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Histamine as a neurotransmitter: Fluorescence immunohistochemistry of its synthetic and degradative enzymes and purification of histamine receptor protein.

组胺作为神经递质：其合成酶和降解酶的荧光免疫组织化学以及组胺受体蛋白的纯化。

基本信息

批准号：
59480132
负责人：
WATANABE Takehiko
金额：
$ 4.16万
依托单位：
TOHOKU UNIVERSITY (1986)Osaka University (1984-1985)
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (B)
财政年份：
1984
资助国家：
日本
起止时间：
1984 至 1986
项目状态：
已结题

项目摘要

Histamine neuron system in rat brain was demonstrated by the fluorescence immunohistochemistry with histidine decarboxylase, a histamine-forming enzyme, as a marker. The cell bodies exist only in the posterior hypothalamus and mammillary body as the tubelomammillary nucleus, and the axons with many varicosities were particularly dense in the hypothalamus, olfactory bulb, cerebral cortex, medial amygdaloid nucleus, med. vestibular n., and central grey matter of the midbrain and pons, and so on. Furthermore, recent extensive studies revealed the presence of nerve terminals in the spinal cord, thalamus, median eminence, posterior pituitary, med. geniculate n., superior colliculus, n. mesencephalic trigeminal nerve,retrochiasmatic area, and so on. In addition to histamine, the neuron system contained GABA, adenosine and galanin, a new peptide found in the intestine, as cotransmitters. On electron microscopic analysis, few typical synaptic formation was found in the varicosities of the neur … More on system, though they contained synaptic vesicles, suggesting the non-synaptic release of histamine. The antibody immunochemically cross-reacted with guinea-pig DOPA decarboxylase but did not with the rat enzyme, indicating that there are some close evolutional relationship among aromatic amino acid decarboxylases. Histamine N-methyltransferase, which is important in the inactivation of histamine released into the synaptic cleft, was purified from rat kidneys. The antibody raised against it showed a positive reaction toward rat kedney (proximal tubules were stained) but no reaction toward the brain. Currently, the monoclonal antibody is being obtained. Histamine <H_1> -receptor protein was solubilized by the mixture of CHAPS-Tween 60-glycerol from rat liver, which contained 100-fold higher [ <^3H> ]-mepyramine binding capacity than the brain, and purified by a series of column chromatographies. The affinity chromatography with diphenhydramine or mepyramine as a ligand seems to be promising for further purification. The molecular weight was determined to be 152 K by target size analysis. Less

以组胺形成酶组氨酸脱羧酶为标记物，采用荧光免疫组化方法对大鼠脑内的组胺神经元系统进行了研究。细胞体仅存在于下丘脑后部和乳状体，如乳状管核，多曲张的轴突在下丘脑、嗅球、大脑皮层、内侧杏仁核、中前庭核、中脑和脑桥中央灰质等部位尤为密集。此外，最近的广泛研究表明，脊髓、丘脑、正中隆起、垂体后叶、膝状中丘、上丘、中脑三叉神经、交叉后区等均存在神经末梢。除了组胺外，神经元系统还含有GABA、腺苷和甘丙氨酸（一种在肠道中发现的新肽）作为共递质。电镜分析显示，在大鼠的静脉曲张中，虽然含有突触囊泡，但没有发现典型的突触形成，提示组胺的非突触释放。该抗体与豚鼠多巴脱羧酶存在免疫化学交叉反应，但与大鼠多巴脱羧酶无交叉反应，说明芳香氨基酸脱羧酶与豚鼠多巴脱羧酶之间存在密切的进化关系。从大鼠肾脏中纯化了组胺n -甲基转移酶，该酶在释放到突触间隙的组胺失活中起重要作用。对其产生的抗体对大鼠肾（近端小管染色）有阳性反应，但对大鼠脑无反应。目前正在获得单克隆抗体。用大鼠肝脏CHAPS-Tween 60-甘油混合溶液溶解组胺<H_1> -受体蛋白，该蛋白的[<^3H>]-甲锥胺结合能力比脑高100倍，并通过一系列柱层析纯化。以苯海拉明或甲皮拉明为配体的亲和层析法似乎有进一步纯化的前景。通过靶尺寸分析确定其分子量为152 K。少