Biomedical subjects

W Van Driessche

Publications and source records attributed to W Van Driessche.

At least 19 recordsLinked to original sources

PubMed1995

Effect of dexamethasone on sodium channel block and densities in A6 cells.

The association (ON) and dissociation (OFF) rates of either positively charged amiloride or its uncharged analogue, CDPC (6-chloro-3, 5-diaminopyrazine-2-carboxamide), with the apical Na+ channel protein of renal A6 cells were analysed during exposure to the synthetic glucocorticoid, dexamethasone, using noise analysis. These rates were further used to reach specific conclusions about single-channel current, channel density and open probability of the channel in the absence of the blocker. Short-term exposure (3 h) to 10(-7) mol/l dexamethasone at the basolateral side increased the short-circuit current, Isc by 85%, without a change in the ON and OFF rates of the interaction between amiloride and the Na+ channel. A longer incubation (24 h) with dexamethasone tripled the current with a notable increase in the ON rate of the interaction between amiloride and the and channel. The OFF rate remained constant. The effects of dexamethasone on the rate constants of the reaction of amiloride with the channel did not match with the expected changes in membrane potential. On the other hand, ON and OFF rates of the interaction between neutral CDPC and the channel were not influenced by a 24-h incubation with dexamethasone. Further calculations disclosed that the gain in macroscopic current after a 24-h incubation with dexamethasone might be explained by an increase in Na+ channel density, and, to a lesser extent, by a rise in single-channel current. This all occurred without a change in the fraction of time spent by the channel in the conducting state in the absence of the blocker.

Amiloride↗

PubMed1994

Effect of insulin on area and Na+ channel density of apical membrane of cultured toad kidney cells.

1. The stimulation of transepithelial Na+ transport caused by insulin in A6 cultured toad kidney cells was investigated by determination of membrane capacitance (Cm), short circuit current (Isc) and current fluctuation analysis. Values of Cm are proportional to membrane area while blocker-induced current fluctuation analysis provides an estimate of the number of active amiloride-sensitive Na+ channels in the apical membrane. 2. Insulin simultaneously increased Cm, Isc and Gt (transepithelial conductance) in epithelia incubated with Na(+)-containing solutions on both sides. 3. Analysis of 6-chloro-3,5-diaminopyrazine-2-carboxamide (CDPC)-induced noise showed that insulin increased the number of active Na+ channels in the apical membrane, without altering the single channel current. 4. When nystatin was used to permeabilize the apical membrane the impedance data revealed the presence of a second time constant. Analysis of these data indicated that the basolateral membrane capacitance (Cb) is much larger than the apical membrane capacitance (Ca). Insulin administered to nystatin-treated epithelia increased the values for both capacitances. 5. We suggest that the stimulation of transepithelial Na+ transport caused by insulin may be associated with the exocytotic delivery of transporters to the apical membrane.

Amiloride↗

PubMed1994

Cell swelling activates a poorly selective monovalent cation channel in the apical membrane of toad urinary bladder.

We measured the effects of conditions that increase cell volume on ion movements through the Ca(2+)-blockable poorly selective monovalent cation channel of the apical membrane of the toad urinary bladder. Three conditions were studied: dilution of the basolateral solution, basolateral perfusion with solutions prepared with solutes of low reflection coefficient and dilution of the apical solution in bladders treated with oxytocin. All three procedures markedly increased K+ movements and elevated the plateau of the Lorentzian component of the power spectrum by enhancing ion currents through the Ca(2+)-blockable pathway. Simultaneously there was a large increase in Ca(2+)-sensitive conductance. The magnitude of this increased conductance strongly suggests that the stimulation of ion flow is due to increased ion permeability and not solely to increases in driving force across the apical membrane. We were not able to detect an increase in the movements of alkali-earth ions induced by the conditions that increase cell volume. We speculate that activation of the Ca(2+)-blockable channel may play an important role in the regulation of cell volume and/or in K+ homeostasis.

