吗啡依赖小鼠十二指肠P2Y_2受体表达及其对十二指肠碳酸氢盐分泌的影响

The expression of P2Y_2 receptor and its effect on duodenum mucosal bicarbonate secretion in morphine dependent mice

目的研究吗啡依赖小鼠十二指肠嘌呤受体P2Y2表达及其对碳酸氢盐分泌的影响。方法小鼠吗啡6 mg/kg颈背部皮下注射,每日1次,连续8 d,建立小鼠吗啡条件性位置偏爱(conditioned place preference,CPP)模型。确认模型建立成功后处死取十二指肠(10只),Western-blot技术检测其P2Y2受体蛋白的表达;选取十二指肠(8只),剥离浆膜后装载于Ussing Chamber上,经ATP和Surmain+ATP分别刺激后,用p H-stat技术测定十二指肠碳酸氢盐的分泌量。结果 CPP模型组小鼠嘌呤受体P2Y2蛋白(0.626 7±0.046 3)在十二指肠中较生理盐水对照组(1.087 0±0.054 9)的表达明显降低(P=0.036 0,*P<0.05);用P2Y2受体特异性激动剂ATP刺激后,CPP模型组小鼠的碳酸根离子分泌(0.426 7±0.060 6)明显低于生理盐水对照组(0.726 7±0.038 4)(P=0.045 0,*P<0.05),Suramin能显著抑制ATP刺激生理盐水对照组(0.216 7±0.020 3)和模型组(0.196 7±0.008 8)的碳酸氢盐分泌。结论在吗啡依赖状态下,十二指肠嘌呤受体P2Y2蛋白的表达水平下调,并由此可介导十二指肠碳酸氢盐分泌的减少,这可能是阿片类依赖者胃肠功能失调的一个重要原因。

Objective To investigate the mechanism of P2Y2 function in duodenal musosal bicarbonate secretion （DMBS） in morphine dependent mice. Methods The conditioned place preference （CPP） model was established by injecting morphine in mouse （6 mg/kg neck subcutaneous） for 8 days. After the CPP model was estabolished, 10 mice were killed. The western - blot was applied to detect the expression of P2Y2 receptor in mouse duodenum between saline group and CPP group. For another 8 mice, the duodenal mucosa were stripped of seromuscular layers and mounted in Ussing chamber for DMBS measurement after application of ATP and Suramin. Results Compared to the saline group （ 1. 087 ± 0. 054 ）, the protein expression of P2Y2 in CPP group （0. 626 ± 0. 046） were significantly decreased （P = 0. 036 0, ＊ P 〈 0.05 ）. After treatment with ATP, a P2Y2 - receptor agonist, the DMBS in saline group （0. 726 ± 0. 038） was higher than the CPP group （0. 426 ± 0. 060） （P = 0. 045 0, ＊P 〈0.05）. And this response was completely abolished by Suramin （100 μmol/L） , a P2Y2 receptor special antagonist in saline group （0. 216 ± 0. 020） and CPP group （0. 196 ±0. 008）. Conclusion The protein expression of P2Y2 was down regulated, causing DMBS secretion decrease in morphine state - dependent. This is an important reason that the gastrointestinal disorders happened in opioid -dependent people.