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目的:观察低渗刺激大鼠后,视上核(SON)内星形胶质细胞调节神经元活动的机制。方法:大鼠分为4组:(1)等渗对照组,经尾静脉注入生理盐水;(2)低渗刺激组,经尾静脉注入低渗盐水(0.83%葡萄糖+0.3%NaCl);(3)氟代柠檬酸(胶质细胞代谢抑制剂,fluorocitrate,FCA)+低渗刺激组,经侧脑室注射FCA(1 nmol/μl/只)2 h后,再经尾静脉注入低渗盐水;(4)甘珀酸(缝隙连接通道阻断剂,carbennoxolon,CBX)+低渗刺激组,经侧脑室注射CBX(50μg/只,10μg/lμl)2 h后,再经尾静脉注入低渗盐水。各组动物刺激后90 min,常规固定、取材、下丘脑连续冠状切片。应用抗牛磺酸(taurine,Tau)、抗加压素(vasopressin,VP)、抗甘氨酸受体(glycine recep-tor,GlyR)、抗缝隙连接蛋白(connexin43,Cx43)的抗体进行标记,以及Fos/胶质原纤维酸性蛋白(GFAP)双标记的免疫荧光染色方法,观察其在SON神经元和星形胶质细胞内的表达。结果:与对照组相比,低渗刺激组大鼠,SON内星形胶质细胞的胞体增大、突起变粗;Tau,Cx43和GFAP的平均荧光强度(MFI)增强,神经元上的GlyR表达增强,但VP和Fos的表达减弱。FCA,而不是CBX,明显减少星形胶质细胞的Tau,GFAP,Cx43的表达。FCA和CBX均可抑制神经元上的VP、GlyR、Fos阳性反应。结论:低渗刺激激活SON内星形胶质细胞,被激活的星形胶质细胞经Cx43半通道释放牛磺酸而抑制神经元的活性,减少VP的释放。
Objective: To investigate the mechanism of neuronal activity in astrocytes of the supraoptic nucleus (SON) after hypotonic stimulation. Methods: The rats were divided into 4 groups: (1) control group of isotonicity, saline injection through caudal vein; (2) hypotonic stimulation group, hypertonic saline (0.83% glucose + 0.3% NaCl) 3) Fluorocitrate (FCA) + hypotonic stimulation group, after intracerebroventricular injection of FCA (1 nmol / μl / only) for 2 h, (4) carbenoxolone (CBX) + hypotonic stimulation group, intracerebroventricular injection of CBX (50μg / 10μg / lμl) for 2 h, and then through the tail vein infusion of hypotonic saline . Each group of animals 90 minutes after stimulation, routine fixation, drawing, continuous coronal section of the hypothalamus. Antibody labeled with taurine (Tau), vasopressin (VP), glycine recep-tor (GlyR) and connexin43 (Cx43) / Glial fibrillary acidic protein (GFAP) double-labeled immunofluorescence staining method to observe its expression in SON neurons and astrocytes. Results: Compared with the control group, the cell bodies of SON in astrocytes increased and the protrusions became thicker in the hypotonic group. The mean fluorescence intensity (MFI) of Tau, Cx43 and GFAP increased, while the GlyR The expression was enhanced, but the expression of VP and Fos was weakened. FCA, but not CBX, significantly reduced the expression of Tau, GFAP and Cx43 in astrocytes. Both FCA and CBX can inhibit VP, GlyR, Fos positive neurons. CONCLUSION: Hypotonic stimulation activates astrocytes in SON and activated astrocytes release taurine via Cx43 hemi-channel to inhibit the activity of neurons and reduce the release of VP.