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, [Ca2+]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 [Ca2+]i. The outward current response disappeared after the removal of K+ from pipette solutions, indicating that caffeine activates a Ca(2+)-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 [Ca2+]i. The reversal potential (Er) of this current was about 0 mV. 4. Substitution of Tris+ or choline+ for external Na+ did not alter the Er. When external Cl- was replaced by thiocyanate-, iodide- or glutamate-, the Er 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 Ca2+ transient was still observed for a few minutes after exposure of the cells to Ca(2+)-free external solution containing 2 mM EGTA. Caffeine failed to produce an inward current and Ca2+ 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.
Ohta
T
T
Department of Pharmacology, Faculty of Veterinary Medicine, Hokkaido University, Sapporo, Japan.
Ito
S
S
Nakazato
Y
Y
D000818:Animals; D002110:Caffeine / Q000494:pharmacology*; D002118:Calcium / Q000378:metabolism / Q000502:physiology; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D018118:Chloride Channels / Q000187:drug effects* / Q000378:metabolism; D002794:Choline / Q000494:pharmacology; D004533:Egtazic Acid / Q000494:pharmacology; D004594:Electrophysiology; D005453:Fluorescence; D016257:Fura-2; D066298:In Vitro Techniques; D007413:Intestinal Mucosa / Q000378:metabolism; D007422:Intestines / Q000187:drug effects; D008297:Male; D008564:Membrane Potentials / Q000187:drug effects; D009130:Muscle, Smooth / Q000187:drug effects / Q000378:metabolism*; D011189:Potassium Chloride / Q000494:pharmacology; D051381:Rats; D017208:Rats, Wistar; D012433:Ryanodine / Q000494:pharmacology
D018118:Chloride Channels; D012433:Ryanodine; D002110:Caffeine; D004533:Egtazic Acid; D011189:Potassium Chloride; D002794:Choline; D002118:Calcium; D016257:Fura-2