GENETIC CONTROL OF MEMBRANE CURRENTS

这个项目的长期目标是确定 基因组决定了可兴奋细胞的膜电流。全 研究是电生理学的，使用吸力电极电压钳 技术(通常称为膜片钳技术，当 电极开口小于2微米)，允许进行非常详细的分析 膜的电学性质。三种方法将是 紧随其后。 第一种方法是寻找控制机制。敏感度 蜗牛神经元膜电流对细胞内信使的影响 将在分离的膜片中进行分析。特别是，一个 将对细胞内钙和氢的明显作用进行研究 阻断钙电流，激活钾电流和非特异性电流。角色 细胞内磷酸化同时控制钙电流和钾电流将 被调查。单通道质子流的大小将是 下定决心。 第二种方法侧重于基因组控制的动态方面。这个 养殖成螺膜兴奋性的变化 (Helisoma)神经元与果蝇胚胎神经和肌肉的发育 将对细胞进行研究。这些研究将确定 分离后的分化膜，随着时间的推移 未分化膜获得兴奋性，而胞体膜 与轴突生长相关的电流。 第三种方法将剖析膜的遗传控制因素。 通过表征由此导致的膜电流的变化来实现兴奋性 来自突变的基因。膜片钳研究将对果蝇神经和 由突变苍蝇卵制成的胚胎培养中的肌肉细胞。 有相当多的膜证据的行为突变体 缺陷，如振动器和Nap，将首先检查。 由于细胞内钙在调节递质中的重要性 释放、肌肉收缩和其他细胞功能，主要 将注意钙电流和重叠的钾电流，它们 测定细胞内钙离子的内流。已发现的机制或 由该项目阐明的将大大有助于 对涉及状态改变的神经和肌肉疾病的理解 膜兴奋性或突触效能。

The long-term goal of this project is to determine the mechanisms by which the genome determines the membranc currents of excitable cells. All studies are electrophysiological, employing suction-electrode voltage-clamp techniques (often referred to as "patch clamp" techniques when the electrode opening is less than 2Mum) that allow a higly detailed analysis of the electrical properties of the membrane. Three approaches will be followed. The first approach is a search for mechanisms of control. The sensitivity of snail (Lymnaea) neuron membrane currents to intracellular messengers will be analyzed in isolated patches of membrane. In particular, an examination will be made of the apparent roles of intracellular Ca2+ and H+ to block Ca current and to activate K and non-specific currents. The role of intracellular phosphorylation in controlling both Ca and K currents will be investigated. The size of single-channel proton currents will be determined. The second approach focuses on the dynamic aspect of genome control. The changes in membrane excitability that occur in cultured adult snail (Helisoma) neurons and developing embryonic Drosophila nerve and muscle cells will be studied. These studies will determine the stability of differentiated membranes after isolation, the time course with which the undifferentiated membrane gains its excitability, and the somal membrane currents associated with neurite growth. The third approach will dissect the elements of genetic control of membrane excitability by characterizing the changes in membrane currents that result from mutations. Patch clamp studies will be made on Drosophila nerve and muscle cells in embryo cultures made from the eggs of mutant flies. Behavioral mutants for which there is considerable evidence of a membrane defect, e.g. Shaker and Nap, will be examined first. Due to the importance of intracellular Ca2+ in controlling transmitter release, muscle contraction, and other cellular functions, primary attention will be given to the Ca current and overlapping K currents, which determine the influx of Ca2+ into the cell. Mechanisms discovered or elucidated by this project will contribute significantly to the understanding of neural and muscular diseases that involve altered states of membrane excitability or synaptic efficacy.