6-羟基多巴胺致帕金森病大鼠模型的建立与评价

应用6-羟基多巴胺(6-hydroxydopamine,6-OHDA)制造帕金森病(Parkinson’s disease,PD)大鼠模型,建模成功后采用组织化学方法检测模型黑质多巴胺能神经元变化,并从行为学及黑质多巴胺能神经元数目的变化对模型进行综合评价。取Wistar大鼠,采用立体定向法,进行右侧纹状体区6-OHDA 双点注射。结果表明,6-OHDA纹状体注射可成功的诱导帕金森大鼠模型。     

睾丸肾上腺素能受体和胆碱能受体的分布及神经递质对睾丸间质细胞增殖的影响

试验旨在研究不同发育时期小鼠睾丸肾上腺素能受体(β1AR、β2AR、β3AR、α1A、α1B和α1D)和胆碱能受体(M1、M2、M3、M4和M5)mRNA的表达,以及神经递质去甲肾上腺素(norepinephrine,NE)和乙酰胆碱(acetylcholine,Ach)对发育期小鼠睾丸间质细胞增殖的影响。RT-PCR结果表明,β1AR和β2AR mRNA在睾丸发育的3个时期都表达,β3AR、α1A和α1B mRNA在小鼠发育早期的睾丸表达,在成年期睾丸不表达,α1D mRNA在睾丸发育的早期不表达,成年期表达;胆碱能受体M1R、M2R、M3R和M5R mRNA在睾丸发育的3个时期都表达,而M4R mRNA主要在成年期表达。另外通过NE和Ach处理体外培养的发育期睾丸间质细胞,发现Ach处理组BrdU阳性细胞的数量明显增加(P＜0.01)。结果表明,肾上腺素能受体和胆碱能受体在小鼠睾丸的整个发育时期都表达,Ach可促进发育期小鼠睾丸间质细胞的增殖。     

中央杏仁核内微量注射地塞米松对NA／NPY在NTS所致心血管效应的影响

运用电生理学技术和心血管实验,观察了中央杏仁核(CeA)神经元与孤束核(NTS)神经元活动之间的相互作用,以及在CeA 内注射地塞米松(Dex)对NTS内去甲肾上腺素(NA)/神经肽Y(NPY)诱导的心血管效应的影响。电生理学实验显示,用L-谷氨酸兴奋CeA 神经元后,NTS内神经元活动主要呈现抑制反应(12/18);向NTS内注射NPY 后,CeA 内神经元活动也以抑制反应为主(13/23)。向CeA 内注射Dex,可以消除NTS内微量注射NA/NPY 所引起的降压和减慢心率效应。NA:动脉血压下降(- 0.37±0.25) kPa,心率下降(- 5±5) 次/m in,对照分别为(- 2.57±0.21) kPa,(- 33±11) 次/m in。NPY:(- 0.47±0.29)kPa,(- 10±8) 次/m in,对照(- 3.37±0.36) kPa, (- 50±9) 次/m in。由此表明,不仅CeA 与NTS在心血管活动调节中存在相互抑制作用,而且当CeA 受到高水平的糖皮质激素作用时,会抑制NTS内降压系统的作用。这可能是应激致发高血压的重要原因     

从下丘脑室旁核向背肩胛棕色组织发送投射的谷氨酸能神经元的鉴定

谷氨酸能神经元可抑制摄食并防止肥胖。有证据表明,在缺乏囊泡型谷氨酸转运蛋白2(VGlut2)的小鼠中,重新表达专一蛋白1(SIM1)神经元上的VGlut2可降低采食。然而,VGlut2神经元的位置和轴突投射尚不清楚。因此,本试验探寻了向背肩胛棕色脂肪组织(IBAT)发送神经元投射的上游脑区,检测了VGlut2和黑皮质素4型受体(MC4R)神经元的脑内定位,确认了胰淀素(amylin)对于VGlut2和MC4R免疫阳性神经元表达的调控作用,拓展了VGlut2神经元发送神经支配的大脑区域。结果显示,PVN^VGlut2神经元向IBAT发送密集的神经支配信号。在前皮质杏仁核(ACo)、纹状体(CPu)和海马齿状回(DG)发现VGlut2与MC4R共定位。amylin注射显著促进PVN^VGlut2::MC4R(P=0.004 8)免疫阳性神经元表达,提示VGlut2与MC4R涉及amylin的调节过程。PVN^VGlut2神经元向下丘脑腹内侧核(VMH)、外侧下丘脑(LH)、中脑导水管周围灰质(PAG)和蓝斑(LC)核团发送神经元投...     