Animals↗

PubMed1994

Responses of endothelial cells to hypotonic solutions: lack of regulatory volume decrease.

Hypotonic stress (HTS) activates a Cl- current and releases intracellular Ca2+ in vascular endothelial cells, but there is no co-activation of K(+)-channels. The concomitant increase in cell volume, as assessed from the changes in endothelial cell thickness, is not followed by a regulatory volume decrease (RVD). This lack compensation of the volume increase may be explained by the absence of a concomitant activation of a K+ outward current, resulting in an insufficient efflux of osmolytes during HTS.

Cell Size↗

PubMed1994

Electrophysiological evidence for the presence of an apical H(+)-ATPase in Malpighian tubules of Formica polyctena: intracellular and luminal pH measurements.

Cellular and luminal pH of isolated ant Malpighian tubules were measured in different bath K+ concentrations using double-barrelled pH microelectrodes. The electrochemical gradient for H+ across the basolateral and the apical cell membranes was estimated. In control Ringer (51 mmol/l K+) cell and luminal pH were alkaline with respect to the basolateral solution: 7.77 and 7.36, respectively, versus 7.25. On lowering basolateral K+ concentration to 5 mmol/l or increasing it to 113 mmol/l, luminal pH and to a lesser extent cell pH followed: luminal pH changed to 7.14 and 7.43 and cell pH to 7.69 and 7.82, respectively. In all conditions a cell inward electrochemical gradient for protons across both membranes was observed. Increasing basolateral K+ concentration, which was positively correlated with secretion rate, decreased the cell inwardly directed apical proton gradient; moreover, the apical membrane potential difference decreased as well, from -93 mV in 5 mmol/l K+ to -65 mV in 113 mmol/l K+. Therefore the turnover rate of the electrogenic active proton pump at the apical membrane is facilitated in a high basolateral K+ concentration. The calculated electromotive force of this pump is -159 mV. Comparing the proton with the K+ electrochemical gradient, taken from another study in the same experimental conditions, we find that the apical proton electrochemical gradient can drive K+ extrusion into the lumen for each value of secretion rate.

Animals↗

PubMed1994

Poorly selective cation channels in the apical membrane of A6 cells.

This paper describes a Ca(2+)-blockable, poorly selective cation pathway in the apical membrane of A6 epithelia. This pathway has properties that resemble the cation-selective channels in the toad urinary bladder and frog skin. Transepithelial short circuit currents (Isc) and power density spectra (PDS) of the fluctuations in current were recorded. The basolateral surface of the tissues was exposed to Cl- or SO4(2-) solutions with Na+ as the major cation. Ca(2+)-blockable inward oriented currents and Lorentzian noise were recorded with isotonic (215 mosmol/kg) mucosal Cl- and hypotonic (144 mos-mol/kg serosal SO4(2-) solution with Na+, K+, Rb+ or Cs+ as the major mucosal cation. Experiments with mucosal K+ demonstrated that the cation-selective channel was markedly activated by serosal hypotonicity. Effects of an increased electrical driving force were excluded on the basis of the results obtained with microelectrode experiments and transepithelial voltage clamping. Cell volume expansion induced by isotonic replacements of serosal sucrose by glycerol or urea also activated the cation-selective pathway. Furthermore, the presence of Cl- in the mucosal solution was a prerequisite for a sustained response to hypotonicity or replacements of the organic compounds. Moreover, we found that the cation-selective channels are mainly expressed in the cells during the early period of epithelial growth.

Animals↗

PubMed1993

Poorly selective cation channels in apical membranes of epithelia.

