甘珀酸对内脏痛大鼠延髓星形胶质细胞和神经元反应的影响

目的:研究侧脑室注射甘珀酸后对福尔马林灌胃致内脏疼痛大鼠的延髓迷走孤束复合体内星形胶质细胞和神经元反应的影响.方法:经侧脑室注射缝隙连接阻断剂甘珀酸(carbenoxolone,CBX)后向大鼠胃内灌入2.5%福尔马林2ml诱发内脏疼痛,用免疫组织化学方法观察延髓迷走孤束复合体(VSC)内抗Fos蛋白(标记神经元)和抗胶质原纤维酸性蛋白(标记星形胶质细胞)的单一或双重标记的免疫荧光染色.结果:福尔马林灌胃后大鼠出现烦躁易激惹,呼吸变快,持续1h;而预先侧脑室注射CBX则动物疼痛行为学反应明显减轻.免疫组织化学染色发现福尔马林灌胃后大鼠VSC中的Fos免疫反应数目增强;大鼠预先侧脑室注射CBX后VSC中的Fos免疫反应数目明显减弱.结论:延髓VSC中的星形胶质细胞和神经元参与福尔马林灌胃致内脏痛的调节,星形胶质细胞可能通过缝隙连接影响神经元对内脏痛的调节功能.     

星形胶质细胞糖代谢对神经元的支持及保护作用

脑组织有着极其复杂的功能,这些功能的完成有赖于神经元细胞与胶质细胞之间的广泛合作。星形胶质细胞作为人脑内数量最多的细胞,其与神经元细胞之间的相互作用就显得十分重要。葡萄糖代谢途径包括糖酵解,有氧氧化及磷酸戊糖三条途径。其为脑组织维持其正常功能的前提。研究表明星形胶质细胞和神经元在糖代谢方面有着各自的特点,神经元在能量底物及抗氧化应激中对星形胶质细胞糖代谢途径存在一定的依赖性,干扰星形胶质细胞与神经元之间的代谢过程会导致疾病的发生。本综述主要从糖酵解及磷酸戊糖两条糖代谢途径阐述了星形胶质细胞与神经元的关系。这或许会对研究脑的代谢,脑疾病中神经元的损伤机制及如何保护神经元提供全新的视角,并可能为一些疾病的治疗开辟了新的途径。     

胃伤害性刺激诱导大鼠脑干星形胶质细胞GFAP蛋白表达及其与神经元的关系

研究大鼠在福尔马林诱发胃伤害性刺激时脑干内星形胶质细胞及神经元的变化。应用免疫组织化学三重标记法在脑原位切片同时显示脑干内Fos蛋白,胶质原纤维酸性蛋白（GFAP）,酪氨酸羟化酶（TH）的表达,结果显示：1、在福尔马林诱发胃伤害性刺激后,脑干胶质细胞GFAP表达阳性,并表现出明显的核团或亚核定位特点,在延髓内脏带（MVZ0,中缝大核（RMg),蓝斑（LC）,臂旁外侧核（LPB）,中缝背核（DR）,中脑导水管周围灰质腹外侧区（vlPAG),上丘中灰层（IngSC)等脑区有较多的Fos阳性细胞,而且Fos阳性表达的分布与上述GFAP阳性分布基本一致;2、MVZ,LC,DR,vlPAG等部位有大量Fos及TH双标阳性神经元,周围有密集的GFAP阳性细胞;3、随着刺激后存活时间的变化,GFAP与Fos阳性细胞的反应均经历逐渐升高后又渐降低直至消失的变化。结果表明：上述核团的神经元和星形胶质细胞可能同时参与了内脏痛及其调节过程。     

成年大鼠神经元和星形胶质细胞细胞周期蛋白表达的差异研究

目的比较研究成年大鼠细胞周期蛋白在神经元和星形胶质细胞的表达差异。方法应用免疫荧光和激光扫描共聚焦显微镜观察成年大鼠生理状态下大脑皮层或海马CA1区神经元和星形胶质细胞细胞周期素D1、E、A、B1、(CyclinD1、E、A、B1)的表达。结果成年大鼠海马CA1区和大脑皮层的神经元有Cyclin D1、E、A和B1的表达,细胞核和细胞浆均有表达,以胞核为主;星形胶质细胞也有上述细胞周期蛋白的表达但细胞数目较少,并且表达这些指标的星形胶质细胞多聚集在海马CA1区。结论成年大鼠大脑皮层和海马区的神经元和星形胶质细胞均表达细胞周期蛋白,而其在神经元的表达较星形胶质细胞更为普遍。     