NO对大鼠中脑腹侧被盖区DA神经元的调节

实验观察了微量注射 NOS抑制剂 L -NAME及微量联合注射 NO的前体 L-精氨酸( L-Arg) L-NAME于大鼠中脑腹侧被盖区( VTA)对该部位多巴胺神经元的调节。发现VTA注射 L -NAME( 1 mg/ 5μL )后 ,伏隔核( Acb)多巴胺 ( DA )代谢产物—双羟苯乙酸( DOPAC)水平升高到注射前的 1 2 2 .5% ( P <0 .0 0 1 ) ,小剂量注射 L-NAME( 0 .2 mg/ 5μL)对伏隔核 DOPAC水平无明显影响 ;同样方法联合注射 L-Arg( 3 0 0 μg/ 5μL) L-NAME( 1 mg/5μL)后 ,伏隔核 DOPAC水平无明显变化。结论 :VTA微量注射 L-NAME兴奋了该部位的DA神经元 ,而 L -Arg L -NAME联合注射 ,却不能影响 DA神经元的活动 ,说明 NO可以通过L-Arg-NOS-NO途径参与 VTA多巴胺神经元的调节     

冷应激反应的神经机制及对免疫系统的影响

一、动物对冷应激反应的神经机制 (一)中枢神经系统反应冷应激下,中枢肾上腺素能神经元被激活,引起外周自主神经(主要是交感神经)的活动加强,使外周神经和交感一肾上腺髓质系统(SAM)处于长时间高度激活状态,E(肾上腺素)和NE(去甲肾上腺素)的合成、释放和周转增强,使机体的神经一内分泌系统和免疫功能之间产生联系,构建了一个完整的调节网络.     

疑核在针刺“足三里”调节机体细胞免疫中的作用

实验用3～4月龄健康家兔(体重2.0～2.5kg)15只。实验组兔经麻醉后固定于大脑立体定位仪上,于A P20-21、L1-1.5处埋置套管(H20-21),以微量注射器注入2%盐酸利多卡因0.5μL 阻断单侧疑核;对照组兔疑核未被阻断,其他处置同实验组。所有动物均电针“足三里”穴0.5 h,并于电针前和电针后即刻及0.5,1.0,1.5,2.0,2.5,3.0 h采血测定PHA诱导的淋巴细胞转化率、T细胞百分率及E-玫瑰花环形成率。结果,实验组兔的上述免疫指标在电针前,后无明显差异,而对照兔电针后的免疫指标显著高于电针前。由此认为,电针“足三里”穴可能通过神经-内分泌-免疫网络调节细胞免疫,而疑核是该调节环路中的一个重要的中继核团。     

营养状况对樱桃谷鸭下丘脑神经肽Y的影响

为了探讨营养状况对家禽下丘脑神经肽Y(neuropeptide Y,NPY)的影响,选用来自同一父母代的樱桃谷鸭为研究对象,按营养状况分成4组对照组(Ⅰ组)、低蛋白组(Ⅲ组)、低能量组(Ⅳ组)和禁食组(Ⅱ组),测定了各组血清总蛋白、甘油三酯的含量,从生化指标上进一步确定了其营养状况的差别;并采用免疫细胞化学ABC法结合计数法,定性定量地观察和比较了在不同营养状况下下丘脑NPY阳性神经元胞体的变化.结果显示(1)Ⅱ组、Ⅲ组和Ⅳ组视上核与下丘脑外侧核内NPY样免疫反应比Ⅰ组显著增强,表现为NPY阳性神经元胞体数目增多,密度增高;(2)Ⅱ组螺旋外侧核和顶盖前核内有少量的NPY阳性神经元胞体,而其他组未观察到此现象.计数法分析进一步从量的角度证实了上述差异的显著性下丘脑外侧核和视上核NPY阳性神经元胞体数,Ⅱ组平均为(48.10±2.17)个/片,Ⅲ组平均为(39.70±1.68)个/片,Ⅳ组平均为(40.45±1.57)个/片,Ⅰ组平均为(18.05±1.37)个/片.经方差分析发现,除Ⅲ组与Ⅳ组差异不显著(P＞0.05)外,其他组间差异极显著(P＜0.01).上述结果表明,营养状况对樱桃谷鸭下丘脑神经肽Y有显著影响,且影响的主要部位在视上核和下丘脑外侧核,禁食、低蛋白和低能量均能使樱桃谷鸭下丘脑NPY神经元胞体数显著增加,其中能量与蛋白质对樱桃谷鸭NPY合成与释放的影响无显著差异.     

