한양대학교 Nano Bio Tech Lab.

Abstract

We investigated the effect of calmodulin (CaM) and myosin light chain kinase (MLCK) on murine ileal myocytes using the whole-cell patch-clamp technique. Under the voltage clamp, at the holding potential of –60 mV, 50 μmol/L carbachol (CCh) induced inward currents (ICCh), and spontaneous decay of ICCh occurred. The peak inward currents induced by the repetitive application of CCh (50 μmol/L) tended to decrease in amplitude. Intracellular application of 0.2 mmol/L guanosine 5′-O-(γ-thio)triphosphate (GTPγS) from the patch electrode induced an inward current at a holding potential of –60mV, and the peak inward currents induced by the repetitive application of Cs tended to decrease slightly in amplitude. The amplitude of ICCh was reduced by pretreatment either with W-7, trifluoroperazine, W-5, and melittin (CaM inhibitors) or with ML-7 and ML-9 (selective MLCK inhibitors), and the inhibitory effects were reversible. However, when we pretreated with 50 μmol/L W-7 or 5 μmol/L ML-7 on GTPγS-induced inward currents, almost no inhibition was observed in the inward currents. Application of both Rho kinase inhibitor and MLCK inhibitor inhibited GTPγS-induced currents. We conclude that CaM and MLCK modulate the activation process of ICCh in murine ileal myocytes and suggest that the classical type transient receptor potential (TRPC) channel 5 might be a candidate for nonselective cationic currents (NSCC) activated by muscarinic stimulation in gastrointestinal smooth muscle cells.