Modulation of Pacemaker Potentials by Pyungwi-San in Interstitial Cells of Cajal from Murine Small Intestine - Pyungwi-San and Interstitial Cells of Cajal -

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  • ABSTRACT

    Objective:

    Pyungwi-san (PWS) plays a role in a number of physiologic and pharmacologic functions in many organs. Interstitial cells of Cajal (ICCs) are pacemaker cells that generate slow waves in the gastrointestinal (GI) tract. We aimed to investigate the beneficial effects of PWS in mouse small-intestinal ICCs.

    Methods:

    Enzymatic digestion was used to dissociate ICCs from the small intestine of a mouse. The wholecell patch-clamp configuration was used to record membrane potentials from the cultured ICCs.

    Results:

    ICCs generated pacemaker potentials in the GI tract. PWS produced membrane depolarization in the current clamp mode. Pretreatment with a Ca2+-free solution and a thapsigargin, a Ca2+-ATPase, inhibitor in the endoplasmic reticulum, eliminated the generation of pacemaker potentials. However, only when the thapsigargin was applied in a bath solution, the membrane depolarization was not produced by PWS. Furthermore, the membrane depolarizations due to PWS were inhibited not by U-73122, an active phospholipase C inhibitor, but by chelerythrine and calphostin C, protein kinase C inhibitors.

    Conclusions:

    These results suggest that PWS might affect GI motility by modulating the pacemaker activity in the ICCs.


  • KEYWORD

    interstitial cells of Cajal , pacemaker cell , Pyungwi-san , traditional herbal medicine , gastrointestinal tract

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  • [Table 1] Amount and composition of PWS
    Amount and composition of PWS
  • [Figure 1] Effects of PWS on pacemaker potentials in cultured ICCs from murine small intestine. A-D: ICCs were isolated from the Balb/c small intestine after primary cell culture for various doses of PSW (5 to 50 mg/mL) and the clamping mode (I = 0). Summary of the responses to PWS in E. The error bars represent mean values ± standard error (SE). **(P < 0.01) Significantly different from the untreated control. CTRL: Control.
    Effects of PWS on pacemaker potentials in cultured ICCs from murine small intestine. A-D: ICCs were isolated from the Balb/c small intestine after primary cell culture for various doses of PSW (5 to 50 mg/mL) and the clamping mode (I = 0). Summary of the responses to PWS in E. The error bars represent mean values ± standard error (SE). **(P < 0.01) Significantly different from the untreated control. CTRL: Control.
  • [Figure 2] Effects of pretreatment with an external Ca2+-free solution or thapsigargin, a Ca2+-ATPase inhibitor, in the endoplasmic reticulum and of U-73122, an active phospholipase C inhibitor, on PWS-induced pacemaker potentials in cultured ICCs. A: ICCs were isolated from the Balb/c small intestine after primary cell culture for various doses of PSW (5 to 50 mg/mL) and the clamping mode (I = 0). The external Ca2+-free solution eliminated the generation of pacemaker potentials. Under these conditions, PWS-induced (30 mg/ml) membrane depolarizations were produced. B: Thapsigargin (5 ㎛) eliminated the generation of pacemaker potentials. Thapsigargin blocked the PWSinduced (30 mg/ml) membrane depolarizations. C: Summary of the responses to PWS in the presence of an external Ca2+-free solution and thapsigargin. Bars represent the mean values ± SE. **(P < 0.01) Significantly different from the untreated control. CTRL: Control.
    Effects of pretreatment with an external Ca2+-free solution or thapsigargin, a Ca2+-ATPase inhibitor, in the endoplasmic reticulum and of U-73122, an active phospholipase C inhibitor, on PWS-induced pacemaker potentials in cultured ICCs. A: ICCs were isolated from the Balb/c small intestine after primary cell culture for various doses of PSW (5 to 50 mg/mL) and the clamping mode (I = 0). The external Ca2+-free solution eliminated the generation of pacemaker potentials. Under these conditions, PWS-induced (30 mg/ml) membrane depolarizations were produced. B: Thapsigargin (5 ㎛) eliminated the generation of pacemaker potentials. Thapsigargin blocked the PWSinduced (30 mg/ml) membrane depolarizations. C: Summary of the responses to PWS in the presence of an external Ca2+-free solution and thapsigargin. Bars represent the mean values ± SE. **(P < 0.01) Significantly different from the untreated control. CTRL: Control.
  • [Figure 3] Effects of PWS on phospholipase C inhibitors in cultured ICCs. A: ICCs were isolated from the Balb/c small intestine after primary cell culture for various doses of PSW (5 to 50 mg/mL) and the clamping mode (I = 0). U-73122 (5 ㎛), a phospholipase C inhibitor, eliminated the generation of pacemaker potentials. U-73122 did not block the PWS-induced (30 mg/ml) membrane depolarizations. B: The application of U-73343 (5 ㎛) did not influence the generation of pacemaker currents. Also, U-73343 did not block the PWS-induced (30 mg/ml) membrane depolarizations. C: Summary of the responses to PWS in phospholipase C inhibitors. Bars represent the mean values ± SE. CTRL: Control.
    Effects of PWS on phospholipase C inhibitors in cultured ICCs. A: ICCs were isolated from the Balb/c small intestine after primary cell culture for various doses of PSW (5 to 50 mg/mL) and the clamping mode (I = 0). U-73122 (5 ㎛), a phospholipase C inhibitor, eliminated the generation of pacemaker potentials. U-73122 did not block the PWS-induced (30 mg/ml) membrane depolarizations. B: The application of U-73343 (5 ㎛) did not influence the generation of pacemaker currents. Also, U-73343 did not block the PWS-induced (30 mg/ml) membrane depolarizations. C: Summary of the responses to PWS in phospholipase C inhibitors. Bars represent the mean values ± SE. CTRL: Control.
  • [Figure 4] Effects of chelerythrine or calphostin C, an inhibitor of protein kinase C, on PWS-induced pacemaker potentials in cultured ICCs of the murine small intestine. A, B: ICCs were isolated from the Balb/c small intestine after primary cell culture for various doses of PSW (5 to 50 mg/mL) and the clamping mode (I = 0). Pacemaker potentials of ICCs exposed to PWS (30 mg/ml) in the presence of chelerythrine (1 ㎛) or calphostin C (10 ㎛) are shown. Under these conditions, PWS did not cause membrane depolarizations. C: Summary of the responses to PWS in the presence of chelerythrine or calphostin C. Bars represent the mean values ± SE **(P < 0.01) Significantly different from the untreated control. CTRL: Control.
    Effects of chelerythrine or calphostin C, an inhibitor of protein kinase C, on PWS-induced pacemaker potentials in cultured ICCs of the murine small intestine. A, B: ICCs were isolated from the Balb/c small intestine after primary cell culture for various doses of PSW (5 to 50 mg/mL) and the clamping mode (I = 0). Pacemaker potentials of ICCs exposed to PWS (30 mg/ml) in the presence of chelerythrine (1 ㎛) or calphostin C (10 ㎛) are shown. Under these conditions, PWS did not cause membrane depolarizations. C: Summary of the responses to PWS in the presence of chelerythrine or calphostin C. Bars represent the mean values ± SE **(P < 0.01) Significantly different from the untreated control. CTRL: Control.