大鼠眼水平同向运动相关核团向动眼神经核投射的神经递质通路研究

发布时间：2018-03-18 06:41

本文选题：大鼠　切入点：眼水平同向运动　出处：《青岛大学》2007年博士论文　论文类型：学位论文

【摘要】： 目的探讨大鼠脑干内向动眼神经核内直肌亚核传入的运动前神经元的定位和分布，建立眼水平同向运动相关的脑干中枢核团与动眼神经核内直肌亚核之间的神经纤维通路。在此基础上，进一步检测这些通路中可能存在的兴奋性和抑制性的神经递质，从而建立眼水平同向运动相关核团向动眼神经核投射的抑制性和兴奋性神经递质通路，为眼水平同向运动障碍性疾病的发生和治疗提供解剖学依据。材料和方法健康成年Wister大鼠30只，220～250g，随机分成2组。第1组14只，大鼠麻醉后，置于脑立体定位仪上，将50％的辣根过氧化物酶(horseradish peroxidase，HRP)溶液0.1μl缓慢注入到大鼠脑的一侧动眼神经核(oculomotor nucleus，OMN)内直肌亚核(medial rectus subdivision)，存活48h后，灌注固定取脑，连续冠状冰冻切片(25～40μm)分5套收集，经TMB呈色反应，1套切片用于观察注射部位和脑干核团中HRP逆行标记细胞的分布。另外4套切片经TMB反应后分别进行抗谷氨酸(glutamate，Glu)，γ-氨基丁酸(γ-amino-butyric acid，GABA)，谷氨酸脱羧酶(glutamic acid decarboxylase，GAD)和谷氨酸受体1型(Glutamate Receptor1，GluR1)血清的SABC免疫组织化学染色，检测这些核团中存在的神经递质。第2组16只，大鼠麻醉后，置于脑立体定位仪上，，将3％的荧光金(fluorogold，FG)0.1μl缓慢注入大鼠脑一侧动眼神经核内直肌亚核。动物存活5～7天，灌注固定取脑，连续冠状冰冻切片(25μm)分5组收集。1套用于观察注射部位和FG逆行标记细胞在脑干核团中的分布，另外4套分别进行抗GABA、Glu、GluR1、GAD血清的免疫荧光组织化学反应，观察这些脑内核团中逆行标记细胞、免疫荧光标记细胞和双标记细胞的分布及其向动眼神经核投射的神经递质联系。结果 (1)逆行标记结果：将HRP或FG注入一侧动眼神经核(OMN)内直肌亚核，在对侧展神经核(abducens nucleus，ABN)、双侧前庭神经内侧核(medial vestibular nucleus，MVN)、脑桥旁正中网状结构(pontine paramedian reticular formation，PPRF)及同侧前庭上核(superior vestibular nucleus，SVN)和舌下前置核(nucleus of prepositus hypoglosal，PH)均可见逆行标记细胞和纤维。 (2)GABA、GAD、Glu、GluR1的免疫组织(荧光)化学反应的切片中分别可见逆行标记细胞，对应的一抗的免疫(荧光)阳性细胞，逆标／免疫(荧光)双标细胞三种标记细胞。在对侧展神经核和同侧舌下前置核主要存在Glu免疫／逆标和GluR1免疫／逆标的双标记细胞，GABA免疫／逆标、GAD免疫／逆标的双标记细胞较少；在同侧前庭神经内侧核主要可见Glu免疫／逆标和GluR1免疫／逆标的双标记细胞，未见明显的GABA免疫／逆标、GAD免疫／逆标的双标记细胞；而对侧前庭内侧核中则四种免疫双标记细胞都有；脑桥旁正中网状结构中GABA免疫／逆标、GAD免疫／逆标、Glu免疫／逆标、GluR1免疫／逆标的双标记细胞分布较为均匀。结论 (1)在对侧展神经核，双侧前庭神经内侧核、同侧前庭上核、舌下前置核以及脑桥旁正中网状结构中均有纤维投射到动眼神经核内直肌亚核，并可能参与协调双眼的水平同向运动。 (2)展神经核核间神经元向对侧动眼神经核投射，其兴奋性投射主要以Glu为神经递质，部分通过GluR1型受体起作用；其抑制性投射中GABA可能不是起主要作用的神经递质。舌下前置核向同侧内直肌运动神经元投射，其兴奋性投射和抑制性投射与展神经核相似。 (3)前庭神经核向同侧动眼神经核内直肌亚核的投射，主要是兴奋性的，用谷氨酸作为神经递质，部分通过GluR1受体起作用；而前庭神经内侧核向对侧内直肌亚核的投射包含兴奋性和抑制性的投射，分别用Glu和GABA作为相应的神经递质；脑桥旁正中网状结构向双侧动眼神经核内直肌亚核的投射同样存在兴奋性和抑制性的投射，其中Glu和GABA分别起一定的作用，是否存在递质共存，需进一步研究。
[Abstract]:objective
To investigate the localization and distribution of rat brain stem within the medial rectus subnucleus of the afferent premotor neurons, a conjugate horizontal eye movement related brainstem nuclei and medial rectus subnucleus between the nerve fiber pathway. On this basis, further testing of these pathways in possible excitatory and inhibitory nerve the transmitter, so as to establish the conjugate horizontal eye movements related nucleus to oculomotor nucleus of inhibitory and excitatory neurotransmitter pathways, as happened to the same level of eye movement disorders and treatment to provide the anatomical basis.
Materials and methods
30 healthy adult Wister rats, 220 ~ 250g, were randomly divided into 2 groups. The first group of 14 rats, rats were anesthetized and placed in a stereotaxic apparatus, 50% horseradish peroxidase (horseradish, peroxidase, HRP) solution 0.1 l slowly injected into the rat brain side of the oculomotor nucleus (oculomotor nucleus. OMN) medial rectus subnucleus (medial rectus subdivision), the survival of 48h after perfusion fixed brains, continuous coronal frozen sections (25~40 m) 5 sets collected by TMB color reaction, 1 sets of sections for observation of the injection site and brainstem nuclei in HRP retrograde labeled cells. The distribution of the other 4 sets section after the TMB reaction were anti glutamic acid (glutamate, Glu), GABA (gamma -amino-butyric acid GABA (glutamic acid), glutamic acid decarboxylase decarboxylase, GAD) and glutamate receptor type 1 (Glutamate Receptor1, GluR1 SABC) immunohistochemical staining to detect the serum. There are some neurotransmitters in the nuclei. The second group of 16 rats, rats were anesthetized and placed in a stereotaxic apparatus, fluorescent