic basis for this stimulation was studied with combined transepithelial and intracellular electrical measurements. In an initial series of experiments, TPA approximately doubled the amiloride-sensitive short-circuit current (ISC), apical Na+ permeability (PapNa), and apical membrane conductance without affecting the basolateral membrane conductance. The apical effects led to a marked depolarization of the short-circuited skin and a small increase in intracellular Na+ concentration. TPAs increase of PapNa was sufficient to explain the stimulation of basolateral Na+ transport when both the voltage and substrate dependence of the pump were taken into account. After the early stimulation, TPA later depressed ISC. Added at this point (congruent to 1-2 h after TPA administration), insulin had no effect on ISC, whereas a partial response to vasopressin was still observed. Measured either early or late after TPA addition, the phorbol ester reduced insulin binding by congruent to 40%. Insofar as 60% of the specific binding is retained, the abolishment of insulin's natriferic response is unlikely to result from the TPA-induced reduction in hormonal binding. The data provide further support for the concept that activation of PKC produces an early stimulation of Na+ transport by increasing apical Na+ permeability, and that part of insulin's natriferic effect may be mediated by PKC activation.

Vasopressin, theophylline, PGE2, and indomethacin on active Na transport in frog skin: studies with microelectrodes

1981 ◽  
Vol 241 (3) ◽  
pp. F279-F288 ◽  
Author(s):  
W. J. Els ◽  
S. I. Helman
Keyword(s):  
Frog Skin ◽  
Apical Membrane ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Membrane Voltage ◽  
Electrical Model ◽  
Na Transport ◽  
Basolateral Membranes ◽  
Transepithelial Voltage

Active transepithelial Na transport in frog skin is influenced by vasopressin, theophylline, indomethacin, and PGE2. During stimulation or inhibition of the short-circuit current, the transapical membrane voltage of short-circuited skins was recorded using an intracellular microelectrode. The microelectrode also permitted determination of the fractional resistance of the apical barrier of the cells (fRo) and the E'1 (transepithelial voltage at which the apical membrane voltage is zero). Analysis of the data according to an electrical model proposed previously indicated that changes of ISC were mediated primarily via changes of the slope resistance Rfo (Vo negative) of the apical barrier of the cells with little or no effect on the Thevenin emf or resistance of the basolateral membranes. These data are in accordance with previous observations that ADH had no effect on the ENa and are discussed in relation to the origin of the ENa at the basolateral membranes of the epithelial cells.

The Sigma-1 Receptor Ligands Chlorpromazine and Trifluoperazine Inhibit Na+ Transport in Frog Skin Epithelium

BIOPHYSICS ◽  
2020 ◽  
Vol 65 (5) ◽  
pp. 784-787
Author(s):  
A. V. Melnitskaya ◽  
Z. I. Krutetskaya ◽  
V. G. Antonov ◽  
N. I. Krutetskaya
Keyword(s):  
Frog Skin ◽  
Receptor Ligands ◽  
Na Transport ◽  
Sigma 1 Receptor ◽  
Skin Epithelium ◽  
Frog Skin Epithelium ◽  
Sigma 1
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