Inward current activated by carbachol in rat intestinal smooth muscle cells.
1. Carbachol (0.1 mM or 10 microM)-evoked inward currents were studied with standard and perforated whole-cell patch clamp techniques in smooth muscle cells isolated from rat small intestine. The intracellular free Ca2+ concentration was monitored simultaneously with the fura-2 method. 2. With a K(+)-containing pipette solution, carbachol produced an inward current at -60 mV and a large outward current at -20 mV. 3. When NaCl was substituted for KCl in the external and pipette solutions, carbachol elicited inward currents at holding potentials more inside-negative than 0 mV. The reversal potential of the carbachol-induced current altered when external chloride (-0.9 mV) was replaced by iodide (-21.2 mV), thiocyanate (-27.0 mV) and glutamate (18.2 mV). The carbachol-induced current at -60 mV was slightly decreased by the replacement of external NaCl with Tris-Cl. 4. The carbachol-induced inward current at -60 mV was accompanied by an increase in the intracellular concentration of free Ca2+. Both responses to carbachol were observed 2 min after exposure of the cells to a Ca(2+)-free solution containing 2 mM EGTA. 5. Intracellular application of heparin inhibited the inward current and Ca2+ transient responses to carbachol but not those to caffeine (10 mM). An inward current and Ca2+ transient were elicited after the patch membrane was ruptured at -60 mV, using a patch pipette containing inositol 1,4,5-trisphosphate (InsP3). 6. It is concluded that the carbachol-induced inward current is due to increases in membrane Cl- and Na+ conductances. Ca2+ released from InsP3-sensitive stores may play a role in increasing both conductances.
Ito
S
S
Department of Pharmacology, Faculty of Veterinary Medicine, Hokkaido, Japan.
Ohta
T
T
Nakazato
Y
Y
D000818:Animals; D002110:Caffeine / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism; D002217:Carbachol / Q000494:pharmacology*; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D004594:Electrophysiology; D016257:Fura-2; D006493:Heparin / Q000494:pharmacology; D007082:Ileum / Q000166:cytology / Q000187:drug effects; D066298:In Vitro Techniques; D015544:Inositol 1,4,5-Trisphosphate / Q000494:pharmacology; D007473:Ion Channels / Q000187:drug effects*; D008297:Male; D008564:Membrane Potentials / Q000187:drug effects; D009130:Muscle, Smooth / Q000166:cytology / Q000187:drug effects*; D051381:Rats; D017208:Rats, Wistar; D011976:Receptors, Muscarinic / Q000187:drug effects
D007473:Ion Channels; D011976:Receptors, Muscarinic; D002110:Caffeine; D015544:Inositol 1,4,5-Trisphosphate; D002217:Carbachol; D006493:Heparin; D002118:Calcium; D016257:Fura-2