盐酸戊乙奎醚对吗啡依赖大鼠相关脑区p-CREB及M5受体的影响

目的 观察盐酸戊乙奎醚(PHC)对吗啡依赖大鼠相关脑区p-CREB及M5受体的影响.方法 将雄性SD大鼠30只随机分为对照组(NS组)、吗啡依赖组(MOR组)和PHC治疗组(PHC组),每组10只.连续7d交替皮下注射吗啡(10 mg/kg,每天1次)或生理盐水,诱导大鼠的吗啡依赖位置偏爱效应.实验第8天停用吗啡,NS组与MOR组腹腔注射等体积的生理盐水；PHC治疗组则腹腔注射PHC 1.5 mg/kg.30 min后各组大鼠行CPP测试.Western blot法检测大鼠中脑腹侧被盖区、伏隔核、前额叶皮层p-CREB的蛋白表达；实时定量聚合酶链反应(PCR)检测大鼠中脑腹侧被盖区、伏隔核、前额叶皮层M5受体的mRNA水平.结果 (1)PHC治疗组的灰区停留时间与MOR组比较显著缩短[(422±103)s 比(622 ±97)s,P＜0.01].(2)MOR组中脑腹侧被盖区、伏隔核、前额叶皮层组织中p-CREB水平显著高于NS组[(0.93±0.06)、(0.67±0.10)、(0.89±0.14)比(0.31±0.02)、(0.20±0.01)、(0.40±0.07),P＜0.01]；与MOR组比较,PHC组中脑腹侧被盖区、伏隔核、前额叶皮层组织中p-CREB水平显著降低[(0.65±0.05)、(0.58±0.04)、(0.67±0.09),P＜0.05或P＜0.01].(3)MOR组中脑腹侧被盖区、伏隔核、前额叶皮层组织中M5受体的mRNA含量较NS组显著增高(1.80、0.22、0.50比1.00、0.13、0.32,P＜0.01)；与MOR组比较,PHC 组能明显降低中脑腹侧被盖区、伏隔核、前额叶皮层组织中M5的mRNA含量(0.65、0.09、0.07,P＜0.01).结论 PHC能抑制吗啡依赖大鼠条件性位置偏爱的表达,该效应可能与PHC下调中脑腹侧被盖区、伏隔核、前额叶皮层组织中M5受体,抑制p-CREB表达有关.

Effect of penehyclidine hydrochloride on expression of p-CREB and M5 receptor of the relevant brain regions in morphine dependent rats

Objective To explore the effect of penehyclidine hydrochloride (PHC) on the expression of phosphorylation of cyclic AMP response element binding protein (p-CREB) and M5 receptor of the relevant brain regions in morphine dependent rats.Methods Thirty male SD rats were randomly assigned to 3 groups ( n =10 each):morphine dependent group ( MOR group),PHC treatment groups ( PHC group),normal saline control group (NS group).The morphine conditioned place preference was induced by alternate subcutaneous injection of morphine for 7 days in rats (10 mg/kg,once daily,09:00) and saline (16:00).At the day 8,the rats received the CPP test.The rats in PHC group were treated with PHC (1.5 mg/kg,ip) prior to the CPP test,whereas the rats were treated with saline in MOR and NS group.Western blotting was used to detect the expression of p-CREB in the relevant brain regions ( including ventral tegmental area,nucleus accumbens,prefrontal cortex).Real-time polymerase chain reaction (PCR)was used to detect the expression of M5 receptor mRNA in the relevant brain regions ( including ventral tegmental area,nucleus accumbens,prefrontal cortex).Results ( 1 ) There were significant differences in the duration spent in the drug-paired side (gray area) between MOR and PHC groups [ (402 ± 103) vs.(622 ±97) s,P＜0.01] ; (2) In MOR group,the p-CREB levels in ventral tegmental area,nucleus accumbens and prefrontal cortex were significantly higher than those in NS group [ (0.93 ±0.06),(0.67 ±0.10),(0.89±0.14) vs.(0.31 ±0.02),(0.20±0.01),(0.40±0.07),P＜0.01].Acute treatment with PHC led to an obvious decrease of p-CREB level in ventral tegmental area,nucleus accumbens and prefrontal cortex [(0.65 ±0.05),(0.58 ±0.04),(0.67 ±0.09),P ＜0.05 or P＜0.01 ]; (3) As compared with NS group,the level of M5 receptor mRNA in ventral tegmental area,nucleus accumbens and prefrontal cortex was increased significantly during morphine dependence ( 1.80,0.22,0.50 vs.1.00,0.13,0.32,P ＜0.01 ).Treatment with PHC could significantly decrease the level of M5 receptor mRNA in ventral tegmental area,nucleus accumbens and prefrontal cortex (0.65,0.09,0.07,P＜0.01).Conclusion PHC could significantly inhibit the expression of CPP on morphine dependent rats,and this effect might be related to inhibition of the expression of p-CREB and down-regulation of M5 receptor in ventral tegmental area,nucleus accumbens and prefrontal cortex.