激光麻醉的镇痛机理研究——氦氖激光照射胫神经前后马脑脊液中乙酰胆碱含量、胆碱酯酶活性变化规律的探讨

本文通过对9匹临床健康马激光麻醉实验研究,用18mWHe-Ne激光照射马胫神经30分钟,确认达到了良好的全身镇痛状态,观察分析了激光照射前后三个不同时期脑脊液(CSF)内乙酰胆碱(Ach)含量、真假胆碱酯酶活性的消长规律。激光照射30分后停照10分,CSF内Ach含量显著增高,即由照前的197.91±17.89pg/ml升高到405.50±23.45pg/ml,激光停照60分时,CSF内Ach含量恢复至与照前值没有显著差异的水平。认为在激光镇痛过程中中枢胆碱能神经元功能增强,积极参与了激光麻醉。正常马CSF中真性胆碱酯酶(AchE)与假性胆碱酯酶(ChE)活性分别为2.21±0.1454、0.36±0.0486μM/ml/37℃/30′。激光照射前后马CSF内AchE和ChE均未发生显著变化。     

纹状体内注射6-羟基多巴胺制备兔帕金森病模型

应用脑立体定位技术微量注射6-OHDA于兔右侧纹状体内。术后每周观察以阿扑吗啡（Apomorphin,APO）诱导的旋转行为,并于术后6周处死兔,以黑质酪氨酸羟化酶（Tyrosine hydroxylase,TH）免疫组化染色,观察黑质多巴胺能神经元的形态、结构及数量变化。结果表明,部分兔在术后即出现行动迟缓、躬身、易激怒等异常行为。术后6周时,20只兔中有16只在阿扑吗啡诱导后30min内的平均旋转圈数大于7r/min,达到成功模型标准。模型成功率达到80%。TH免疫组化染色可见正常对照组、假手术组及模型组未损侧黑质内有胞浆浓染、突起明显的TH免疫反应阳性神经元分布,神经元数量较多,轴突长度较长,且3者差异不显著（P〉0.05）;而模型组损毁侧黑质内TH免疫反应阳性神经元与上述3者相比,数目明显减少（P〈0.05）,残存的细胞染色较浅,胞体轮廓和突起均不清晰,轴突长度明显变短（P〈0.05）。结果提示,将6-OHDA注射于兔单侧纹状体是一种制备帕金森病（Parkinson’s disease,PD）模型的有效方法,此法操作简便,动物死亡率低,模型制作成功率高。     

The role of noradrenaline in the generation of the preovulatory LH surge in the ewe

Increasing plasma estrogen (E) levels during the follicular phase of the estrous cycle trigger the pre-ovulatory surge of gonadotropin-releasing hormone (GnRH)/LH. Noradrenaline (NA)-producing cells of the brain stem are involved in regulating GnRH cells and project to the preoptic area (POA) and bed nucleus of stria terminalis (BnST). Input to GnRH cells may be direct or indirect, via relay neurons in the POA/BnST. To investigate this, we ascertained whether an ₁-adrenergic antagonist would block/delay the LH surge in ovariectomised (OVX), E-treated ewes. E benzoate (EB) (50 μg) was injected (i.m.) and Doxazosin (100 nmol/h) or vehicle was infused into the third ventricle 2–26 h after EB injection. Doxazosin reduced the magnitude of the LH surge, but did not affect timing. To determine if NA is released in the POA/BnST of cyclic ewes, we immunostained dopamine-β-hydroxylase (DBH) in terminal fields. Reduced numbers of varicosities staining for DBH indicates release of NA. The number of varicosities immunostained for DBH was reduced in the dorsal and lateral BnST during the follicular phase and during the preovulatory LH surge compared to the luteal phase. These data suggest that noradrenergic mechanisms are involved in generation of the GnRH/LH surge via projections to the BnST and relay to GnRH cells. Since Doxasozin reduced the magnitude of the LH surge in the E-treated OVX ewe, and release of NA in cyclic ewes occurred during the follicular phase of the estrous cycle, we speculate that NA is a permissive factor in surge generation. Thus, increased noradrenergic activity is not a trigger mechanism for initiation of the surge.     

