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L Reuss

Publications and source records attributed to L Reuss.

At least 73 records · Page 4Linked to original sources
PubMed1989

pH effects on basolateral membrane ion conductances in gallbladder epithelium.

The pH sensitivity of the basolateral membrane voltage of Necturus gallbladder epithelial cells (Vcs) was evaluated with conventional and pH-sensitive intracellular microelectrodes. Elevating solution CO2 from 1 to 5% (at constant [HCO3-] = 10 mM) caused a depolarization of Vcs from -76 +/- 3 to -60 +/- 2 mV and a decrease in intracellular pH (pHi) from 7.36 +/- 0.04 to 7.05 +/- 0.03. Serosal exposure to a 50 mM HCO3(-)-5% CO2 solution [at constant extracellular pH (pHo)] caused a similar cell acidification (delta pHi = 0.27), whereas at 3 min Vcs was unchanged. Exposure to 1 mM HCO3- (at constant CO2) depolarized Vcs from -77 +/- 2 to -56 +/- 2 mV and caused a small decrease in pHi (from 7.36 +/- 0.03 to 7.33 +/- 0.03). These results indicate that the observed depolarizations of Vcs are attributable to changes in pHo and not in pHi. Basolateral membrane potassium conductance (GK) congruent to chloride conductance (GCl) congruent to 0.50 mS/cm2 in 10 mM HCO3(-)-1% CO2 Ringer. The depolarization of Vcs caused by elevation of serosal [K+] in 50 mM HCO3(-)-5% CO2 was similar to that observed under control conditions. In contrast, the depolarization of Vcs elicited by elevating serosal [K+] was reduced by about two-thirds in 1 mM HCO3-, whereas the depolarization caused by reduction of serosal [Cl-] was increased twofold in 1 mM HCO3-, compared with control. Inasmuch as the apparent ratio of membrane resistances remained unchanged during serosal solution acidification, the most likely explanation for the observed decrease in Vcs is a reduction of basolateral K+ permeability concomitant with an increase in Cl- permeability.

Animals↗
PubMed1989

Electrophysiological effects of mucosal Cl- removal in Necturus gallbladder epithelium.

The factors responsible for the cell membrane hyperpolarization elicited in Necturus gallbladder epithelium on Cl- removal from the mucosal bathing solution were evaluated with conventional and ion-sensitive microelectrode techniques. Cl- removal causes reversal of apical Cl- -HCO3- exchange, resulting in a fall in intracellular Cl- activity (aiCl) and an increase in intracellular pH (pHi). Concomitantly, the cell membranes hyperpolarize to values close to the K+ equilibrium potential (EK), aiNa falls, and aiK rises. The observed changes in membrane voltage are not attributable to a pHi-dependent increase in cell membrane K+ permeability (PK), because 1) the cell membrane resistances increased and 2) elevating solution partial pressure of CO2 (PCO2) to counterbalance the cellular alkalinization on mucosal Cl- removal caused a further hyperpolarization of the cell membranes to values greater than EK. This additional hyperpolarization was related to the activity of the Na+ pump, inasmuch as it was accompanied by an increase in aiNa and was ouabain sensitive. These results are consistent with, but do not prove, pump electrogenicity. During the period of Cl- removal from the mucosal bathing solution, the cell membrane depolarization caused by raising serosal K+ concentration was increased, whereas the depolarization caused by lowering serosal Cl- concentration was decreased, compared with substitutions under control conditions. These results indicate that mucosal Cl- removal causes a decrease in basolateral PCl, which we speculate could be due to a decrease in cell volume. We conclude that the hyperpolarization of the cell membranes on mucosal Cl- removal is primarily due to the combined effects of the fall in basolateral PCl and the increase in basolateral ECl.

Animals↗
PubMed1988

Dependence of cell membrane conductances on bathing solution HCO3-/CO2 in Necturus gallbladder.

