Contractile effects and intracellular Ca2+ signalling induced by emodin in circular smooth muscle cells of rat colon

AIM: To investigate whether emodin has any effects on circular smooth muscle cells of rat colon and to examine the mechanism underlying its effect. METHODS: Smooth muscle cells were isolated from the circular muscle layer of Wistar rat colon and the cell length was measured by computerized image micrometry. Intracellular Ca2+ ([Ca2+](i)) signalling was studied in smooth muscle cells using Ca2+ indicator Fluo-3 AM on a laser-scanning confocal microscope. RESULTS: Emodin dose-dependently induced smooth muscle cells contraction. The contractile responses induced by emodin were inhibited by preincubation of the cells with ML-7 (an inhibitor of MLCK). Emodin caused a large, transient increase in [Ca2+](i) followed by a sustained elevation in [Ca2+](i). The emodin -induced increase in [Ca2+](i) was unaffected by nifedipine, a voltage-gated Ca2+-channel antagonist, and the sustained phase of the rising of [Ca2+](i) was attenuated by extracellular Ca2+ removal with EGTA solution. Inhibiting Ca2+ release from ryanodine-sensitive intracellular stores by ryanodine reduced the peak increase in [Ca2+](i). Using heparin, an antagonist of IP3R, almost abolished the peak increase in [Ca2+](i). CONCLUSION: Emodin has a direct excitatory effect on circular smooth muscle cells in rat colon mediated via Ca2+/CaM dependent pathways. Furthermore, emodin-induced peak [Ca2+](i) increase may be attributable to the Ca2+ release from IP3 sensitive stores, which further promote Ca2+ release from ryanodine-sensitive stores through CICR mechanism. Additionally, Ca2+ influx from extracellular medium contributes to the sustained increase in [Ca2+](i).