The apical membrane of frog skin contains two types of pathways which allow the passage of several monovalent cations in the absence of external Ca2+. Differences between the two pathways concern their open-close kinetics, selectivity, and the affinity for several blocking agents. Type S channels open and close relatively slowly, whereas type F channels display fast open-close kinetics. Both channel types allow the passage of Na+, K+, and Rb+ currents which are blocked by divalent cations and La3+ added to the extracellular side. Type F channels are permeable for Cs+ which is, however, excluded from type S channels. Shifts in open-close kinetics induced by Mg2+ occur at concentrations below 5 microM for type F channels, whereas more than a tenfold higher dose is required for the type S pathway. UO2(2+) concentrations up to 100 microM only occlude type S channels while 100 microM tetracaine selectively blocks type F channels. Apical membranes of toad urinary bladder, cultured amphibian renal epithelia (A6), and toad colon contain only type F channels. In toad bladder and A6 cells volume expansion strongly activates this pathway. Macroscopic currents carried by Ba2+ and Ca2+ could be recorded after activation of toad bladders with oxytocin and treatment of the apical surface with nanomolar concentrations of Ag+, which seems to interact with a site located at the channel interior.

Amiloride↗

PubMed1993

Ca(2+)-blockable, poorly selective cation channels in the apical membrane of amphibian epithelia. Tetracaine blocks the UO2(2+)-insensitive pathway.

We examined the effect of the local anesthetic tetracaine on the Ca(2+)-blockable, poorly selective cation channels in the isolated skin of Rana temporaria and the urinary bladder of Bufo marinus using noise analysis and microelectrode impalements. Experiments with frog skin demonstrated that mucosal concentrations of the compound up to 100 microM did not affect the Na+ current through type S channels (slowly fluctuating, UO2(2+)-blockable channels) and the associated noise. On the other hand, 20 microM mucosal tetracaine already suffices to inhibit approximately 50% of the current carried by Cs+ and Na+ through channel type F (fast fluctuating, UO2(2+)-insensitive channel) and So of the associated Lorentzian component. With 100 microM of the inhibitor the current and So values were reduced by at least 70-80%. The time course of the response to serosal tetracaine was markedly slower and the effects on the current and So were smaller. Possible effects on the basolateral K+ conductance were excluded on the basis of the lack of response of transepithelial K+ movements to 100 microM tetracaine. UO2(2+) and tetracaine together blocked the poorly selective cation pathways almost completely. Moreover, both agents retain their inhibitory effect in the presence of the other. In toad urinary bladder, the Ca(2+)-blockable channel is also tetracaine blockable. The concentration required for half-maximal inhibition is approximately 100 microM in SO4(2-) and approximately 20 microM in Cl-. The data with tetracaine complement those obtained with UO2(2+) and support the idea that the Ca(2+)-blockable current proceeds through two distinct classes of cation channels. Using tetracaine and UO2(2+) as channel-specific compounds, we demonstrated with microelectrode measurements that both channel types are located in the granulosum cells.

Animals↗

PubMed1993

Ca(2+)-blockable, poorly selective cation channels in the apical membrane of amphibian epithelia. UO2(2+) reveals two channel types.

This study deals with the effect of mucosal UO2(2+) on the Ca(2+)-blockable, poorly selective cation channels in the apical membrane of frog skin and toad urinary bladder. Our data show that UO2(2+) inhibits the Na+ currents through the amiloride-insensitive cation pathway and confirm a previously described stimulatory effect on the amiloride-blockade Na+ transport. Noise analysis of the Ca(2+)-blockable current demonstrates that the divalent also depresses the low-frequency Lorentzian (fc = 11.7 Hz) in the power density spectrum (PDS) and reveals the presence of high-frequency relaxation noise (fc = 58.5 Hz). The action of UO2(2+) is not reversed upon washout and is not accompanied by noise, typically induced by reversible blockers. The divalent merely depresses the plateau of the low-frequency Lorentzian, demonstrating a decrease in the number of conductive cation channels. Similarly, with mucosal K+ and Rb+, UO2(2+) also unmasks the high-frequency Lorentzian by depressing the noise from the slowly fluctuating cation channels (type S). In all experiments with mucosal Cs+, the PDS contains high-frequency relaxation noise (fc = 75.1 Hz in Rana temporaria, and 65.4 Hz in Rana ridibunda). An effect of UO2(2+) on the Cs+ currents and Lorentzian plateaus could not be demonstrated, suggesting that this monovalent cation does not pass through type S channels. Experiments with the urinary bladder revealed only a UO2(2+)-insensitive pathway permeable for Na+, K+, Rb+, and Cs+. We submit that in frog skin two cation-selective channels occur, distinguished by their spontaneous gating kinetics, their sensitivity to UO2(2+), and their permeability for Cs+. In toad urinary bladder, only one kind of cation-selective channel is observed, which resembles the UO2(2+)-insensitive channel in frog skin, with fast open-closed kinetics (type F).