The effects of bathing solution HCO3-/CO2 concentrations on baseline cell membrane voltages and resistances were measured in Necturus gallbladder epithelium with conventional intracellular microelectrode techniques. Gallbladders were bathed in either low HCO3-/CO2 Ringer's solutions (2.4 mM HCO3-/air or 1 mM HEPES/air) or a high HCO3-/CO2 Ringer's (10 mM HCO3-/1% CO2). The principal finding of these studies was that the apical membrane fractional resistance (fRa) was higher in tissues bathed in the 10 mM HCO3-/CO2 Ringer's, averaging 0.87 +/- 0.06, whereas fRa averaged 0.63 +/- 0.07 and 0.48 +/- 0.08 in 2.4 mM HCO3- and 1 mM HEPES, respectively. Intraepithelial cable analysis was employed to obtain estimates of the individual apical (Ra) and basolateral membrane (Rb) resistances in tissues bathed in 10 mM HCO3-/1% CO2 Ringer's. Compared to previous resistance measurements obtained in tissues bathed in a low HCO3-/CO2 Ringer's, the higher value of fRa was found to be due to both an increase in Ra and a decrease in Rb. The higher values of fRa and lower values of Rb confirm the recent observations of others. To ascertain the pathways responsible for these effects, cell membrane voltages were measured during serosal solution K+ and Cl- substitutions. The results of these studies suggest that an electrodiffusive Cl- transport mechanism exists at the basolateral membrane of tissues bathed in a 10 mM HCO3-/1% CO2 Ringer's, which can explain in part the fall in Rb. The above observations are discussed in terms of a stimulatory effect of solution [HCO3-]/PCO2 on transepithelial fluid transport, which results in adaptive changes in the conductive properties of the apical and basolateral membranes.

Animals↗
PubMed1988

Voltage- and time dependence of apical membrane conductance during current clamp in Necturus gallbladder epithelium.

The effects of short (1 sec) and long (1 min) transepithelial current clamps on membrane voltages and resistances of Necturus gallbladder were investigated. Transepithelial and cell membrane current-voltage relationships determined from 1-sec clamps revealed that: a) depolarization of the apical membrane voltage (Vmc) results in a marked decrease in apical membrane fractional resistance (fRa), whereas hyperpolarization of Vmc results in either no change in fRa or a small increase, and b) the voltage-dependent changes in fRa are essentially complete within 500 msec. Exposure of the tissue to 5 mM TEA+ on the mucosal side caused no significant change in baseline Vmc (-69 +/- 2 mV) and yet virtually abolished the voltage dependence of fRa. A possible interpretation of these results is that two types of K+ channels exist in the apical membrane, with different voltage dependencies and TEA+ sensitivities. Acidification or Ba2+ addition to the mucosal solution also reduced the voltage-dependent changes in fRa. The time courses of the changes in fRa and in the cable properties of the epithelium were assessed during 1-min transepithelial current clamps (+/- 200 microA/cm2). No secondary change in fRa was observed with mucosa-to-serosa currents, but a slow TEA+-sensitive decrease in fRa (half-time of seconds) was evident with serosa-to-mucosa currents. Cable analysis experiments demonstrated that the initial (less than 500 msec) voltage-dependent decrease in fRa is due to a fall in apical membrane resistance. The later decrease in fRa is due to changes in both cell membrane resistances attributable to the increase in transcellular current flow resulting from a fall in paracellular conductance. The voltage dependence of the apical membrane conductance is a more significant problem in estimating fRa than the current-induced effects on the lateral intercellular spaces. In principle, TEA+ can be used to prevent the nonlinear behavior of Ra during measurements of the voltage divider or membrane resistance ratio.

Animals↗
PubMed1987

Cyclic AMP inhibits Cl-/HCO3- exchange at the apical membrane of Necturus gallbladder epithelium.