Ⅰ型糖尿病胃轻瘫大鼠动物模型建立与优化

目的探讨I型糖尿病胃轻瘫大鼠模型的建立方法,为糖尿病胃肠动力障碍研究提供合理的平台.方法链脲佐菌素(STZ)一次性腹腔注射方法建立I型糖尿病动物模型,观察大鼠一般状况、血糖、胃排空、小肠传输速率.结果 ①模型组大鼠于造模后均出现糖尿病症状;平均体重明显低于对照组(P＜0.01);平均血糖浓度显著高于对照组(P＜0.01).②模型组大鼠胃内色素残留率明显高于对照组(P＜0.01);小肠传输速率显著慢于对照组(P＜0.01).结论 链脲佐菌素一次性腹腔内注射10周后能成功诱导出糖尿病胃轻瘫大鼠模型.     

莫沙必利对糖尿病胃轻瘫大鼠胃窦Cajal间质细胞的影响

目的:研究莫沙必利对糖尿病胃轻瘫大鼠胃窦Cajal间质细胞(ICC)的影响。方法:将48只SD雄性大鼠随机分为正常组、模型组、莫沙必利组,各组分别16只。模型组、莫沙必利组大鼠给予一次性腹腔注射链脲佐菌素后建立糖尿病胃轻瘫模型,建模第10天起,莫沙必利组大鼠给予莫沙必利注灌胃治疗。建模30周后,检测各组大鼠的血糖水平、胃残留率,通过采用电生理学方法检测各组大鼠胃动力,并采用免疫组化染色方法检测各组胃窦ICC的相对数量。结果:模型组大鼠胃残留率明显高于正常组大鼠(P0.05),而给予莫沙必利后胃残留率显著降低(P0.05);模型组大鼠胃电图波幅和频率均明显低于正常组大鼠(P0.05),而给予莫沙必利后胃电图波幅和频率明显增加(P0.05);模型组大鼠胃窦ICC明显低于正常组大鼠(P0.05),而给予莫沙必利后胃窦ICC有所增加(P0.05)。结论:莫沙必利可显著改善糖尿病胃轻瘫大鼠的胃动力,可能与其增加胃窦Cajal间质细胞有关。     

电针足三里穴对胃电的影响

目的:探讨电针大鼠足三里穴对胃动力的调节.方法:将大鼠随机分成足三里组、非经非穴组、非针刺对照组.观察电针后胃电的变化.结果:选取胃动过缓分布＞1/3(Ⅰ)及胃动过速分布＞1/3(Ⅱ)的各组数据进行统计学分析,胃电图表明:①各组Ⅰ的例数均比Ⅱ的例数多;②在Ⅰ或Ⅱ中,足三里组比其余两组的例数多、平均频率分布率多;③在Ⅰ中,足三里组比其余两组的平均主频慢.结论:电针足三里穴对大鼠胃电具有双向调节作用,以抑制作用为主.     

大鼠脑缺血再灌注后神经元和星形胶质细胞凋亡的比较研究

目的比较研究大鼠局灶性脑缺血再灌注后神经元和星形胶质细胞的凋亡规律。方法建立大鼠大脑中动脉阻塞(middle cerebral artery occlusion,MCAO)再灌注模型,在缺血再灌注后1、3、7、14d断头取脑,应用流式细胞分选技术和原位末端标记法分别检测各组MCAO后不同时期神经元和星形胶质细胞凋亡情况。结果局灶性脑缺血再灌注后,海马区星形胶质细胞凋亡数量超过神经元,其凋亡以再灌注3d最为显著,而神经元则以7d最为显著;而皮层区神经元凋亡数量超过星形胶质细胞,两种细胞凋亡均在再灌注后7d达高峰。结论脑缺血再灌注后,皮层和海马区的神经元及星形胶质细胞均可发生凋亡,海马区星形胶质细胞比皮层区更易凋亡,而皮层区神经元比海马区更易凋亡。     

