Cell Biology of the Retina

描述（由申请人提供）：我们的研究目的是了解哺乳动物的视网膜如何分析视觉世界，并将其空间，时间和色彩对比编码为发送给大脑的动作电位信息。在视网膜中，多巴胺是导致神经适应光线的许多事件的调节器。为了确定控制多巴胺释放的机制和神经网络，我们在小鼠中标记了遗传多巴胺能无分泌细胞（DA细胞），从而可以在体外研究视网膜分离后的生理学。我们还开发了一种技术来研究单个神经元的整体基因表达。目前的应用有以下目的：(i)我们将测试DA细胞除了多巴胺外，还通过胞外作用在其整个表面和与AII无突细胞的突触释放GABA的假设，AII无突细胞是将杆状信号传递给锥体双极细胞的神经元。（ii）我们将通过完整小鼠视网膜的细胞内记录来研究DA细胞的光反应。（iii）通过在覆盖转录小鼠基因组的高密度寡核苷酸探针阵列上分析基因表达，我们将编制一份完整的DA细胞转录本清单。了解DA细胞中包含的离子通道、传递受体、g蛋白偶联第二信使通路组分和分泌的神经活性分子，将使我们能够对新功能提出有意义的假设，这些假设可以通过实验进行测试。最后，我们将比较DA细胞的转录组与另外两种视网膜神经元的转录组，2型儿茶酚胺能神经元和杆状双极神经元。除了新分子的存在，我们还将寻找神经元细胞类型概念的核心问题的答案：我们希望揭示细胞类型特定的分子，并确定三种类型的神经元在处理光信号时所执行的独特功能的“分子特征”。（iv）我们将通过电子显微镜鉴定在光照和黑暗条件下控制DA细胞活动的神经元。这些研究对于理解视网膜的视觉机制至关重要。

DESCRIPTION (provided by applicant): The objective of our research is to understand how the retina of mammals analyzes the visual world and encodes its spatial, temporal and chromatic contrast into a message of action potentials which is sent to the brain. In the retina, dopamine is the modulator responsible for many of the events that lead to neural adaptation to light. To identify the mechanisms and neural networks that control the release of dopamine, we labeled genetically dopaminergic amacrine cells (DA cells) in the mouse and could therefore investigate their physiology in vitro after dissociation of the retina. We also developed a technique to study global gene expression in single neurons. The present application has the following aims: (i) we will test the hypothesis that DA cells, in addition to dopamine, release GABA by exocytosis both over their entire surface and at their synapses with AII amacrine cells, the neuron that transfer rod signals to cone bipolar cells. (ii) We will investigate the light responses of DA cells by intracellular recordings in the intact mouse retina. (iii) By profiling gene expression on high-density arrays of oligonucleotide probes covering the transcribed mouse genome, we will compile a comprehensive list of the transcripts contained in DA cells. Knowledge of the repertory of ion channels, transmitter receptors, components of G-protein-coupled second messenger pathways and secreted neuroactive molecules that are contained in DA cells will allow us to formulate meaningful hypotheses about novel functions that can be tested experimentally. Finally, we will compare the transcriptome of DA cells with that of two other types of retinal neurons, type 2 catecholaminergic amacrines and rod bipolars. In addition to the presence of novel molecules, we will be searching for answers to questions that are at the very heart of the concept neuronal cell type: we want to uncover cell-type specific molecules and identify a "molecular signature" for the unique functions that are carried out by the three types of neurons in the processing of light signals. (iv) We will identify by electron microscopy the neurons that control the activity of DA cells in light and darkness. These studies are crucial to the understanding of the mechanism of vision in the retina.