Intracellular microelectrode techniques were employed to study the effect of cyclic AMP on apical membrane Cl-/HCO3- exchange and electrodiffusive HCO3- transport in Necturus gallbladder epithelium. Intracellular cAMP levels were raised by addition of either the phosphodiesterase inhibitor theophylline (3 X 10(-3) M) or the adenylate cyclase activator forskolin (10(-5) M) to the serosal bathing solution. Measurements of pH in a poorly buffered control mucosal solution upon stopping superfusion show acidification, owing to secretion of both H+ and HCO3-. When the same experiment is performed after addition of amiloride or removal of Na+ from the mucosal bathing medium, alkalinization is observed since H+ transport is either inhibited or reversed, whereas HCO3- secretion persists. The changes in pH in both amiloride or Na-free medium were significantly decreased in theophylline-treated tissues. Theophylline had no effect on the initial rates of fall of intracellular Cl- activity (aCli) upon reducing mucosal solution [Cl-] to either 10 or 0 mM, although membrane voltage and resistance measurements were consistent with stimulation of apical membrane electrodiffusive Cl- permeability. Estimates of the conductive flux, obtained by either reducing simultaneously mucosal [Cl-] and [HCO3-] or lowering [Cl-] alone in the presence of a blocker of anion exchange (diphenylamine-2-carboxylate), indicate that elevation of intracellular cAMP inhibited the anion exchanger by approximately 50%. Measurements of net Cl- uptake upon increasing mucosal Cl- from nominally zero to levels ranging from 2.5 to 100 mM suggest that the mechanism of inhibition is a decrease in Vmax. Consistent with these results, the rate of intracellular alkalinization upon reducing external Cl- was also inhibited significantly by theophylline. Reducing mucosal solution [HCO3-] from 10 to 1 mM under control conditions caused intracellular acidification and an increase in aCli. Theophylline inhibited both changes, by 62 and 32%, respectively. These data indicate that elevation of intracellular cAMP inhibits apical membrane anion (Cl-/HCO3-) exchange. Studies of the effects of rapid changes in mucosal [HCO3-] on membrane voltages and the apparent ratio of membrane resistances, both in the presence and in the absence of theophylline, with or without Cl- in the mucosal solution, do not support the hypothesis that cAMP produces a sizable increase in apical membrane electrodiffusive HCO3- permeability.

Animals↗
PubMed1987

Diphenylamine-2-carboxylate blocks Cl(-)-HCO3- exchange in Necturus gallbladder epithelium.

Intracellular microelectrode techniques were employed to study the effects of diphenylamine-2-carboxylate (DPC) on ion transport in Necturus gallbladder epithelium. Under control conditions, addition of DPC to the mucosal bathing solution caused a concentration-dependent, reversible hyperpolarization of both cell membranes with no measurable resistance changes. In addition, DPC caused the following effects, all consistent with inhibition of apical membrane Cl(-)-HCO3- exchange: fall in intracellular Cl- activity (aCli), increase in intracellular pH (pHi), reduction of the changes in aCli and pHi produced by lowering mucosal solution [Cl-], and reduction of the change in pHi produced by lowering mucosal solution [HCO3-]. Similar studies in theophylline-treated preparations indicate that DPC also inhibits anion exchange under these conditions, but has no effect on the apical membrane electrodiffusive Cl- permeability induced by cyclic AMP. Under these conditions, DPC caused cell membrane hyperpolarization but had no effect on the apparent ratio of membrane resistances. In addition, DPC had no effects on the rapid changes in apical membrane voltage elicited by altering mucosal [Cl-], but caused significant reductions of the slower, secondary voltage changes observed in response to changes in mucosal [Cl-], and the changes in aCli and pHi produced by lowering mucosal [Cl-]. Because others have demonstrated that DPC blocks Cl- channels in other epithelia (Distefano, A., M. Wittner, E. Schlatter, H. J. Lang, H. Englert, and R. Greger. Diphenylamine-2-carboxylate, a blocker of the Cl(-)-conductive pathway in Cl(-)-transporting epithelia. Pfluegers++ Arch. 405: S95-S100, 1985), it is possible that the structures of those channels and that induced by cyclic AMP in Necturus gallbladder are different. Because of its relatively high affinity and rapid reversibility, DPC may become useful in studies of anion exchange in other cells.

Animals↗
PubMed1987

Protamine alters apical membrane K+ and Cl- permeability in gallbladder epithelium.

