CHLORIDE CURRENTS ACTIVATED BY CAFFEINE IN RAT INTESTINAL SMOOTH-MUSCLE CELLS

1. Current responses to caffeine in single smooth muscle cells isolated from rat intestine were studied with the whole-cell patch clamp technique. Intracellular calcium concentration, [Ca 2+]i, was simultaneously monitored with fura-2 (0.1 mm) introduced into the cell through a patch pipette. 2. With a potassium-containing pipette solution, caffeine (10 mm) produced an outward current at a holding potential of 0 mV and an inward current at - 60 mV, both of which were accompanied by parallel increases in [Ca 2+]i. The outward current response disappeared after the removal of K+ from pipette solutions, indicating that caffeine activates a Ca2+-activated K+ conductance. 3. When NaCl was present in both pipette and external solutions as the major constituent, caffeine evoked an inward current at - 60 mV simultaneously with a rise in [Ca 2+]i. The reversal potential (E(r)) of this current was about 0 mV. 4. Substitution of Tris+ or choline+ for external Na+ did not alter the E(r). When external Cl- was replaced by thiocyanate-, iodide- or glutamate-, the E(r) changed to respectively - 55, - 38 and + 35 mV. 5. The current response to caffeine decreased with increasing concentration of EGTA in the pipette solution. The caffeine-induced current and the intracellular Ca 2+ transient was still observed for a few minutes after exposure of the cells to Ca2+-free external solution containing 2 mm EGTA. Caffeine failed to produce an inward current and Ca 2+ transient after treatment with extracellular ryanodine. 6. It is concluded that caffeine caused an increase in membrane Cl- conductance and in K+ conductance resulting from a rise in [Ca2+]i derived from ryanodine-sensitive intracellular Ca2+ stores in isolated smooth muscle cells of the rat intestine.