Interstitial Cells of Cajal (ICCs) were considered as the pacemaker cells in gastrointestinal(GI) tract.We isolated ICCs from the small intestine of Balb/C mouse and measured the membrane potential and ionic current after 1 or 2 day culture using whole-cell patch clamp technique.In current clamping mode (I =0), substance P(SP) induced membrane depolarization and reduced amplitude.The effect of SP on membrane potentials was blocked by Gd3+ or by pre-incubating cells with a competitive peptide or a TACR1 antagonist (L703606).Reducing the external concentration of Na+ from 135 mM to 5 mM abolished the generation of pacemaker potentials.Under this condition, SP did not produce membrane depolarization.To determine the role of G proteins in the membrane depolarization by SP, we locked G proteins in active or inactive states with non-hydrolysable analogues of GTP (GTPγS) or GDP (GDPβS), respectively, applied by means of patch pipettes.Surprisingly, SP still induced membrane depolarization of comparable sizes.We also experinented the effect of SP in single ICC.The effect of SP in single ICC was similar with that of SP in ICC clusters.The biophysical and pharmacological properties of the NALCN (Sodium Leak Channel) currents closely resemble those of the SP-activated cation channel currents studied in ICCs.Reverse-transcription polymerase chain reaction and immunohistochemistry all showed abundant and localized expression of NALCN mRNA and protein in ICCs.These results suggest that NALCN may be a possible candidate for the SP-activated cation channel in ICCs from murine small intestine.