Role of noradrenergic receptors in the bed nucleus of the stria terminalis in regulating pulsatile luteinizing hormone secretion in female rats

The bed nucleus of the stria terminalis (BNST) is one of the brain areas densely innervated by noradrenergic neurons originating in the brain stem. The present study aims to determine the role of noradrenergic receptors in the BNST in regulating pulsatile luteinizing hormone (LH) secretion in female rats. Ovariectomized (OVX) or estrogen-primed OVX (OVX+E2) rats received three 1-h-interval injections of 0.05 micromol of noradrenaline (NA), phenylephrine (alpha1-adrenergic receptor agonist), clonidine (alpha2-agonist), or isoproterenol (beta-agonist) into the BNST. Injection of NA or alpha1-adrenergic agonist into the BNST strongly suppressed pulsatile LH secretion in OVX+E2 rats with a significant (P < 0.05) decrease in the mean LH level for 3 h and LH pulse frequency, but alpha2-and beta-agonists did not affect any of the LH pulse parameters. In OVX animals, alpha1- and alpha2-adrenergic agonists caused a significant change in LH pulse frequency and amplitude, respectively, though the effect was not as apparent as the NA- or alpha1-agonist-induced changes in OVX+E2 animals. These results indicate that NA inputs to the BNST suppress pulsatile LH secretion via alpha1-adrenergic receptors and that estrogen enhances this suppression.     

Absence of postmyelographic adverse effects and high radiographic resolution of iotrolan used in the cervical myelography for the clinically normal cat: an open-placebo controlled study

Iotrolan solution at 300 mgI/ml (Isovist 300, Schering AG), was injected into the cerebromedullary cistern in 8 cats (Isovist group), at a volume of 0.5 ml/Kg BW and over one minute. In 3 cats (control group), normal saline was also injected at a same volume and speed. During the subarachnoid injection and at 1, 5 and 10 min thereafter all the cats were being monitored for evidence of respiratory arrest, bradycardia or changes in the arterial blood pressure. The cats were also checked for postmyelographic adverse effects for at least 3 h after their complete recovery from anaesthesia. Postmyelographic neurologic examination was done for 3 consecutive days and the myelographic quality of the films taken at 5 and 15 min postinjection was evaluated. No significant differences were detected between the Isovist and the control groups for all the variables evaluated immediately before, during and up to 3 days after the injection. No actual post-myelographic adverse effects were seen in the animals of both groups. At 15 and 30 min postinjection the opacity, the distention of the subarachnoid space and the detail of all the myelograms obtained were assessed as highly diagnostic.     

纹皮蝇HypoderminC基因真核表达质粒构建及小鼠免疫试验

根据GenBank发表的Hypodermin C(HC)基因序列设计引物,以原核表达载体pET-HC为模板,PCR扩增HC基因,将克隆基因插入到真核表达载体pVAX1中,构建重组真核表达质粒pVAX1-HC。将重组质粒pVAX1-HC转染小鼠胎儿成纤维细胞,以间接免疫荧光法检测HC基因在细胞中表达。用真核表达质粒pVAX1-HC经双侧胫前肌注射昆明小鼠,对小鼠进行体液免疫和细胞免疫研究。结果表明,小鼠血清抗体水平和淋巴细胞增殖水平明显高于非免疫对照组。本研究成功构建了真核表达质粒pVAX1-HC,为纹皮蝇核酸疫苗研发奠定了基础。     

Light Microscopy of the Enteric Nervous System of Horses with or without Equine Dysautonomia (Grass Sickness): its Correlation with the Motor Effects of Physostigmine

