NEUROTRANSMITTER RELEASE

本项目拟研究光调制的神经递质 从分离的脊椎动物光感受器释放。 最近的证据表明 酸性氨基酸，谷氨酸，可能作为神经递质， 包括金鱼在内的许多物种的光感受器。 然而，在这方面， 光调节视细胞终末内源性谷氨酸的释放 尚未得到证实。 我们建议建造一个 用于谷氨酸检测的谷氨酸敏感电极 金鱼的光感受器释放出的光， 从视网膜分离出来。 描述了两种方法 这种电极的构造，两者都依赖于膜片钳 技术和含有谷氨酸受体的gigaseals的形成。 一种方法是使用生化分离的谷氨酸受体 另一种是从大脑中提取谷氨酸敏感的细胞， 孤立的水平细胞的膜。 灵敏度、选择性和 电极的稳定性将通过比较 产生的谷氨酸及其类似物对响应特性的影响 谷氨酸盐通道，即，它们的电导、平均开放时间和百分比 在封闭状态下度过的时间。 电极，放置在并置， 然后将使用感光器末端来确定（1）程度 内源性递质模仿谷氨酸对 通道特性，（2）光强度变化对 发射机释放，和（3）发射机释放的机制， 受控的，例如，钙依赖性、哇巴因敏感性等。 这项研究的结果将直接关系到突触的性质， 一般神经元的传递和视觉信息的处理 尤其是视网膜细胞。

The present project proposes to examine light-modulated neurotransmitter release from isolated vertebrate photoreceptors. Recent evidence suggests that the acidic amino acid, glutamate, may act as the neurotransmitter from photoreceptors in a variety of species, including goldfish. However, light-modulated release of endogenous glutamate from visual cell terminals has not yet been demonstrated. We propose to construct a glutamate-sensitive electrode to be used in the detection of glutamate release from goldfish photoreceptors that have been enzymatically dissociated from the retina. Two methods are described with respect to the construction of such electrodes, both of which rely on patch-clamp technology and the formation of gigaseals containing glutamate receptors. One method involves the use of biochemically isolated glutamate receptors from brain; the other employs outside-out patches from glutamate-sensitive membranes of isolated horizontal cells. The sensitivity, selectivity, and stability of the electrodes will be characterized by comparing the effects produced by glutamate and its analogues on the response properties of glutamate channels, i.e., their conductance, mean open time, and percentage time spent in the closed state. The electrodes, placed in apposition to the photoreceptor terminals, will then be used to determine (1) the degree to which the endogenous transmitter mimics the effects of glutamate on channel properties, (2) the effects of variation in photic intensity on transmitter release, and (3) the mechanisms whereby transmitter release is controlled, e.g., calcium dependence, ouabain sensitivity, etc. The results of this study will have direct bearing upon the nature of synaptic transmission by neurons in general and the processing of visual information by retinal cells in particular.