Amiloride↗

PubMed1993

Current-noise analysis of Na absorption in the embryonic coprodeum: stimulation by aldosterone and thyroxine.

The mechanism and regulation of sodium transport in the embryonic coprodeum of chicken were investigated with isolated epithelia in vitro by electrophysiological techniques. Electrogenic sodium transport (INa) was measured in Ussing chambers by the short-circuit current (Isc) technique and identified by the diuretic amiloride or by removal of sodium from the apical medium. Apical sodium channels and the kinetics of amiloride binding were investigated by current-noise analysis. Isc and INa were measured under control conditions and under the influence of in vitro incubation with aldosterone and thyroxine. At 20 days the embryonic coprodeum has an Isc of 12.6 +/- 1.4 microA/cm2 and a transepithelial resistance of 519 +/- 40 omega.cm2. Amiloride blocks 9.0 +/- 1.3 microA/cm2 of the Isc, which represents electrogenic Na+ absorption and can be inhibited by serosal ouabain. Aldosterone does not stimulate Isc or INa, whereas thyroxine increases Isc and INa about threefold. Aldosterone in combination with thyroxine increases Isc and INa further to about five- to sixfold. In both cases the hormonal stimulation can be totally blocked by spironolactone. Current-noise analysis of the apical Na+ entry step reveals amiloride-sensitive Na+ channels with a single-channel current of approximately 2.3 pA and a channel density of 9-16 million/cm2 under stimulated conditions. Half-maximal amiloride block occurs at 0.8-1 microM. The hormones stimulate Na+ absorption by increasing the Na+ channel density and not the single-channel current.(ABSTRACT TRUNCATED AT 250 WORDS)

Absorption↗

PubMed1992

Verapamil blocks basolateral K+ channels in the larval frog skin.

The short-circuit current (Isc) across isolated skin from larval frogs (Rana catesbeiana) was measured when the tissue was bathed with Na2SO4 Ringer solution on the serosal side and with a Ringer solution containing K+ as the primary cation on the mucosal side. When 150 U/ml nystatin was added to the mucosal solution, the Isc increased from 1.4 +/- 0.1 to 35.4 +/- 4.8 microA/cm2. When verapamil was added to the mucosal and serosal Ringer solutions in concentrations between 2.5 and 80 microM, Isc was inhibited in a stepwise manner. At 80 microM, Isc was reduced by 75.3% to 8.74 +/- 1.14 microA/cm2. Analysis of the inhibition of Isc with the direct linear plot method showed that the blockage of Isc could be described by pseudo-first-order kinetics with a Michaelis constant (Km) of 9.59 +/- 2.20 microM. Fluctuation analysis revealed a Lorentzian component in power spectra obtained from preparations treated with 10-80 microM verapamil. The corner frequency of these Lorentzian components increased in a linear manner over this range of verapamil concentrations. The Km calculated from the ratio of the dissociation and association rate constants (k10/k'01) was 39.5 microM. The single-channel currents (i) calculated from the fluctuation analysis parameters decreased significantly between verapamil concentrations of 10 and 80 microM. It appears that the inhibition of K+ channels in the basolateral membrane of this tissue has at least two components.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1992

Effect of forskolin on conductive anion pathways of toad skin.