Protamine addition to the solution bathing the mucosal side of Necturus gallbladder epithelium (25-100 mg/l) caused depolarization of both cell membranes, a mucosa-negative change in transepithelial voltage, an increase in the apical membrane resistance (Ra) followed by a decrease, and a monotonic increase in transepithelial resistance (Rt). In protamine (25 mg/l), the change in apical membrane voltage elicited by elevating mucosal solution [K+] from 2.5 to 92.5 mM was reduced from 66 +/-2 to 38 +/- 5 mV (P less than 0.001). The K+-induced fall in Ra was also reduced in protamine. These effects could also be elicited by elevating mucosal solution [K+] simultaneously with the addition of protamine and by transient addition of protamine during exposure to the high K+ medium. The effect of protamine on the electrodiffusive Cl- permeability of the apical membrane (PCl) was studied both in control and forskolin-treated tissues. In the absence of forskolin, the hyperpolarization of Vmc produced by lowering mucosal [Cl-] to 10 mM was reversed to a small depolarization; in forskolin, the initial depolarization produced by lowering [Cl-] was significantly increased. Finally, exposure to protamine in the absence of forskolin produced an initial fall in intracellular Cl- activity. Our results indicate that protamine decreases apical membrane K+ permeability and increases apical membrane PCl. The time course of the effects of protamine suggests the possibility of an initial effect on surface potential, followed by secondary actions mediated by intracellular events.

Animals↗
PubMed1985

Changes in cell volume measured with an electrophysiologic technique.

Epithelial cells of the gallbladder of Necturus maculosus were loaded with tetramethylammonium (Me4N+) by transient exposure of the apical (lumen-facing) surface to a solution of high Me4N+ concentration containing also the polyene antibiotic nystatin. Upon removal of nystatin, in the continued presence of Me4N+, spontaneous restoration of the native ionic permeability of the apical cell membrane was observed. At this time, external Me4N+ was removed; intracellular [Me4N+] measured with ion-sensitive microelectrodes was 2-15 mM and remained unchanged for several hours. Changes in cell volume were estimated from the changes in intracellular [Me4N+] produced by alterations in the osmolality of the mucosal bathing solution. Assuming that such changes are caused entirely by water fluxes across the apical membrane, the minimum value of its hydraulic permeability coefficient (Lp) was 1-3 X 10(-3) cm.sec-1.(osmoles/kg)-1, suggesting that an osmolality difference across the apical membrane as small as 1-3 milliosmoles/kg could explain the average rate of transepithelial water transport. These results agree with optical measurements [Persson, B. O. & Spring, K. R. (1982) J. Gen. Physiol. 79, 481-505]. The effective thickness of the apical unstirred layer was estimated from the time courses of both the apical membrane voltage and the response of an extracellular K+-sensitive microelectrode to an increase in [K+] in the mucosal bath. Since changes in concentration of the osmotically active solute at the membrane surface were thus shown to be significantly delayed by diffusion, the Lp value, calculated assuming a step-change in osmolality, is an underestimate.

Animals↗
PubMed1985

Cyclic AMP inhibits Na+/H+ exchange at the apical membrane of Necturus gallbladder epithelium.

The effects of elevating intracellular cAMP levels on Na+ transport across the apical membrane of Necturus gallbladder epithelium were studied by intracellular and extracellular microelectrode techniques. Intracellular cAMP was raised by serosal addition of the phosphodiesterase inhibitor theophylline (3 mM) or mucosal addition of either 8-Br-cAMP (1 mM) or the adenylate cyclase activator forskolin (10 microM). During elevation of intracellular cAMP, intracellular Na+ activity (alpha Nai) and intracellular pH (pHi) decreased significantly. In addition, acidification of the mucosal solution, which contained either 100 or 10 mM Na+, was inhibited by approximately 50%. The inhibition was independent of the presence of Cl- in the bathing media. The rates of change of alpha Nai upon rapid alterations of mucosal [Na+] from 100 to 10 mM and from 10 to 100 mM were both decreased, and the rate of pHi recovery upon acid loading was also reduced by elevated cAMP levels. Inhibition was approximately 50% for all of these processes. These results indicate that cAMP inhibits apical membrane Na+/H+ exchange. The results of measurements of pHi recovery at 10 and 100 mM mucosal [Na+] and a kinetic analysis of recovery as a function of pHi suggest that the main or sole mechanism of the inhibitory effect of cAMP is a reduction in the maximal rate of acid extrusion. In conjunction with the increase in apical membrane electrodiffusional Cl- permeability, produced by cAMP, which causes a decrease in net Cl- entry (Petersen, K.-U., and L. Reuss, 1983, J. Gen. Physiol., 81:705), inhibition of Na+/H+ exchange contributes to the reduction of fluid absorption elicited by this agent. Similar mechanisms may account for the effects of cAMP in other epithelia with similar transport properties. It is also possible that inhibition of Na+/H+ exchange by cAMP plays a role in the regulation of pHi in other cell types.

