Characteristics of chloride currents activated by noradrenaline in rabbit ear artery cells

Abstract

1. Responses to noradrenaline were studied in isolated rabbit ear artery cells with the nystatin method of whole-cell patch-clamp recording. With this technique it was possible to obtain reproducible responses to noradrenaline which was not possible with traditional whole-cell recording. 2. With NaCl as the major constituent of the bathing solution (potassium-free pipette and external solutions) the reversal potential (Er) of the noradrenaline-evoked current was about 0 mV. When external chloride was replaced by thiocyanate, iodide, nitrate and bromide, Er was shifted to more negative potentials which indicates that a chloride conductance increase contributes to the current activated by noradrenaline. 3. When sodium was substituted by Tris, N-methyl-D-glucamine, choline or barium, Er of the noradrenaline-evoked current did not alter. This result suggests that a cation conductance is not implicated in the response to noradrenaline recorded with the nystatin method of whole-cell recording. 4. The chloride current activated by noradrenaline was blocked by the selective alpha 1-adrenoceptor antagonist prazosin but was not affected by the alpha 2-adrenoceptor antagonist yohimbine. 5. When cells were exposed to zero calcium bathing solutions the amplitude of the current elicited by noradrenaline was unaffected when measured within 1-2 min in zero calcium conditions. Continued exposure to 0 Ca + 1 mM-EGTA solution reversibly abolished the chloride current to noradrenaline. 6. In the presence of caffeine, which releases Ca2+ from internal stores and itself induced an inward current (at a holding potential of -50 mV), noradrenaline did not elicit a current. These data suggest that the chloride current evoked by noradrenaline results from an increase in intracellular concentration of calcium derived from internal stores. 7. The chloride channel blocking agents 4,4'-diisothiocyanatostilbene-2,2'-disulphonic acid (DIDS; 0.5 mM) and furosemide (0.5 mM) produced partial reduction of the noradrenaline-evoked chloride current whereas 4-acetamido-4'-isothiocyanatostilbene-2,2'-disulphonic acid (SITS), anthracene-9-carboxylic acid (A-9-C) and picrotoxin were ineffective in concentrations of up to 0.5 mM. However DIDS and furosemide were non-selective blockers as both agents were more effective against the adenosine triphosphate-induced cation current.