gold 3% (Fluorogold, FG) 0.1 l slowly injected into the rat brain side of medial rectus subnucleus. The animal survived for 5~7 days, take continuous coronary perfusion fixed brain frozen section (25 m) were divided into 5 groups to observe the distribution of.1 from the injection site and FG retrograde labeled neurons in the brainstem nuclei, the other 4 were anti GABA, Glu, GluR1, immunohistochemical reaction of GAD in serum, to observe the retrograde labeling of cells in these nuclei, the distribution of immunofluorescence the labeled cells and double labeled cells and projecting to the oculomotor nucleus of the neurotransmitter.
Result
Results: (1) retrogradely labeled HRP or FG into the unilateral oculomotor nucleus (OMN) medial rectus subnucleus, contralateral abducens nucleus (abducens, nucleus, ABN), bilateral medial vestibular nucleus (medial vestibular nucleus, MVN), PPRF (pontine paramedian reticular formation, PPRF) and the same the lateral superior vestibular nucleus (superior vestibular nucleus, SVN) and hypoglossal nucleus (nucleus of prepositus hypoglosal, PH) were retrogradely labeled cells and fibers.
(2) GABA, GAD, Glu, GluR1 immunohistochemistry (Ying Guang) chemical reaction sections were observed retrogradely labeled cells, a corresponding antibody (Ying Guang) positive cells labeled / immune (Ying Guang) three kinds of double labeled cells labeled cells in the contralateral abducens nucleus and ipsilateral hypoglossal pre nuclear are Glu / GluR1 and immune labeled immune / retrogradely labeled double labeled neurons and GABA labeled GAD / immune, immune / retrogradely labeled double labeled neurons in the ipsilateral vestibular less; medial nucleus mainly visible Glu and GluR1 retrogradely labeled immune / immune / retrogradely labeled double labeled cells, no obviously GABA / GAD retrogradely labeled immune, immune / retrogradely labeled double labeled neurons; while the contralateral medial vestibular nucleus in four immune cells have double labeling; PPRF GABA / GAD retrogradely labeled immune, immune / inverse standard, Glu / GluR1 retrogradely labeled immune, immune / inverse calibration the double labeled cells The distribution is more uniform.
conclusion
(1) in the lateral abducens nucleus, bilateral medial vestibular nucleus, ipsilateral superior vestibular nucleus, sublingual anterior nucleus and para Pontus medialis reticular structure, all fibers projected into the rectus subnucleus of the oculomotor nucleus, and may be involved in coordinating the horizontal horizontal movement of the eyes.
(2) the abducens nucleus nucleus neurons to the lateral projection of the oculomotor nucleus, the excitatory projection takes Glu as the main neurotransmitter part via GluR1 receptors; the inhibition of GABA projection may not mainneurotransmitter. Hypoglossal nucleus to the ipsilateral medial rectus motoneurons projecting, the the excitatory and inhibitory projection projection and abducens nucleus is similar.
(3) vestibular nuclei projecting to the ipsilateral medial rectus subnucleus, predominantly excitatory, use glutamate as neurotransmitter, partly mediated by GluR1 receptors; and the medial vestibular nucleus to the lateral projection of the medial rectus subnucleus contains excitatory and inhibitory projection, respectively Glu and GABA as the corresponding neurotransmitters; PPRF projecting to the bilateral medial rectus subnucleus have excitatory and inhibitory projections, which Glu and GABA play a role, whether there is coexistence of transmitters, to be further studied.

【学位授予单位】：青岛大学
【学位级别】：博士
【学位授予年份】：2007
【分类号】：R77;R33

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