Electrical properties of the cellular transepithelial pathway inNecturus gallbladder: III. Ionic permeability of the basolateral cell membrane

Abstract

The ionic permeability of the basolateral membrane of Necturus gallbladder epithelium was studied with intracellular microelectrode techniques. After removal of most of the subepithelial tissue (to reduce unstirred layer thickness), impalements were performed from the serosal side, and ionic substitutions were made in the serosal solution while a microelectrode was kept in a cell. Thus, it was possible to obtain continuous (and reversible) records of transepithelial and cell membrane potentials and to measure intermittently the transepithelial resistance and the ratio of cell membrane resistances. From these data and the mean value of the equivalent resistance of the cell membranes in parallel (obtained from cable analysis in a different group of tissues), absolute cell membrane and shunt resistances and equivalent electromotive forces (emfs) were calculated. From the changes of basolateral membrane emf (Eb) produced by the substitutions, the conductance (G) and permeability (P) of the membrane for K, Cl and Na were estimated. Potassium-for-sodium substitutions produced large reductions of both cell membrane potentials, of Eb, and of the resistance of the basolateral membrane (Rb), indicating high GK and PK. Chloride substitution with isethionate or sulfate resulted in smaller changes of cell membrane potentials and Eb and in no significant change of Rb, indicating small but measurable values of GCl and PCl. Sodium substitutions with N-methyl-D-glucamine (NMDG) resulted in cell potential changes entirely attributable to the biionic potential produced in the shunt pathway (PNa greater than PNMDG), and in no significant changes of Rb or Eb, indicating that GNa and PNa are undetectable. The question of the mechanism of Cl transport across the basolateral membrane was addressed by comparing the mean rate of transepithelial Cl transport : formula, see text: and the predicted passive Cl flux across the basolateral membrane (from the membrane Cl conductance, potential, and Cl equilibrium potential). The conclusion is that only a very small fraction of the Cl flux across the basolateral membrane can be electrodiffusional. Since the paracellular Cl conductance is also too low to account for : formula, see text:, these results suggest the presence of a neutral mechanism of Cl extrusion from the cells. This could be a NaCl pump, a downhill KCl transport mechanism, or a Cl-HCO3 exchange mechanism.

Mechanism of the effect of cyanide on cell membrane potentials in Necturus gall‐bladder epithelium.

“…First, its rapidity: at high cyanide concentration the effect is maximal within 30 sec. This time course is consistent with the apparent thickness of the unstirred layer in such preparations (Reuss, 1979 even at 10 mm concentration. Thirdly, there is a consistent lack of initial depolarization: in squid giant axon, cyanide produces hyperpolarization, but this effect takes a longer time and is preceded by a transient small depolarization, interpreted as being caused by inhibition of the (electrogenic) sodium pump (De Weer, 1978).…”

“…Potentials and resistances were measured as previously described (Reuss & Finn, 1975a, b;Reuss, 1979;Reuss & Grady, 1979a). Glass micro-electrodes with an inner fibre were filled with 3 M-potassium chloride or 4 M-potassium acetate and selected for tip resistances of 20-50 MCI and low tip potentials.…”

“…During exposure to cyanide, sodiumRinger and potassium-Ringer solutions had the same cyanide concentration. In experiments in which the time course of the effect of cyanide or the selectivity of the basolateral membrane were studied, the tissues were mounted serosal side up, the subepithelial layers removed as decribed before (Reuss, 1979), and the cells impaled from the basolateral surface.…”

Active sodium transport and fluid secretion in the gall‐bladder epithelium of Necturus.

“…If all the entry of Na is to occur by electrodiffusion a permeability coefficient of 08-P17 x 10-6 cm s-1 can be calculated from the constant field theory (Hodgkin & Katz, 1949). This is much higher than the values estimated by cable analysis and ionic substitutions (Van Os & Slegers, 1975;Reuss & Finn, 1975b;Graf & Giebisch, 1979). This consideration supports the idea of a carrier-mediated entry of Na across the mucosal membrane.…”

“…This means that the steady-state value for Er in K Ringer solution was about 10 mV more negative than Em (average Er = -18-6 + 1-6 mV). These changes in cell potentials in high K are well documented, and the current view is that they reflect the high K permeability of the cells (Reuss & Finn, 1975b;Van Os & Slegers, 1975;Graf & Giebish, 1979). The decrease in aca after K substitution followed an exponential time course and the over-all transient could be described by a single rate constant.…”