Synaptic Mechanisms in the Mammalian Retina

哺乳动物视网膜的突触机制

• 批准号: 9157532
• 负责人: JEFFREY S DIAMOND
• 金额: $ 122.8万
• 依托单位: NATIONAL INSTITUTE OF NEUROLOGICAL DISORDERS AND STROKE
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/9157532
• 关键词: Address; Amacrine Cells; Behavior; Biological Neural Networks; Calcium Channel; Cells; Characteristics; Collaborations; Communication; Cone; Coupled; Development; Diamond; Docking; Electron Microscopy; Feedback; Filament; Freezing; Glutamate Receptor; Goals; Image; Inner Plexiform Layer; Manuscripts; Mediating; Membrane; Mus; N-Methyl-D-Aspartate Receptors; Neurons; Output; Pathway interactions; Photons; Physiological; Physiology; Presynaptic Terminals; Process; Property; Proteins; Publications; Resolution; Retina; Retinal; Retinal Ganglion Cells; Shapes; Signal Transduction; Specificity; Staging; Stem cells; Structure; Synapses; Synaptic Transmission; Synaptic Vesicles; Techniques; Time; Tissues; Vesicle; Visual; Work; absorption; cell type; ganglion cell; luminance; postsynaptic; pressure; presynaptic; prevent; reconstruction; research study; response; retinal rods; ribbon synapse; signal processing; synaptic function; tomography; vesicular release; visual process; visual processing; voltage

Our work focuses on specialized synapses in the inner retina. We have expanded our study of inhibitory synaptic connections made by amacrine cells within the inner retina, to understand how feedforward and feedback inhibition contributes to signal processing in this network. We previously discovered that A17 amacrine cells provide rapid GABAergic feedback to rod bipolar cell ribbon synapses via a release process that is independent of membrane depolarization or voltage-gated calcium channels (Chavez, et al., 2006). This rapid feedback may be essential to prevent the rapid depletion of readily-releasable vesicles from the rod bipolar cell synaptic terminal (Singer and Diamond, 2006). Our most recent work indicates that reciprocal feedback from A17s extends the range over which these synapses encode luminance and compute contrast (Oesch and Diamond, submitted). Feedback from other amacrine cells weakly modulates the synaptic gain but does not change the operating range. We also find that A17-mediated feedback inhibition enhances the gain of synaptic responses in the rod pathway to the absorption of single photons (Grimes, et al., 2015). Our understanding of ribbon synaptic physiology in bipolar cells is limited to rod bipolar cells. We don't know as much about synaptic transmission from cone bipolar cells, because it is very difficult to obtain synaptically coupled cone bipolar - ganglion cell pairs. We have crossed mouse lines with genetically encoded markers identifying specific types of bipolar and ganglion cells that are very likely to be connected. In particular, we have utilized mouse lines in which type two cone bipolar cells (CBC2) can be visualized. We find that CBC2s make reciprocal synaptic connections with AII amacrine cells. We are studying bidirectional communication between these two cell types, and we are also studying how the rod bipolar cell signal is shaped by the AII amacrine cell before it is passed to the CBC2. Our preliminary findings suggest that AIIs preferentially transmit information about contrast, but not luminance, to CBCs. We are examining the cellular and synaptic processes that underlie this transformation. We plan to expand these experiments to examine release from ON CBCs and their postsynaptic partners in an effort to compare the dynamics of vesicle release from ON and OFF bipolar cells. We have also collaborated with Donald Zack (Johns Hopkins) to examine the physiological characteristics of retinal ganglion cells derived from stem cells. We find that these cells develop divers firing properties that are consistent with different ganglion cell subtypes at different stages of development. We also found that ESC-derived ganglion cells express functional glutamate receptors. A revised manuscript has been submitted for publication. Finally, we are extending our electron microscopy studies, in collaboration with Tom Reese and Richard Leapman, to explore the detailed ultrastructure of synaptic ribbons in photoreceptors and rod bipolar cells. So far, EM tomography enables us to detect protein filaments that tether synaptic vesicles to the ribbon and the presynaptic membrane. This approach may enable us to discern morphologically, for the first time, docked and primed synaptic vesicles. We have obtained detailed reconstructions of entire, intact ribbons (Graydon, et al., 2014) and are now working to obtain higher resolution images to examine quantitatively the tethers that connect synaptic vesicles to the ribbon and to the presynaptic membrane. To this end, we have recently obtained high-resolution images of ribbon synapses using high-pressure freezing EM techniques. This approach has been used to great effect in cultured mammalian neurons, but it has proved quite difficult in intact tissue. A great deal of effort has finally enabled us to obtain high-quality freezing the in the inner plexiform layer of intact retina, which will enable us to address numerous questions of synaptic function.

