INTRACELLULAR MESSENGER FUNCTIONS OF INOSITOL PHOSPHATES

肌醇磷酸盐被用作细胞内的信使 用于控制许多不同细胞的信号通路 功能，如神经递质和激素反应， 分泌、肌肉收缩和光传导。精神障碍 这一信号通路的一部分与疾病有关 包括肿瘤发生和躁郁症，以及 这一系统与临床研究的相关性几乎可以肯定 随着我们对它的了解越来越多，它就会成长。有了很大的进步 最近在生物化学解释中做出了关于 导致几种不同肌醇形成的途径 磷酸盐化合物。然而，对老年人的生理学研究 这些化合物的作用没有那么先进，尽管它 认为它们的作用主要是通过提高细胞内游离 钙水平，对许多人的具体作用知之甚少 或者关于它们调节钙的机制 跨细胞内膜和细胞表面膜的通量。主 这项建议的目的是研究信使功能 肌醇磷酸盐，通过使用电生理和光学 膜电流和细胞内电流的测量技术 钙。非洲爪哇卵母细胞将作为一种方便的模型细胞 系统，因为这些细胞具有很好的磷脂酰肌醇特性 信令系统，其庞大的体积方便了许多程序 包括电压钳记录和微量注射。 最初的目的是表征不同肌醇的能力。 磷酸盐释放细胞内钙，激活钙离子内流 钙穿过质膜，并激活细胞膜 与钙无关的电导。随后， 肌醇磷酸盐激活的钙通道的特性将是 通过电压和膜片钳记录详细检查 穿过质膜的电流，并通过重组 从内膜到脂质双层的通道。这将是 给出关于单分子的动力学和电导的信息 肌醇磷酸盐对通道的调节及其离子特异性 和其他第二信使，并被钙拮抗剂阻断 和其他药理药剂。类似的研究也将在 由钙激活的氯离子通道组成，介导 肌醇磷脂激活后的最终电反应 卵母细胞，以及任何已发现的钙非依赖性电流 被肌醇磷酸盐调节。录音带 细胞内钙离子将被用来确定细胞内的 肌醇磷酸盐对膜振荡电流的响应 产生于钙的振荡释放，反馈 负责这一过程的机制将被研究。这个 卵母细胞也将被用作外源基因的翻译系统 信使核糖核酸，看看钙通道或其他成分 磷脂酰肌醇信号系统，可以在功能上 通过大脑或唾液腺的mRNA表达。

Inositol phosphates are used as intracellular messengers within a signalling pathway which serves to control many diverse cellular functions, such as neurotransmitter and hormone responses, secretion, muscle contraction and phototransduction. Disorders of this signalling pathway have been implicated in diseases including tumorigenesis and manic depressive illness, and the relevance of this system to clinical studies will almost certainly grow as we come to understand it more. Great progress has recently been made in the biochemical elucidation of the pathways leading to the formation of several different inositol phosphate compounds. However, the physiological study of the actions of these compounds is less well advanced, and although it is thought that they act primarily by raising intracellular free calcium levels, little is known about the specific actions of many of them, or about the mechanisms by which they regulate calcium fluxes across intracellular and cell surface membranes. The main object of this proposal is to study the messenger functions of inositol phosphates, by using electrophysiological and optical techniques to measure membrane currents and intracellular calcium. Xenopus oocytes will be used as a convenient model cell system, as these cells have a well characterized phosphoinositide signalling system, and their large size facilitates many procedures including voltage-clamp recording and micro-injection. The initial aim is to characterize the abilities of different inositol phosphates to liberate intracellular calcium, to activate influx of calcium across the plasma membrane, and to activate membrane conductances independent of calcium. Subsequently, the properties of inositol phosphateactivated calcium channels will be examined in detail by voltage- and patch-clamp recording of currents across the plasma membrane, and by reconstituting channels from internal membranes into lipid bilayers. This will give information about the kinetics and conductances of single channels, their ionic specificity, modulation by inositol phosphates and other second messengers, and blocking by calcium antagonists and other pharmacological agents. Similar studies will also be made of the calcium-activated chloride channels which mediate the final electrical response to phosphoinositide activation in the oocyte, and of any calcium-independent currents which are found to be modulated by inositol phosphates. Recordings of intracellular calcium will be made to determine whether the oscillatory membrane current response to inositol phosphates arises from an oscillatory liberation of calcium, and the feedback mechanism responsible for this process will be studied. The oocyte will also be used as a translation system for exogenous mRNA, to see whether calcium channels, or other components of the phosphoinositide signalling system, can be functionally expressed by mRNA from brain or salivary gland.