Light microscopy was undertaken on sections from the caudal flexure of the duodenum and the terminal ileum proximal to the ileocaecal fold in 5 control horses, 5 horses with acute grass sickness (AGS), and 5 horses with chronic grass sickness (CGS). With the exception of the ileal submucous plexus of the CGS group, the AGS group had the lowest number of neurons as measured using a subjective scoring scheme. The proportion of abnormal neurons in the AGS group was similar in both plexuses and both regions, whereas the values for the CGS group were much higher in the duodenal region than in the ileal region. The motility of tissue adjacent to that used for histology was measured isometrically in vitro. The increase in the rate of contractions following exposure to physostigmine was greatest for the AGS group, both from the duodenal and from the ileal region. The latency was longest for the AGS group, suggesting that the material from this group had the least number of active cholinergic neurons. The studies with physostigmine thus indicated that the most severe functional damage occurred in cases of AGS. These findings confirm that extensive damage occurs in the enteric neurons in equine grass sickness. There was good correlation between the functional cholinergic responses and the extent of neuronal degeneration.     

Estimation of body composition of neonatal pigs via deuterium oxide dilution: validation of technique

Five studies were conducted to determine 1) the time period required for the equilibration of deuterium oxide (D2O) in body water in neonatal pigs following the intravenous (iv), intramuscular (im) or intraperitoneal (ip) injection of D2O and 2) the accuracy and precision of estimating body water, protein, fat and ash in neonatal pigs from the body D2O pool space. Deuterium oxide administered by iv and im injection equilibrated with body water by 40 and 20 min postinjection, respectively. Body D2O space determined from individual samples of blood fluids drawn at 40, 80 or 120 min postinjection accurately reflected body water determined by desiccation. The difference between D2O pool space determined at 120 min postinjection and body water in 4-kg pigs injected iv averaged - .050 kg, and the magnitude of the difference was relatively constant (standard deviation [SD] = .121 kg). The D2O pool space determined from im injection overestimated body water slightly (.163 kg), but the precision of the estimate was good (SD = .019 kg). Deuterium oxide injected ip did not consistently equilibrate by 200 min postinjection, and the estimated D2O pool space was not indicative of body water. The 90% confidence limits for estimating body water, protein, fat and ash in 4-kg pigs as determined by im and iv injection of D2O were .020 and .092 kg, .014 and .021 kg, .030 and .043 kg and .007 and .047 kg, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)     

Intrinsic innervation of the horse ileum

This paper describes the morphology and distribution of the enteric nervous system (ENS) cells and fibres immunoreactive for choline acetyltransferase (ChAT), neuronal nitric oxide synthase (nNOS), substance P (SP), calcitonin gene-related peptide (CGRP), NF200 kDa (NF200), and S100 protein. The percentages of subclasses of enteric neurons in the total neuronal population were investigated by the use of anti-PGP 9.5 or anti-NSE antibodies.ChAT-IR myenteric plexus (MP) and submucosal plexus (SMP) neurons were 66 ± 7% and 74 ± 15%, respectively, whereas those cells expressing nNOS-IR were 38 ± 7% and 5 ± 1%, respectively. MP and SMP neurons expressing both phenotypes were also present. SP-IR was expressed by 14 ± 13% of MP and 66 ± 8% of SMP neurons whereas CGRP-IR was observed only in the SMP (43 ± 6%). NF200-IR was expressed by 61 ± 15% and 91 ± 10% of the MP and SMP neurons, respectively. The majority of the CGRP-IR SMP neurons expressed also SP-IR. Almost all SP-IR neurons in both the plexuses were cholinergic. The present study quantifies the main neuronal subpopulations of the ENS of the horse ileum; these data might be utilized to understand the neuronal modifications which occur in several gastrointestinal tract disorders.     

Hypothalamus involvement in the reticulo-rumen motor and behavioural disturbances induced by morphine in sheep

Morphine (20 and 40 µg/kg) administered into the cerebral ventricle of conscious sheep caused significant inhibition of the mean frequency and the average amplitude of primary ruminal contractions by 45 min after injection. Between 90 and 120 min, morphine (40 µg) provoked a significant increase in the amplitude (p<0.01). At both doses it caused strong psychomotor excitability that lasted for more than 140 min. Isolation of the hypothalamus prevented both the inhibitory effects of morphine on rumen motility and the drug-induced psychomotor excitability. Histopathological analysis of slices of the hypothalamus, pons and medulla indicated descending degenerative changes in the nervous pathways connecting the hypothalamus with lower structures in the brain. These results suggest either that hypothalamic isolation caused degeneration of inhibitory descending pathways that connect the hypothalamus with the gastric centres or that structures of importance for forestomach motility are not located within the gastric centres but elsewhere in the brain, for example in the hypothalamus.Abbreviations Ach acetylcholine - CSF cerebrospinal fluid - ICV intracerebroventricular(ly) - NA noradrenalin - 6-OHDA 6-hydroxydopamine - PAGM periaqueductal grey matter     