LPS诱导海马星形胶质细胞活化与活化的星形胶质细胞对海马神经元的影响

探讨脂多糖(Lipopolysaccharide,LPS)对长时间存活大鼠海马内星形胶质细胞的反应以及对神经元的影响。方法:本实验用10只健康成年雄性SD大鼠,海马CA3区注射LPS 10μ1．7和14d后,尼氏染色观察神经元的变化,免疫组织化学染色结合图像分析方法观察海马CA3区注射部位胶质纤维酸性蛋白(glial fibrillary acidic protein GFAP)、的表达变化。结果:脂多糖可促进海马星形胶质细胞的活化,但并不能引起海马区神经元的损伤。结论:星形胶质细胞在脑损伤后的脑内炎症反应起了一定的作用,但并不能引起神经元的损伤。     

糖尿病胃轻瘫患者的护理进展

正2型糖尿病并发上消化道动力异常并不少见,出现的胃症状较多,称之为糖尿病性胃病。当无幽门或十二指肠梗阻时,胃排空延长被称为胃轻瘫(Gastroparesis),其发生率占糖尿病患者的50%~76%,其中10%患者出现临床表现[1]。随着糖尿病发病率的增加,因糖尿病胃轻瘫(Diabetic Gastroparesis,DGP)住院的人数也显著增加[2],严重影响患者的     

miR-103a对癫痫大鼠海马组织星形胶质细胞活化的影响

为了考察miR-103a对癫痫大鼠海马组织星形胶质细胞活化的影响。本研究通过腹腔注射氯化锂和毛果芸香碱诱导癫痫大鼠模型,对大鼠脑室内注射miR-103a抑制剂来敲低miR-103a的表达;采用免疫组织化学染色检测大鼠海马组织中胶质纤维酸性蛋白(GFAP)的阳性表达;采用RT-qPCR和Western blotting方法检测大鼠海马组织中miR-103a、脑源性神经营养因子(BDNF)、GFAP、TNF-α和IL-6的m RNA和蛋白表达;苏木精-伊红(HE)染色评价海马组织病变程度;Nissl染色检测神经元存活情况;TUNEL染色检测神经元的凋亡。结果显示,癫痫大鼠海马组织中miR-103a被上调。下调miR-103a抑制癫痫大鼠海马组织中GFAP的mRNA和蛋白表达,且抑制癫痫大鼠海马神经元的病理损伤,但能促进癫痫大鼠海马神经元的存活并抑制其凋亡。此外,下调miR-103a还抑制癫痫大鼠海马组织中IL-6和TNF-α的表达,并促进癫痫大鼠海马组织中BDNF的表达。本研究表明,靶向沉默miR-103a可以抑制癫痫大鼠海马组织中星形胶质细胞的活化并改善神经元的病理损伤。     

Chronic Activation of the Cyclic AMP Signaling Pathway Promotes Development and Long-Term Survival of Mesencephalic Dopaminergic Neurons

Abstract: Dibutyryl cyclic AMP (dbcAMP), a permeant analogue of cyclic AMP (cAMP), prevented, for at least 3 weeks, the death of tyrosine hydroxylase (TH)-immunopositive dopaminergic neurons, which occurred spontaneously by apoptosis in mesencephalic cultures. Treatment with the cyclic nucleotide analogue also led to a significant increase in the uptake of [³H]dopamine, attesting that the rescued TH⁺ neurons were fully functional and differentiated. dbcAMP was most effective when added immediately after plating, but delayed treatment could still arrest the ongoing degenerative process. Trophic/survival effects were long-lasting, declining only progressively after withdrawal of dbcAMP from the culture medium. They were independent of cell density and still detectable in the absence of serum proteins. The effects of dbcAMP were mimicked by depolarizing concentrations of potassium and by agents that increase endogenous production of cAMP, such as forskolin or 3-isobutyl-1-methylxanthine, but not by native cAMP, which cannot cross cell membranes. Elimination of glial cells by arabinoside-C did not reduce the activity of dbcAMP. GABAergic neurons, also present in these cultures, were much less dependent on the cyclic nucleotide analogue for their survival, and serotoninergic cells were not dependent at all. Therefore, cAMP-dependent signaling may be particularly crucial for the maturation and long-term survival of mesencephalic dopaminergic neurons.     

