小檗碱通过阿片类δ受体抑制小鼠结肠动力

Mechanism of berberine inhibiting colonic motility through delta-opioid receptor

目的探讨小檗碱抑制结肠动力的作用机制。方法选取体重为20~25g的雄性C57BL小鼠。肠道平滑肌组织的分离和分离肠灌流槽实验中,当小檗碱和高选择性δ受体激动剂DeltorpinⅡ浓度为1×10^(-10)~1×10^(-6) mol/L时,观察肠道组织的收缩情况。应用非选择性阿片类受体拮抗剂纳络酮(1×10^(-6) mol/L)、δ受体拮抗剂纳曲吲哚(1×10^(-6) mol/L)、κ受体拮抗剂nor-BNI(1×10^(-6) mol/L)和μ受体拮抗剂β-FNA(1×10^(-6) mol/L),检测其拮抗小檗碱抑制肠道动力的作用,取每次实验的最后3次收缩值的平均值进行分析。结肠玻璃珠推排实验中,于小鼠腹腔内分别注射小檗碱1、3、5mg/kg,记录玻璃珠排出时间。于小檗碱(3 mg/kg)注射前15 min,分别于小鼠腹腔内注射纳络酮(1 mg/kg)和δ受体拮抗剂纳曲吲哚(1mg/kg),检测小檗碱抑制结肠动力的作用变化。小鼠腹腔内分别注射小檗碱(2和5mg/kg),采用免疫组织化学方法检测小鼠回肠和结肠中δ受体表达量。结果随着小檗碱和高选择性δ受体激动剂DeltorpinⅡ浓度的依次升高,结肠组织的收缩值依次减弱;小檗碱组1×10^(-6) mol/L时的结肠组织收缩值显著低于同组1×10^(-10) mol/L时(P<0.01),DeltorpinⅡ组1×10^(-7) mol/L、1×10^(-6) mol/L时的结肠组织收缩值均显著低于同组1×10^(-10) mol/L时。小檗碱浓度为1×10^(-7)、1×10^(-6) mol/L时,小檗碱+纳络酮组、小檗碱+纳曲吲哚组结肠组织的收缩值均显著高于小檗碱组相同浓度时(P值分别<0.01、0.05)。小檗碱3 mg/kg组注射后15min和小檗碱5mg/kg组注射后15、45min的结肠玻璃珠排出时间均显著长于空白对照组同时间(P值分别<0.01、0.05),小檗碱5mg/kg组注射后15、45min的结肠玻璃珠排出时间均显著长于于小檗碱1mg/kg组和小檗碱3mg/kg组同时间(P值均<0.01)。注射小檗碱后15和45min,小檗碱+DMSO组的结肠玻璃珠排出时间显著长于小檗碱+纳络酮组和小檗碱+纳曲吲哚组(P值分别<0.01、0.05)。檗碱2mg/kg组和小檗碱5mg/kg组小鼠回肠中δ受体表达量均显著高于空白对照组(P值均<0.01),小檗碱5mg/kg组小鼠结肠中δ受体表达量显著高于空白对照组(P<0.05)。结论小檗碱能通过阿片类δ受体抑制小鼠结肠动力。

Objective To elucidate the mechanism of berberine in inhibiting colonic motility. Methods C57BL mice weighing 20- 25 g were collected in this study. Berberine and 6-opioid receptor agonist Deltorpin Ⅱ（both 1×10^10 - 1×10^-6 mol/L） were added cumulatively into the organ baths and intestinal structure contractions were recorded. In separate experiments, nonselective opioid receptor antagonist naloxone （1×10^-6 mol/L）, δ-opioid receptor antagonist naltrindole （1 ×10^-6mol/L）, κ-opioid receptor antagonist nor-BNI （1 ×10^-6mol/L） and μ-opioid receptor antagonist β - FNA （1×10^-6mol/L） were added. The mean of the last 3 successive contractions of every concentration was analyzed. Berberine （ 1, 3 and 5 mg/kg, respectively） was injected intraperitoneally in mice. The time of bead expulsion was recorded. Then Naloxone （1 mg/kg） and naltrindole （1 mg/kg） were administered intraperitoneally 15 rain prior to berberine （3 mg/kg） injection. Berberine （2 and 5 mg/kg, respectively） was intraperitoneally injected, and δ-opioid receptor expression in ileum and Colon was tested by immunohistochemstry. Results With Berberine and Deltorpin Ⅱ concentrations increasing, colon contraction became weak gradually. Colon contraction after 1 ×10^-6 mol/Lberberine exposure was significantly weaker than that after 1×10^-6 mol/L berberine exposure （P〈 0. 01 ）. Colon contraction after 1 ×10^-7mol/L or 1 ×10^-6mol/L Deltorpin Ⅱ exposure was significantly weaker than that after 1 ×10^-6 mol/L Deltorpin Ⅱ exposure. When the concentration of berberine was 1 ×10^-7 mol/L or 1 ×10^-6 mol/L, conlon contraction in the mice given berberine＋ naloxone and those given berberine＋ naltrindole was significantly stronger than that in these given berberine only （P〈0.01, 0.05）. The time of colonic bead expulsion was significantly prolonged 15 min after 3 mg/kg berberine was injected, 15 and 45 min after 5 mg/kg berberine were injected as compared with black control group （P〈0.01, 0.05）. The time of colonic bead expulsion 15 and 45 min after 5 mg/kg berberine exposure were significantly longer than that 1 mg/kg and 3 mg/kg berberine exposure （all P〈0. 01 ）. The time of colonic bead expulsion in berberine＋ DMSO group was significantly longer than that in blank control group, berberine＋ naloxone group, and berberine＋ naltrindole group 15 and 45 min after berberine injection （all P〈0. 01）. The expression of 6-opioid receptor in ileum and colon of mice given 2 and 5 mg/kg berberine was significantly higher than that in the blank control group （both P〈0. 01）. The expression of δ-opioid receptor in colon of mice given 5 mg/kg berberine was also significantly higher than that in the blank control group （P〈0.05）. Conclusion Berberine can inhibit colonic motility in mice through δ-opioid receptor.