Intramuscular Selenium Administration in Selenium-Deficient Cattle

Nine recently weaned Hereford heifers were randomly assigned to a control group (n = 3) or a treatment group (n = 6). The animals were selenium (Se) deficient (mean ± SD blood Se concentration = 0.024 ± 0.012 μg/mL). They were maintained on a selenium-deficient diet, and on day 0 of the study the treatment group was given 0.05 mg Se/kg body weight intramuscularly, while the control group received a placebo. The Se concentration of blood, serum, and urine as well as the glutathione peroxidase (GSH-Px) activity of blood and serum was measured over an 84-day period. Peak blood Se and serum Se concentrations (mean ± SD) in the treatment group occurred at 5 hours postinjection and were 0.131 ± 0.028 μg/mL and 0.154 ± 0.027 μg/mL, respectively. The mean blood Se concentration of the treatment group was greater (P < .05) than that of the control group for the first 28 days after injection. The mean serum Se concentration of the treatment group was greater (P < .05) than that of the control group for all times after injection, except for day 56. The mean (±SD) blood GSH-Px activity of the treatment group (12.0 ± 2.3 mU/min/mg hemoglobin) was increased (P < .05) over the control group (2.0 ± 1.4 mU/min/ mg hemoglobin) by day 28 and continued to be greater (P < .05) throughout the 84 day postinjection period. The blood GSH-Px activity and the blood Se concentrations in the treatment group heifers did not reach concentrations considered indicative of Se adequacy (30 mU/min/mg hemoglobin and 0.10 μg/mL, respectively) except briefly, at 5 hours postinjection when the blood Se concentration of the treatment group was 0.131 ± 0.028 μg/mL. The mean serum GSH-Px activity of the treatment group did not differ at any time from that of the control group (P≥ .17). The mean (±SD) fractional excretion (FE) of Se, as an estimate of Se excretion, was greater (P < .05) in the treatment group heifers (n = 5; 6.2 ± 2.5%) than in the control heifers (n = 3; 1.3 ± 0.6%) at 24 hours postinjection. The mean (±SD) weight gain, from day 0 to day 84, for the treatment group heifers was 63.0 ± 18.1 kg and the mean weight gain for the control group heifers was 53.1 ± 7.3 kg at 84 days postinjection and there was no difference between the groups (P < .39). Conclusions drawn from this study include: 1) the increase in blood GSH-Px activity occurs approximately 28 days after Se injection given to Se-deficient heifers, 2) a single label dose of injectable Se does not result in blood Se concentrations or blood GSH-Px activity normally considered to be adequate, 3) the label dose of injectable Se, although therapeutically beneficial for nutritional myodegeneration (NMD), does not seem to be a desired method for long-term Se supplementation of cattle consuming a Se-deficient diet, and 4) blood Se is a better predictor of Se status than serum Se. (Journal of Veterinary Internal Medicine 1993; 7:342–348. Copyright © 1993 by the American College of Veterinary Internal Medicine.)     

疑核在下丘脑外侧区调节机体细胞免疫中的作用

实验用3～4月龄、1.5～2.5kg健康青紫蓝家兔16只,麻醉后固定于脑立体定位仪上.对其中11只刺激下丘脑外侧区;5只损毁疑核后,再刺激下丘脑外侧区.在刺激前和刺激后不同时间采血,检测外周血中T细胞百分率和PHA诱导的淋巴细胞转化率.结果:刺激下丘脑外侧区,能迅速提高外周血中的T细胞百分率,过一段时间,T细胞转化率也显著升高.而损毁疑核后再刺激下丘脑外侧区,外周血中T细胞百分率和淋巴细胞转化率均无显著变化.由此认为,下丘脑外侧区具有增强机体细胞免疫功能的作用,疑核是下丘脑外侧区调节机体免疫反应的中继核.