溶酶体抑制剂对小鼠行为学与黑质多巴胺神经元的影响

目的初步探讨溶酶体抑制剂对小鼠行为学及多巴胺神经元功能的影响。方法于小鼠右侧中脑黑质（SN）区,立体定向微量注入溶酶体抑制剂和蛋白酶体抑制剂。观察阿朴吗啡诱导的小鼠旋转行为改变以及行为学变化;检测黑质区酪氨酸羟化酶（TH）阳性细胞数。结果阿朴吗啡未能诱导出小鼠旋转行为。蛋白酶体抑制剂组为10～15圈。多巴胺损害程度与溶酶体的剂量相关。磷酸氯喹25μmol/L时多巴胺神经元几乎没有损害作用;50μmol/L时局部区域多巴胺神经元有轻微损害;100μmol/L时损害明显;200μmol/L时黑质区注射局部神经元损害最严重,黑质注射区未见有神经元存在。1～4周呈现出较为明显逐渐恢复的特点。结论溶酶体功能抑制与多巴胺神经元凋亡相关,不同剂量溶酶体抑制剂在不同时间对多巴胺神经功能的损害不同。     

Identification of Intact Neurotensin in the Substantia Nigra After Its Retrograde Axonal Transport in Dopaminergic Neurons

Two hours after the injection of (3-[125I]iodotyrosyl3) neurotensin into the striatum, a labeling was observed in the ipsilateral substantia nigra. In the present study, we demonstrated by HPLC that this radioactivity corresponded to intact neurotensin and to degradation products of this peptide. This finding provides the first clearcut evidence that a neuropeptide can be internalized and retrogradely transported in brain neurons. Therefore, the fact that intact neurotensin can be seen to exist over a long period of time in the cell body suggests that the retrograde transport process could perhaps be involved in the long-term effects of neuropeptides.     

帕金森病模型大鼠行为学改变与黒质致密部多巴胺能神经元存活率之间的相关性

目的探讨线刀损毁内侧前脑束建立的帕金森病模型大鼠行为学改变与黒质致密部多巴胺能神经元存活率之间的相关性。方法采用可伸缩线刀切断大鼠内侧前脑束建立单侧损伤的帕金森病大鼠模型;皮下注射阿朴吗啡后测试大鼠的旋转行为;取中脑黑质切片进行酪氨酸羟化酶免疫组织化学染色,通过计数黑质致密部酪氨酸羟化酶阳性神经元的数目得到多巴胺能神经元存活率。结果模型组大鼠损伤侧酪氨酸羟化酶阳性神经元的数目明显下降,而由阿朴吗啡诱导的旋转行为明显增加。结论线刀损毁术后4周,帕金森病模型大鼠由阿朴吗啡诱导的旋转行为和黒质致密部多巴胺能神经元存活率之间存在着明显的负相关。     

Toxic Effects of Iron for Cultured Mesencephalic Dopaminergic Neurons Derived from Rat Embryonic Brains

Iron, a transition metal possibly involved in the pathogenesis of Parkinson's disease, was tested for its toxic effects toward cultures of dissociated rat mesencephalic cells. When cultures were switched for 24 h to serum-free conditions, the effective concentrations of ferrous iron (Fe2+) producing a loss of 50% of dopaminergic neurons, as quantified by tyrosine hydroxylase (TH) immunocytochemistry, TH mRNA in situ hybridization, and measurement of TH activity, were on the order of 200 microM. High-affinity dopamine (DA) uptake, which reflects integrity and function of dopaminergic nerve terminals, was impaired at significantly lower concentrations (EC50 = 67 microM). Toxic effects were not restricted to dopaminergic neurons inasmuch as trypan blue dye exclusion index and gamma-aminobutyric acid uptake, two parameters used to assess survival of other types of cells present in these cultures, were also affected. Protection against iron cytotoxicity was afforded by desferrioxamine and apotransferrin, two ferric iron-chelating agents. Normal supplementation of the culture medium by serum proteins during treatment was also effective, presumably via nonspecific sequestration. Potential interactions with DA were also investigated. Fe2+ at subtoxic concentrations and desferrioxamine in the absence of exogenous iron added to the cultures failed to potentiate or reduce DA cytotoxicity for mesencephalic cells, respectively. Transferrin, the glycoprotein responsible for intracellular delivery of iron, was ineffective in initiating selective cytotoxic effects toward dopaminergic neurons preloaded with DA. Altogether, these results suggest (a) that ferrous iron is a potent neurotoxin for dopaminergic neurons as well as for other cell types in dissociated mesencephalic cultures, acting likely via autoxidation into its ferric form, and (b) that the presence of intra- and extracellular DA is not required for the observed toxic effects.     