我们的工作集中在视网膜内层的特殊突触上。 我们已经扩大了我们的研究抑制性突触连接的无长突细胞内视网膜，了解前馈和反馈抑制如何有助于在这个网络中的信号处理。 我们先前发现，A17无长突细胞通过独立于膜去极化或电压门控钙通道的释放过程向杆状双极细胞带状突触提供快速GABA能反馈（Chavez，et al.，2006年）。 这种快速反馈对于防止杆双极细胞突触末端的易释放囊泡的快速耗尽可能是必不可少的（Singer和Diamond，2006）。 我们最近的工作表明，来自A17的相互反馈扩展了这些突触编码亮度和计算对比度的范围（Oesch和Diamond，提交）。 来自其他无长突细胞的反馈微弱地调节突触增益，但不改变操作范围。 我们还发现，A17介导的反馈抑制增强了视杆通路中对单光子吸收的突触反应的增益（Grimes，et al.，2015年）。 我们对双极细胞中带状突触生理学的理解仅限于杆状双极细胞。我们对来自锥双极细胞的突触传递了解不多，因为很难获得突触耦合的锥双极-神经节细胞对。我们已经将小鼠品系与遗传编码标记进行了杂交，这些标记识别了极有可能连接的特定类型的双极细胞和神经节细胞。特别地，我们已经利用了小鼠品系，其中可以可视化2型锥双极细胞（CBC 2）。我们发现CBC 2与AII无长突细胞形成相互的突触连接。我们正在研究这两种细胞类型之间的双向通讯，我们也在研究视杆双极细胞信号在传递到CBC 2之前是如何被AII无长突细胞塑造的。我们的初步研究结果表明，AIIs优先传输信息的对比度，而不是亮度，CBCs。我们正在研究这种转变背后的细胞和突触过程。 我们计划扩大这些实验，以检查从ON CBCs和它们的突触后伙伴的释放，努力比较从ON和OFF双极细胞释放囊泡的动力学。 我们还与唐纳德·扎克（约翰霍普金斯）合作，研究干细胞衍生的视网膜神经节细胞的生理特征。 我们发现，这些细胞开发潜水员发射特性是一致的不同的神经节细胞亚型在不同的发展阶段。我们还发现，ESC衍生的神经节细胞表达功能性谷氨酸受体。修订稿已提交出版。 最后，我们正在与Tom Reese和Richard Leapman合作，扩展我们的电子显微镜研究，以探索光感受器和视杆双极细胞中突触带的详细超微结构。 到目前为止，电磁断层扫描使我们能够检测到蛋白质丝，这些蛋白质丝将突触囊泡拴在丝带和突触前膜上。 这种方法可能使我们能够从形态学上第一次辨别对接和启动的突触囊泡。 我们已经获得了完整的完整条带的详细重建（Graydon等人，2014年），现在正在努力获得更高分辨率的图像，以定量检查连接突触囊泡到丝带和突触前膜的系绳。为此，我们最近获得了高分辨率图像的带状突触使用高压冷冻EM技术。这种方法已经在培养的哺乳动物神经元中取得了很大的效果，但在完整的组织中却被证明是相当困难的。大量的努力终于使我们能够获得高质量的冷冻完整视网膜的内丛状层，这将使我们能够解决突触功能的许多问题。