丙泊酚抑制2型糖尿病大鼠颈动脉窦压力感受性反射的机制:疑核含GluR2亚基的AMPA受体

Mechanism of propofol inhibiting carotid sinus baroreflex in rats with type 2 diabetes mellitus:GluR2 subunit-containing AMPA receptors in nucleus ambiguus

目的评价丙泊酚抑制2型糖尿病(T2DM)大鼠颈动脉窦压力感受性反射(CSR)的机制与疑核含GluR2亚基的AMPA受体的关系。方法选取SPF级健康雄性SD大鼠,3周龄,高糖高脂喂养6周后腹腔注射链脲佐菌素30 mg/kg制备T2DM大鼠模型。取T2DM造模成功的大鼠24只,采用随机数字表法分为4组(n=6):糖尿病-生理盐水组(DN组)、糖尿病-丙泊酚组(DP组)、AMPA受体激动剂-生理盐水组(AN组)和AMPA受体激动剂-丙泊酚组(AP组)。另取正常大鼠12只,采用随机数字表法分为2组(n=6):正常-生理盐水组(NN组)和正常-丙泊酚组(NP组)。AN组和AP组于颈动脉窦隔离灌流前30 min,利用微量进液器向疑核内微量注射AMPA受体激动剂mibampator 1 nmol/L(50 nl)。NP组、DP组、AP组经股静脉泵注丙泊酚45 mg·kg^(-1)·h^(-1),输注时间2 h,其余组静脉泵注等容量生理盐水。于丙泊酚或生理盐水泵注结束后20 min时建立颈动脉窦隔离灌流模型,绘制窦内压(ISP)-MAP曲线,记录CSR参数:曲线最大斜率(PS)、阈压(TP)、饱和压(SP)、平衡压(EP)、MAP反射性下降最大值(RD)和颈动脉窦压力感受器工作范围(OR)。灌流结束后,取脑组织,采用Western blot法和免疫荧光法测定疑核GluR2亚基的表达水平。结果与相应的生理盐水组(NN组、DN组、AN组)相比,各丙泊酚组(NP组、DP组、AP组)PS和RD降低,TP、SP和OR升高(P<0.05),ISP-MAP曲线上移,NP组和DP组疑核GluR2亚基表达下调(P<0.05),AP组疑核GluR2亚基表达差异无统计学意义(P>0.05)。与NP组相比,DP组PS和RD降低,TP、SP和OR升高(P<0.05),ISP-MAP曲线上移,疑核GluR2亚基表达下调(P<0.05);与DP组相比,AP组PS和RD升高,TP、SP和OR降低(P<0.05),ISP-MAP曲线下移,疑核GluR2亚基表达上调(P<0.05)。结论丙泊酚抑制T2DM大鼠CSR的机制可能与疑核含GluR2亚基的AMPA受体表达下调有关。

Objective To evaluate the relationship between the mechanism of propofol inhibiting carotid sinus baroreflex(CSR)and GluR2 subunit-containing AMPA receptors in the nucleus ambiguus of rats with type 2 diabetes mellitus(T2DM).Methods SPF healthy male Sprague-Dawley rats,aged 3 weeks,were selected and fed a high glucose and high fat diet for 6 weeks,and then streptozotocin 30 mg/kg was intraperitoneally injected to prepare a T2DM model of rats.Twenty-four T2DM rats were divided into 4 groups(n=6 each)using a random number table method:diabetes mellitus-normal saline group(DN group),diabetes mellitus-propofol group(DP group),AMPA receptor agonist-normal saline group(AN group),and AMPA receptor agonist-propofol group(AP group).Another 12 normal rats were selected and divided into 2 groups(n=6 each)using a random number table method:normal-normal saline group(NN group)and normal-propofol group(NP group).AMPA receptor agonist mibamitor 1 nmol/L(50 nl)was injected into the nucleus ambiguus using a micropipette at 30 min before perfusion of isolated carotid sinus in AN and AP groups.Propofol 45 mg·kg^(-1)·h^(-1)was infused for 2 h via the femoral vein in NP group,DP group and AP group,and the equal volume of normal saline was given instead in the other groups.A model for perfusing isolated carotid sinus was developed at 20 min after infusion of propofol or normal saline,the intracarotid sinus pressure(ISP)-mean arterial blood pressure(MAP)curve was drawn,and CSR parameters such as maximum slope(PS),threshold pressure(TP),saturation pressure(SP),equilibrium pressure(EP),maximum decrease in MAP reflexivity(RD),and carotid sinus baroreceptor operating range(OR)were recorded.Brain tissues were taken at the end of perfusion,and the expression of GluR2 subunit in the nucleus ambiguus was detected by Western blot and immunofluorescence.Results Compared with the corresponding normal saline groups(NN group,DN group,AN group),PS and RD were significantly decreased,TP,SP and OR were increased(P<0.05),and the ISP-MAP curve was shifted upward in propofol groups(NP group,DP group,AP group),the expression of GluR2 subunit in the nucleus ambiguus was down-regulated in NP and DP groups(P<0.05),and no significant change was found in the expression of GluR2 subunit in the nucleus ambiguus in AP group(P>0.05).Compared with NP group,PS and RD were significantly decreased,TP,SP and OR were increased(P<0.05),the ISP-MAP curve was shifted upward,and the expression of GluR2 subunit in the nucleus ambiguus was down-regulated in DP group(P<0.05).Compared with DP group,PS and RD were significantly increased,TP,SP and OR were decreased(P<0.05),the ISP-MAP curve was shifted downward,and the expression of GluR2 subunit in the nucleus ambiguus was up-regulated in AP group(P<0.05).Conclusions The mechanism by which propofol inhibits CSR may be related to down-regulation of the expression of GluR2 subunits-containing AMPA receptors in the nucleus ambiguus of rats with T2DM.