Hwangryunhaedok-tang induces the depolarization of pacemaker potentials through 5-HT(3) and 5-HT(4) receptors in cultured murine small intestine interstitial cells of Cajal

AIM: To investigate the effects of a water extract of Hwangryunhaedok-tang (HHTE) on the pacemaker potentials of mouse interstitial cells of Cajal (ICCs). METHODS: We dissociated ICCs from small intestines and cultured. ICCs were immunologically identified using an anti-c-kit antibody. We used the whole-cell patch-clamp configuration to record the pacemaker potentials generated by cultured ICCs under the current clamp mode (I = 0). All experiments were performed at 30 °C-32 °C RESULTS: HHTE dose-dependently depolarized ICC pacemaker potentials. Pretreatment with a 5-HT(3) receptor antagonist (Y25130) or a 5-HT(4) receptor antagonist (RS39604) blocked HHTE-induced pacemaker potential depolarizations, whereas pretreatment with a 5-HT(7) receptor antagonist (SB269970) did not. Intracellular GDPβS inhibited HHTE-induced pacemaker potential depolarization and pretreatment with a Ca(2+)-free solution or thapsigargin abolished the pacemaker potentials. In the presence of a Ca(2+)-free solution or thapsigargin, HHTE did not depolarize ICC pacemaker potentials. In addition, HHTE-induced pacemaker potential depolarization was unaffected by a PKC inhibitor (calphostin C) or a Rho kinase inhibitor (Y27632). Of the four ingredients of HHT, Coptidis Rhizoma and Gardeniae Fructus more effectively inhibited pacemaker potential depolarization. CONCLUSION: These results suggest that HHTE dose-dependently depolarizes ICC pacemaker potentials through 5-HT(3) and 5-HT(4) receptors via external and internal Ca(2+) regulation and via G protein-, PKC- and Rho kinase-independent pathways.