Establishment of a method how to calculate the absolute number of neurotransmitter receptors acuroadiogaphically imaged

体外宏观自显影是测定神经递质受体在中枢神经系统各离散区分布的最佳和唯一方法。本研究的目的是建立一种利用氚化放射性标准计算大脑放射自显像中受体绝对数量的方法。实际上，标准化是通过结合3H-QNB来完成的，3H-QNB是一种麝香碱乙酰胆碱受体的拮抗剂。制备正常大鼠脑匀浆，部分isa与^3H-QNB孵育，计算以mol/mg蛋白表达的结合。将剩余的孵育液离心，得到的微球按普通的显微放射自显影工艺涂在h敏感膜上。利用蔡司公司的II计算机系统，测定了自射线照相光密度与放射性的关系。作为结果，我们注意到，当转换为In-In尺度时，光密度和组织结合放射性之间存在线性关系。通过这些实验，我们获得了体外放射自显影计算的受体绝对数量与脑匀浆结合实验的非常好的一致性。这意味着我们制备的标准品质量良好，足以作为定量体外放射自显影的标准品。通过我们建立的这项技术，我们检测了mptp诱导的猴子帕金森病、人类脊髓小脑变性和阿尔茨海默病以及这些神经系统疾病的动物模型中各种神经递质受体的分布。此外，我们还观察了大鼠脑中麝香碱乙酰胆碱受体亚型和中性紧张素受体的致瘤发展。通过这些实验，证实了体外定量自图术是一种可靠且广泛应用于神经科学的实验技术。

in vitro Macro-autoradigraphy is the best and only method to determine distrbution of neurotransmitter receptors in each discete region within the central nervous system. The purpose of this research is to establish the method to calculate the absolute number of receptors in autoradiographic images of the brain using tritiated rediactive standards. Actually, Standaedizartion was done by binding of 3H-QNB which is an antabonist of muscatineic acetylcholine receptor. Homogenate of the normal rat brain was prepared adn part of isa was incubated with^3H-QNB to calculate the binding expressed as mol/mg protein. The rest of incubated solution was centriguged and the obtained pellet was applied to a H-sensitive film according to the ordinary process of macro-autoradiography. We utilized the IBAS II computer system of Zeiss company to determine relationship between autoradiographic optical density and radioactivity. As the results, we noticed that there was a linewar relationship between optical density and tissue-bound radioactivity when converted to a In-In scale. With these experiments, we obrained a very good consistence between the absolute number of receptore calculated from in vitro autoradioragraphy and that from brain homogenate binding experiments. This means that the standard prepared by us has a good quality enough to use as a standard of quantitative in vitro autoradiography. Through this rechnique esteblished by us, we examined distributions of various neurotransmitter receptors in the brains of MPTP-induced monkey parkinsonism, human cases of spinocerebellar degenetation and Alzheimer disease, and animal models of these neurological desorders. Besids, ontogenic developments of muscatinic acetylcholine receptor subtypes and neutotensin receptors in rat brains were examined. Throught these experiments, is was confirmed thast quantitative in vitro autoradigrraphy is a reliable and widely applicable experimental technique in neuroscience.