The effect of the diterpene, forskolin, on pathways for conductive Cl- transport was analyzed using isolated skins of Bufo viridis. Forskolin did not stimulate the voltage-activated Cl- movement from mucosa to serosa; the Lorentzian component in the power density spectrum, which was present at serosa positive clamp potentials under control conditions, decreased significantly. The observation that stimulation of cytosolic adenosine 3'-5'-cyclic monophosphate (cAMP) by forskolin has no effect on the voltage-activated Cl- transport argues against control of this pathway by cAMP. Our data further demonstrate that the forskolin-activated Cl- conductive pathway is also permeable for NO3-. This pathway was studied in absence of mucosal Cl-, which eliminates Cl- movement through the voltage-activated pathway. With SO4(2-) and Cl- on the mucosal and serosal sides, respectively, this forskolin-induced pathway displayed a linear current-voltage relationship. The associated Lorentzians increased at serosa negative clamp potentials. Transepithelial current and plateau value of the Lorentzian were related by a quadratic function, which suggests voltage-independence of number and open-close probability of these conductance sites. Morphological sites for voltage-activated and forskolin-induced conductive Cl- transport remain to be identified.

Animals↗

PubMed1991

Development of aldosterone-stimulation of short-circuit current across larval frog skin.

The short-circuit current (SCC) across isolated skin from bullfrog larvae in developmental stage XXI was small and insensitive to amiloride. Overnight incubation of this tissue with 10(-6) M aldosterone stimulated the SCC from 1.35 +/- 0.55 to 14.55 +/- 4.12 microA.cm-2 with 11.18 +/- 4.46 microA.cm-2 being blocked by 100 microM amiloride. Histologic examination of aldosterone-treated skins revealed a separation of the apical cell layer from the underlying epidermis that was not seen in untreated preparations. The onset of amiloride-sensitive Na+ transport thus coincided with the exposure of the apical surface of newly differentiated epithelial cells. Similar results were obtained with skin from stage XXI larvae whose rate of metamorphosis had been stimulated by 10 micrograms.1-1 thyroxine (T4) but not with skin from T4-treated larvae in stages XIX and XX. Fluctuation analysis of the amiloride-sensitive SCC of the above preparations failed to show a consistent Lorentzian component in the power-density spectrum. Fluctuation analysis was possible on skins from larvae whose development had been accelerated by 7-9 days treatment with 10 micrograms.l-1 triiodothyronine (T3). Aldosterone treatment of these tissues resulted in a significant increase in Na+ channel density.

Aldosterone↗

PubMed1991

Amiloride blockage of Na+ channels in amphibian epithelia does not require external Ca2+.

Noise analysis was used to study the influence of external Ca2+ on the blockage of Na+ transport by amiloride. Experiments were done using frog skin (Rana temporaria and Rana catesbeiana), toad urinary bladder (Bufo marinus) and epithelia of A6 cells. In nondepolarized skins and bladders, removal of Ca2+ from the mucosal bath diminished markedly the inhibitory effect of amiloride. Ca2+ depletion also gave rise to the appearance of an additional noise component related to cation movement through the poorly selective cation channel in the apical membrane [Aelvoet I, Erlij D, Van Driessche W (1988) J Physiol (Lond) 398:555-574; Van Driessche W, Desmedt L, Simaels J (1991) Pflügers Arch 418:193-203]. The amplitude of this Ca(2+)-blockable noise component was elevated by amiloride and markedly exceeded the amiloride-induced Lorentzian noise levels as recorded in the presence of Ca2+. On the other hand, in K(+)-depolarized skins and bladders as well as in nondepolarized epithelial of A6 cells, the Ca(2+)-blockable noise was absent or of much smaller amplitude. Depolarization of frog skin and toad urinary bladder apparently inactivated the poorly selective channels, whereas in A6 cells they were not observed. Under these conditions the typical amiloride-induced blocker noise could also be analysed in the absence of Ca2+ and demonstrated that the on and off rates for amiloride binding were not significantly altered by external Ca2+.(ABSTRACT TRUNCATED AT 250 WORDS)

Amiloride↗

PubMed1991

Blockage of Na+ currents through poorly selective cation channels in the apical membrane of frog skin and toad urinary bladder.

