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Molecular cytobiological study on hippocampal LTP and LTD

海马LTP和LTD的分子细胞生物学研究

基本信息

批准号：
09044324
负责人：
MIYAMOTO Eishichi
金额：
$ 4.1万
依托单位：
Kumamoto University
依托单位国家：
日本
项目类别：
Grant-in-Aid for international Scientific Research
财政年份：
1997
资助国家：
日本
起止时间：
1997 至 1998
项目状态：
已结题

项目摘要

The purposes of the study is to elucidate the mechanisms of learning and memory of animals on the basis of molecular cytobiological methodology. When electrical high frequency stimulation (HFS) is given to synapses, potentiation of synaptic activities lasts for several hours or longer. This phenomenon is called long-term potentiation(LTP) and considered to be a model for learning and memory of animals. The elucidation of the mechanisms of LTP will give a clue to understand the higher activity of the brain. It has been recently reported that protein phosphorylatiorr especially Ca^2/calmodulin-dependent protein kinaseII (CaM kinase II) is involved in induction and maintenance of LTP in the hippocampus. In an attempt to examine the molecular mechanisms of LTP in the hippocampus, we analyzed LTP.1)Activation of CaM kinase II and mitogen-activated protein kinase (MAP kinase) was analyzed during LTP induction by the in-gel assay with myelin basic protein as a substrate. MAP kinase was activated during LTP induction. The peak of the activation was observed 3 min after HFS and rapidly decreased to the original level within 10 mm. When 50 muM PD098059, a MAP kinase kinase inhibitor, was previously given to hippocampal slices, LTP induction and activation of both CaM kinase II and MAP kinase were inhibited. However, addition of 30 muM PD098059 to slices did not affect both LTP and activation of CaM kinase II but did inhibit activation of MAP kinase. These results indication that LTP induction is correlated with activation of CaM kinase II, but not with activation of MAP kinase.2)Calyculin A-sensitive protein phosphatase (PP) activity, which includes PP1 and PP2 was inhibited during LTP induction. The autoradiography and immunoblot analyses showed that phosphorylation of the regulatory subunit of PP2A is phosphorylated with a concomitant decrease in PP activity.

该研究的目的是在分子细胞生物学方法的基础上阐明动物学习和记忆的机制。当对突触进行高频电刺激（HFS）时，突触活动的增强会持续几个小时或更长时间。这种现象称为长时程增强（LTP），被认为是动物学习和记忆的模型。 LTP机制的阐明将为理解大脑的高级活动提供线索。最近报道蛋白质磷酸化尤其是Ca^2/钙调蛋白依赖性蛋白激酶II(CaM激酶II)参与海马LTP的诱导和维持。为了研究海马LTP的分子机制，我们分析了LTP。1)以髓磷脂碱性蛋白为底物，通过凝胶内测定，在LTP诱导过程中分析了CaM激酶II和丝裂原激活蛋白激酶（MAP激酶）的激活。 MAP 激酶在 LTP 诱导过程中被激活。 HFS 后 3 分钟观察到激活峰值，并在 10 mm 内迅速降至原始水平。当先前将 50 muM PD098059（一种 MAP 激酶激酶抑制剂）给予海马切片时，LTP 诱导以及 CaM 激酶 II 和 MAP 激酶的激活均受到抑制。然而，向切片中添加 30 muM PD098059 不会影响 LTP 和 CaM 激酶 II 的激活，但会抑制 MAP 激酶的激活。这些结果表明LTP诱导与CaM激酶II的激活相关，但与MAP激酶的激活无关。2)Calyculin A敏感的蛋白磷酸酶(PP)活性，包括PP1和PP2在LTP诱导过程中受到抑制。放射自显影和免疫印迹分析表明，PP2A 调节亚基的磷酸化伴随着 PP 活性的降低。