Animals↗
PubMed1985

Electrophysiological effects of propionate and bicarbonate on gallbladder epithelium.

The effects of propionate and HCO3- on cell membrane potentials in Necturus gallbladder epithelium were investigated using microelectrode techniques in vitro. Bilateral exposure to either propionate or HCO3- (25 mM each) hyperpolarized both cell membranes by about 12 mV. Mucosal addition of either substance produced cyclic changes in voltage of both cell membranes, which attenuated spontaneously, whereas serosal addition caused sustained hyperpolarization. By intraepithelial cable analysis it was shown that both cell membrane conductances rose during the hyperpolarization. Experiments using substitution of mucosal K+ for Na+ revealed that the relative K+ permeability (PK) of the apical membrane was enhanced during the hyperpolarization induced by mucosal (or serosal) propionate (or HCO3-). These effects are mediated by increases in PK at both membranes, with a larger basolateral effect. We suggest that this mechanism accounts for the higher cell membrane potential values measured in epithelia bathed in HCO3--or propionate-containing solutions. Inasmuch as both propionate and HCO3- stimulate fluid absorption in gallbladder epithelium, the increase in cell membrane PK may represent an adaptive response of the cells to regulate their solute content.

Animals↗
PubMed1984

Na+-H+ exchange and Na+ entry across the apical membrane of Necturus gallbladder.

The role of Na+-H+ exchange in Na+ transport across the apical membrane was evaluated in Necturus gallbladder epithelium by means of intracellular Na+ activity (aNai) and 22Na+ uptake measurements. Under control conditions, complete replacement of Na+ in the mucosal solution with tetramethylammonium reduced aNai from 14.0 to 6.9 mM in 2 min (P less than 0.001). Mucosal addition of the Na+-H+ exchange inhibitor amiloride (10(-3) M) reduced aNai from 15.0 to 13.3 mM (P less than 0.001), whereas bumetanide (10(-5) and 10(-4) M) had no effect. Na+ influx across the apical membrane was studied by treating the tissues with ouabain, bathing them in Na-free solutions, and suddenly replacing the mucosal solution with an Na-containing solution. When the mucosal solution was replaced with Na-Ringer's, aNai increased at approximately 11 mM/min. This increase was inhibited by 54% by amiloride (10(-3) M, P less than 0.001) and was unaffected by bumetanide (10(-5) M). Amiloride-inhibitable Na+ fluxes across the apical membrane were also induced by the imposition of pH gradients. Na+ influx was also examined in tissues that had not been treated with ouabain. Under control conditions, 22Na+ influx from the mucosal solution into the epithelium was linear over the first 60 s and was inhibited by 40% by amiloride (10(-3) M, P less than 0.001) and by 19% by bumetanide (10(-5) M, P less than 0.025). We conclude that Na+-H+ exchange is a major pathway for Na+ entry in Necturus gallbladder, which accounts for at least half of apical Na+ influx both under transporting conditions and during exposure to ouabain. Bumetanide-inhibitable Na+ entry mechanisms may account for only a smaller fraction of Na+ influx under transporting conditions, and cannot explain influx in ouabain-treated tissues. These results support the hypothesis that NaCl entry results primarily from the operation of parallel Na+-H+ and Cl--HCO-3 exchangers, and not from a bumetanide-inhibitable NaCl cotransporter.