脂多糖对大鼠多巴胺能神经元毒性作用的研究

目的　建立新的帕金森病 (Parkinson’sdisease ,PD)动物模型 ,探讨其发病机制。方法　在大鼠脑黑质(substantianigra ,SN)内注射脂多糖 (Lipopolysaccharide ,LPS)后 ,按大鼠不同存活期用高效液相色谱 (HPLC)来测定脑内多巴胺 (Dopamine,DA)及其代谢产物的含量 ;用免疫组化法观察酪氨酸羟化酶 (Tyrosinehydroxylase ,TH)阳性神经细胞、小胶质细胞的形态及数量变化。结果　DA及其代谢产物的含量在LPS注射侧随时间不同有不同程度下降 ,于第 14天达到最低 (P <0 0 1) ;注射侧黑质TH阳性神经元可以达到全部消失 ,该处可见大量被激活并有形态改变的小胶质细胞。结论　LPS可导致大鼠黑质多巴胺能神经元的损害     

Tyrosine Hydroxylase mRNA Concentration in Midbrain Dopaminergic Neurons Is Differentially Regulated by Reserpine

Tyrosine hydroxylase (TH)-mRNA, assayed by in situ hybridization combined with TH immunocytochemistry, showed a selective increase in the ventral tegmental area (A-10) but not in the substantia nigra (A-9) midbrain dopaminergic (DAergic) neurons 3 days after reserpine treatment. TH-mRNA in locus ceruleus noradrenergic (A-4) neurons was increased by reserpine, as confirmed by RNA blot hybridization. These findings show that TH-mRNA is differentially regulated in midbrain DAergic neurons in response to reserpine.     

Pyruvate Carboxylase Activity in Primary Cultures of Astrocytes and Neurons

Abstract: The activity of the pyruvate carboxylase was determined in brains of newborn and adult mice as well as primary cultures of astrocytes, of cerebral cortex neurons, and of cerebellar granule cells. The activity was found to be 0.25 ± 0.14, 1.24 ± 0.07, and 1.75 ± 0.13 nmol · min⁻¹· mg⁻¹ protein in, respectively, neonatal brain, adult brain, and astrocytes. Neither of the two types of neurons showed any detectable enzyme activity (i.e., < 0.05 nmol · min⁻¹· mg⁻¹). It is therefore concluded that pyruvate carboxylase is an astrocytic enzyme.     

Toxicity of 1-Methyl-4-Phenylpyridinium for Rat Dopaminergic Neurons in Culture: Selectivity and Irreversibility

Cultures of dissociated embryonic rat mesencephalic cells were exposed to 10 microM 1-methyl-4-phenylpyridinium (MPP+), a concentration shown earlier to result in loss of greater than 85% of tyrosine hydroxylase (TH)-positive neurons without affecting the total number of cells observed by phase-contrast microscopy. To characterize better the selectivity of the toxic action of MPP+, other parameters were measured reflecting survival and function of dopaminergic or nondopaminergic neurons. Exposure of cultures to 10 microM MPP+ for 48 h reduced TH activity to 11% of control values without reducing protein levels. [3H]Dopamine uptake was reduced to less than 4% of control values, whereas the uptake of gamma-[3H]aminobutyric acid ([3H]GABA) was not affected in these cultures. This same treatment failed to reduce the number of cholinergic cells visualized in septal cultures and did not affect either choline acetyltransferase activity or high-affinity choline uptake. To assess for possible recovery of dopaminergic neurons, cultures were exposed to 10, 1.0, or 0.1 microM MPP+ for 48 h and then kept for up to 6 days in MPP(+)-free medium. After exposure to 10 microM MPP+, the number of TH-positive neurons, their neurite density, TH activity, and [3H]dopamine uptake remained at constant, reduced levels throughout the period of observation after termination of exposure, whereas GABA uptake remained normal. Treatment with lower concentrations of MPP+, i.e., 1.0 and 0.1 microM, induced less pronounced dopaminergic toxic effects. However, no recovery was seen after posttreatment incubation in toxin-free medium. These findings provide evidence that MPP+ treatment results in highly selective and irreversible toxicity for cultured dopaminergic neurons.