Effects of Lizhong Tang on cultured mouse small intestine interstitial cells of Cajal

Abstract

To investigate the effects of Lizhong Tang, an herbal product used in traditional Chinese medicine, on mouse small intestine interstitial cells of Cajal (ICCs). Enzymatic digestions were used to dissociate ICCs from mouse small intestine tissues. The ICCs were morphologically distinct from other cell types in culture and were identified using phase contrast microscopy after verification with anti c-kit antibody. A whole-cell patch-clamp configuration was used to record potentials (current clamp) from cultured ICCs. All of the experiments were performed at 30-32 °C. ICCs generated pacemaker potentials, and Lizhong Tang produced membrane depolarization in current-clamp mode. The application of flufenamic acid (a nonselective cation channel blocker) abolished the generation of pacemaker potentials by Lizhong Tang. Pretreatment with thapsigargin (a Ca²⁺-ATPase inhibitor in the endoplasmic reticulum) also abolished the generation of pacemaker potentials by Lizhong Tang. However, pacemaker potentials were completely abolished in the presence of an external Ca²⁺-free solution, and under this condition, Lizhong Tang induced membrane depolarizations. Furthermore, When GDP-β-S (1 mmol/L) was in the pipette solution, Lizhong Tang still induced membrane depolarizations. In addition, membrane depolarizations were not inhibited by chelerythrine or calphostin C, which are protein kinase C inhibitors, but were inhibited by U-73122, an active phospholipase C inhibitors. These results suggest that Lizhong Tang might affect gastrointestinal motility by modulating pacemaker activity in interstitial cells of Cajal.