Amiloride↗
PubMed1984

Cl-/HCO3- exchange at the apical membrane of Necturus gallbladder.

The hypothesis of Cl-/HCO3- exchange across the apical membrane of the epithelial cells of Necturus gallbladder was tested by means of measurements of extracellular pH (pHo), intracellular pH (pHi), and Cl- activity (alpha Cli) with ion-sensitive microelectrodes. Luminal pH changes were measured after stopping mucosal superfusion with a solution of low buffering power. Under control conditions, the luminal solution acidifies when superfusion is stopped. Shortly after addition of the Na+/H+ exchange inhibitor amiloride (10(-3) M) to the superfusate, alkalinization was observed. During prolonged (10 min) exposure to amiloride, no significant pHo change occurred. Shortly after amiloride removal, luminal acidification increased, returning to control rates in 10 min. The absence of Na+ in the superfusate (TMA+ substitution) caused changes in the same direction, but they were larger than those observed with amiloride. Removal of Cl- (cyclamate or sulfate substitution) caused a short-lived increase in the rate of luminal acidification, followed by a return to control values (10-30 min). Upon re-exposure to Cl-, there was a transient reduction of luminal acidification. The initial increase in acidification produced by Cl- removal was partially inhibited by SITS (0.5 mM). The pHi increased rapidly and reversibly when the Cl- concentration of the mucosal bathing solution was reduced to nominally 0 mM. The pHi changes were larger in 10 mM HCO3-Ringer's than in 1 mM HEPES-Ringer's, which suggests that HCO3- is transported in exchange for Cl-. In both HEPES- and HCO3-Ringer's, SITS inhibited the pHi changes. Finally, intracellular acidification or alkalinization (partial replacement of NaCl with sodium propionate or ammonium chloride, respectively) caused a reversible decrease or increase of alpha Cli. These results support the hypothesis of apical membrane Cl-/HCO3- exchange, which can be dissociated from Na+/H+ exchange and operates under control conditions. The coexistence at the apical membrane of Na+/H+ and Cl-/HCO3- antiports suggests that NaCl entry can occur through these transporters.

Animals↗
PubMed1984

Independence of apical membrane Na+ and Cl- entry in Necturus gallbladder epithelium.

Transepithelial fluid transport (Jv) and intracellular Na+ and Cl- activities (aNai, aCli) were measured in isolated Necturus gallbladders to establish the contribution of different proposed apical membrane entry mechanisms to transepithelial salt transport. In 10 mM HCO3-Ringer's, Jv was 13.5 +/- 1.1 microliter X cm-2 X h-1, and was significantly reduced by a low bicarbonate medium and by addition of amiloride (10(-3)M) or SITS (0.5 X 10(-3)M) to the mucosal bathing solution. Bumetanide (10(-5)M) was ineffective. Bilateral Na+ removal abolished Jv. The hypothesis of NaCl cotransport was rejected on the basis of the following results, all obtained during mucosal bathing solution changes: during Na+ removal, aNai fell 4.3 times faster than aCli; during Cl- removal, aCli fell 7.5 times faster than aNai; amiloride (10(-3) M) reduced aNai at a rate of 2.4 +/- 0.3 mM/min, whereas aCli was not changed; bumetanide (10(-5) M) had no significant effects on Jv or aCli. The hypothesis of Na-K-Cl cotransport was rejected for the same reasons; in addition, K+ removal from the mucosal bathing solution (with concomitant Ba2+ addition) did not alter aNai or aCli. The average rate of NaCl entry under normal transporting conditions, estimated from Jv, assuming that the transported fluid is an isosmotic NaCl solution, was 22.5 nmol X cm-2 X min-1. Upon sudden cessation of NaCl entry, assuming no cell volume changes, aNai and aCli should fall at an average rate of 4.8 mM/min. To compare this rate with the rates of Na+ and Cl- entry by ion exchange, the Na+ or Cl- concentration in the mucosal bathing solution was reduced rapidly to levels such that electroneutral cation or anion exchange, respectively, should cease. The rate of Na+ or Cl- entry before this maneuver was estimated from the initial rate of fall of the respective intracellular ionic activity upon the mucosal solution substitution. aNai and aCli decreased at initial rates of 3.7 +/- 0.4 and 5.9 +/- 0.8 mM/min, respectively. The rate of fall of aNai upon reduction of external [Na] was not affected by amiloride (10(-3) M), and the rate of fall of aCli upon reduction of external [Cl] was unchanged by SITS (0.5 X 10(-3) M), which indicates that net cation or anion exchange was, in fact, abolished by the changes in Na+ and Cl- gradients, respectively. I conclude that double exchange (Na+/H+ and Cl-/HCO-3) is the predominant or sole mechanism of apical membrane NaCl entry in this epithelium.

