CHEMICAL SYNAPSES, BIOPHYSICAL STUDIES

这个项目涉及神经递质受体的功能， 直接选通离子通道。核酸技术将被用于 结合电生理测量(电压钳、贴片 Clip)来测试关于特定多肽结构域的作用的假设 以及激动剂结合和通道门控中的单个氨基酸。这个 分子生物学操作将使用cdna克隆来 受体；将构建这些克隆的定点突变。 可用的酶方法将被用来转录正常和 变异的克隆人。将通过微量注射或通过 膜融合到非洲爪哇卵母细胞或哺乳动物细胞的培养中。这个 靶细胞将在细胞膜上表达受体； 功能离子通道将进行电生理学研究。这项工作 将包括Torpedo Electric的烟碱型乙酰胆碱受体 器官(现已有克隆，正在研究中)和小鼠的受体 肌肉(克隆人预计很快就会出现)。 类似的研究将继续进行的GABA受体-通道复合体 老鼠和小鸡的大脑。将继续进行检测分离的mRNA的工作 表达GABA通道的制备及cDNA克隆的筛选 由活性馏分组成的。当测序的克隆 Available，电压钳和膜片钳具体分析 构建的突变通道将开始。

This project concerns the function of neurotransmitter receptors that directly gate ion channels. Nucleic acid techniques will be used in conjunction with electrophysiological measurements (voltage clamp, patch clamp) to test hypotheses about the role of specific polypeptide domains and of individual amino acids in agonist binding and channel gating. The molecular biological manipulations will employ cDNA clones for the receptors; site-specific mutations of these clones will be constructed. Available enzymatic methods will be used to transcribe the normal and mutated clones. The mRNA will be introduced by microinjection or by membrane fusion into Xenopus oocytes or mammalian cells in culture. The target cells will express the receptors in the cell membrane; the functional ion channels will be studied electrophysiologically. The work will include both the nicotinic acetylcholine receptor of Torpedo electric organ (clones now available and under study) and the receptor from mouse muscle (clones expected soon). Analogous studies will be pursued on the GABA receptor-channel complex of rat and chick brain. Work will continue on assaying fractionated mRNA preparations that express GABA channels and on screening cDNA clones constructed from the active fractions. When sequenced clones are available, voltage-clamp and patch-clamp analysis of specifically constructed mutant channels will begin.