The blockage of Na+ movements through the poorly selective cation channels in the apical membrane of frog skin (Rana temporaria) and toad urinary bladder (Bufo marinus) was investigated with noise, impedance analysis and microelectrode techniques. Na+ currents through this pathway were studied with NaCl Ringer solutions on both sides. After removal of Ca2+ and other divalent cations from the mucosal compartment, a considerable part of Isc became insensitive to amiloride. In frog skin, the inhibitory effect of amiloride in mucosal Ca(2+)-free solutions was highly variable. In some experiments a complete lack of inhibition was observed. Similarly, in the absence of amiloride, the inhibitory effect of mucosal Ca2+ varied strongly among frogs. In the absence of mucosal Ca2+, analysis of the fluctuation in Isc revealed a Lorentzian component in the power density spectrum. The corner frequency (fc) of this spontaneous Lorentzian was 12.3 Hz in frog skin and 347 Hz in the toad urinary bladder. In frog skin, nanomolar concentrations of mucosal Ca2+ induced an additional Lorentzian noise component. Its corner frequency shifted upwards with increasing mucosal Ca2+ concentration ([Ca2+]m). The relation between 2 pi fc and [Ca2+]m was linear at small [Ca2+]m whereas a parabolic increase of fc was observed at the highest [Ca2+]m. In the bladder, nanomolar concentrations of mucosal Ca2+ did not induce an additional noise component but modified the spontaneous Lorentzian noise by increasing fc proportionally with [Ca2+]m. Microelectrode recordings demonstrated that at least part of the Ca(2+)-blockable current passes through the granulosum cells and confirmed the apical localization of the poorly selective cation channel. The lack of the inhibitory effect of amiloride in Ca(2+)-free solutions seems to originate from the parallel arrangement of the amiloride- and Ca(2+)-blockable pathways and from influences of the blockage of apical channels on the basolateral membrane conductances. The latter cross-talk seems to find its origin in the voltage dependence of the basolateral membrane conductance [Garty H (1984) J Membr Biol 77:213-222; Nagel W (1985) Pflügers Arch 405 [Suppl 1]:S39-S43].

Amiloride↗

PubMed1991

Chloride-related current fluctuation in amphibian skin.

Fluctuation in transepithelial current was analysed in skins of Rana esculenta and Bufo viridis. Perturbation of the tissues to serosa + 40 mV or + 80 mV activated a pathway for conductive Cl transport and led to the appearance of a Lorentzian component in the power density spectrum. The presence of Cl on the mucosal side was mandatory for the observation of Lorentzians. Corner frequencies, fc, ranged between 48 Hz and 102 Hz. The mean value was slightly larger at 40 mV than at 80 mV (88.3 +/- 4.4 versus 72.5 +/- 6.4 Hz). Plateau values, So, were proportional to the magnitude of the activated Cl current. Single-channel currents of 48.3 +/- 5.2 fA and 93.0 +/- 8.7 fA were calculated at 40 mV and 80 mV, respectively, with the assumption of equal open and closed probabilities of the channels. From these data, a voltage-dependent open-channel conductance of about 1.2 pS is obtained. Activation of Cl conductance was associated with an increase in channel density. Stimulation of Cl conductance by procaine did not affect So and fc despite considerable stimulation of transepithelial Cl current. This could indicate that non-gated channels participate in the voltage-sensitive Cl conductance. Mucosal application of inhibitors of Cl conductance (3',5-dichlorodiphenylaminocarboxy acid, MK-196) decreased So in a concentration-dependent manner, but had no effect on fc. The data could support the hypothesis that the voltage-sensitive transepithelial Cl transport is localized to a cellular compartment, which is most likely the mitochondria-rich cells.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1991

Microelectrode study of voltage-dependent Ba2+ and Cs+ block of apical K+ channels in the skin of Rana temporaria.