Absorption↗
PubMed1983

Cyclic AMP-induced chloride permeability in the apical membrane of Necturus gallbladder epithelium.

The effects of theophylline, 8-Br-cAMP, and cAMP on necturus gallbladder epithelium were investigated using microelectrode techniques. Each of these substances depolarized the cell membranes by approximately 15 mV and decreased the apparent ratio of apical to basolateral membrane resistances to a value not significantly different from zero. Examination of the ionic selectivity of the apical membrane by ion substitutions in the mucosal bathing medium revealed a large increase in Cl permeability with no apparent changes in K and Na permeabilities. Intracellular Cl activity ((a)CL(i)) was measured using Cl- sensitive liquid ion-exchanger microelectrodes. Under control conditions, (a)Cl(i) was approximately 20 mM, 2.5 times higher than the value expected for equilibrium distribution ((a)Cl(i/eq). After addition of 8-Br-cAMP, (a)Cl(i) decreased within less than 60 s to approximately 13 mM, a value not significantly different from ((a)Cl(i/eq)). Virtually identical results were obtained with theophylline. Under control conditions, luminal Cl removal caused (a)Cl(i) to fall at an initial rate of 1.8 mM/min, whereas in tissues exposed to 8-Br- cAMP or theophylline a rate of 11.6 mM/min was observed. The apical membrane Cl transference number was estimated from the change of (a)Cl(i) upon exposure to 8-Br-cAMP as well as from the changes in apical membrane potential during variation of the luminal Cl concentration. The results, 0.91 and 0.88, respectively, are indicative of a high Cl permeability of the apical membrane during cAMP. This effect may explain, at least in part, the complete inhibition of fluid absorption produced by theophylline in this tissue. Moreover, enhancement of apical membrane Cl permeability may account for a variety of cAMP effects in epithelial tissues.

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed1983

Intracellular ion activities and Cl-transport mechanisms in bullfrog corneal epithelium.

Cell membrane potentials, cell membrane resistances, and intracellular ionic activities were measured in bullfrog corneal epithelium. Equivalent circuit analysis was performed by adding adenosine to the apical surface and assuming that only the apical membrane is initially affected. From single-ion substitutions in the apical bathing solution, the apical membrane was found to have a high Cl- permeability, a low K+ permeability, and an unmeasurably small Na+ permeability. Under control conditions intracellular Cl- activity (aCli) was 22 +/- 2 (SE) mM, intracellular Na+ activity (aNai) was 14 +/- 3 mM, and intracellular K+ activity (aKi) was 106 +/- 5 mM. The electrical potential differences across apical and basolateral membranes were about 50 and 67 mV, respectively, both cell negative. aCli and aKi are higher, whereas aNai is much lower than predicted for equilibrium distribution. Inasmuch as Cl- is transported from the basolateral (stromal) to the apical (tear) side, basolateral entry of this anion is uphill and apical exit is downhill. Basolateral entry is Na+ dependent, as evidenced by a fall of aCli to near-equilibrium values after basolateral Na+ removal. The electrochemical gradient for Cl- efflux across the apical membrane is large enough to account for Cl- transport by electrodiffusion only. Na+ removal from the basolateral solution causes a reversible decrease of apical membrane Cl- permeability. The results support the hypothesis that net transepithelial Cl- transport results from coupled NaCl entry (or an equivalent process) at the basolateral membrane and electrodiffusional Cl- exit at the apical membrane.

Animals↗