Cl⁻ currents of unstimulated T84 intestinal epithelial cells studied by intracellular recording

Abstract

The ionic currents spontaneously present in T84 intestinal epithelial cells, a line of colonie carcinoma origin, have been studied using the whole-cell recording mode of the patch-clamp technique and the single-electrode voltage-clamp method. Patch-clamp experiments showed that nonstimulated T84 cells already possess large currents but that these tend to disappear during the course of the experiments, presumably through the dialysis of some essential cytoplasmic component against the micropipette solution. The main charge carrier in these experiments appears to be Cl⁻ as judged from ion replacement. Microelectrode impalement of T84 cells gave a membrane potential of around −30 mV, similar to the equilibrium potential for Cl⁻ estimated from previously published values for intracellular Cl⁻ concentration. Voltage-clamp experiments with a single microelectrode revealed three kinetically distinguishable current patterns; currents decaying during hyperpolarizing voltage pulses, currents slowly activating during hyperpolarizing pulses and time-independent currents. The appearance of these distinct kinetic patterns was not predictable from cell to cell, and was not dependent on extracellular Ca²⁺. Ionic replacement experiments suggest that the charge carrier was always Cl⁻, regardless of the kinetic pattern observed. No K⁺ currents appear to be present in the nonstimulated T84 cells. Exposure of T84 cells to the muscarinic agonist carbachol induced a shift in the membrane potential towards more negative values, consistent with an activation of a K⁺ conductance. Thus, we suggest that the resting membrane potential in T84 cells is determined by the distribution of Cl⁻. This might imply that activation of K⁺ conductance could by itself support secretion by T84 monolayers through tonically active Cl⁻ channels.