The blockage of the apical K+ channels in frog species Rana temporaria by Ba2+ and Cs+ is strongly voltage-dependent. The interaction of both blockers with the K+ channels was studied by recording relations between the K+ currents (IK) and the transepithelial and intracellular potential. Mucosal Ba2+ and Cs+ depress IK, hyperpolarize the cell and induce pronounced nonlinearities in the current/voltage (I/V) relations. The nonlinearities are caused by the voltage-dependent interaction of Ba2+ and Cs+ with the binding site. Consequently, the apical membrane resistance not only depends on the blocker concentration but also on the apical membrane potential. Also the fractional resistance, fRa, and the voltage divider ratio, fVa, will change with blocker concentration and voltage. Owing to this non-ohmic behaviour, measurements of fVa in the presence of Ba2+ deviate markedly from the expected fRa values. The inhibitory effect of Ba2+ and Cs+ was analysed at different transepithelial and apical membrane voltages. The relation between the Michaelis-Menten constants and the voltage could be fitted with equations based on Eyring rate theory with the assumption of a single binding site. With this model we calculated the relative electrical position of the binding site for the blocker (delta), referred to the extracellular side of the channel. We obtained for Ba2+, delta = 0.34 +/- 0.05 and for Cs+, delta = 0.81 +/- 0.01. Comparison of the results from apical and transepithelial I/V relations demonstrates that the analysis of the transepithelial data provides overestimated values of the Hill coefficient and results in an underestimation of delta.

Animals↗

PubMed1991

K+ secretion across frog skin. Induction by removal of basolateral Cl-.

We examined the development of K+ secretion after removing Cl- from the basolateral surface of isolated skins of Rana temporaria using noise analysis. K+ secretion was defined by the appearance of a Lorentzian component in the power density spectrum (PDS) when Ba2+ was present in the apical bath (0.5 mM). No Lorentzians were observed when tissues were bathed in control, NaCl Ringer solution. Replacement of basolateral Cl- by gluconate, nitrate, or SO4- (0-Clb) yielded Lorentzians with corner frequencies near 25 Hz, and plateau values (So) that were used to estimate the magnitude of K+ secretion through channels in the apical cell membranes of the principal cells. The response was reversible and reproducible. In contrast, removing apical Cl- did not alter the PDS. Reduction of basolateral Cl- to 11.5 mM induced Lorentzians, but with lower values of So. Inhibition of Na+ transport with amiloride or by omitting apical Na+ depressed K+ secretion but did not prevent its appearance in response to 0-Clb. Using microelectrodes, we observed depolarization of the intracellular voltage concomitant with increased resistance of the basolateral membrane after 0-Clb. Basolateral application of Ba2+ to depolarize cells also induced K+ secretion. Because apical conductance and channel density are unchanged after 0-Clb, we conclude that K+ secretion is "induced" simply by an increase of the electrical driving force for K+ exit across this membrane. Repolarization of the apical membrane after 0-Clb eliminated K+ secretion, while further depolarization increased the magnitude of the secretory current. The cell depolarization after 0-Clb is most likely caused directly by a decrease of the basolateral membrane K+ conductance. Ba2(+)-induced Lorentzians also were elicited by basolateral hypertonic solutions but with lower values of So, indicating that cell shrinkage per se could not entirely account for the response to 0-Clb and that the effects of 0-Clb may be partly related to a fall of intracellular Cl-.

Amiloride↗