Department of Ophthalmology & Visual Science and Department of Cellular & Molecular Physiology, Yale University School of Medicine, New Haven, Connecticut 06511, USA.
J Neurosci. 2013 Jul 3;33(27):10972-85. doi: 10.1523/JNEUROSCI.1241-13.2013.
Alpha/Y-type retinal ganglion cells encode visual information with a receptive field composed of nonlinear subunits. This nonlinear subunit structure enhances sensitivity to patterns composed of high spatial frequencies. The Y-cell's subunits are the presynaptic bipolar cells, but the mechanism for the nonlinearity remains incompletely understood. We investigated the synaptic basis of the subunit nonlinearity by combining whole-cell recording of mouse Y-type ganglion cells with two-photon fluorescence imaging of a glutamate sensor (iGluSnFR) expressed on their dendrites and throughout the inner plexiform layer. A control experiment designed to assess iGluSnFR's dynamic range showed that fluorescence responses from Y-cell dendrites increased proportionally with simultaneously recorded excitatory current. Spatial resolution was sufficient to readily resolve independent release at intermingled ON and OFF bipolar terminals. iGluSnFR responses at Y-cell dendrites showed strong surround inhibition, reflecting receptive field properties of presynaptic release sites. Responses to spatial patterns located the origin of the Y-cell nonlinearity to the bipolar cell output, after the stage of spatial integration. The underlying mechanism differed between OFF and ON pathways: OFF synapses showed transient release and strong rectification, whereas ON synapses showed relatively sustained release and weak rectification. At ON synapses, the combination of fast release onset with slower release offset explained the nonlinear response of the postsynaptic ganglion cell. Imaging throughout the inner plexiform layer, we found transient, rectified release at the central-most levels, with increasingly sustained release near the borders. By visualizing glutamate release in real time, iGluSnFR provides a powerful tool for characterizing glutamate synapses in intact neural circuits.
α/Y 型视网膜神经节细胞通过由非线性亚基组成的感受野对视觉信息进行编码。这种非线性亚基结构增强了对高空间频率模式的敏感性。Y 细胞的亚基是突触前双极细胞，但非线性的机制仍不完全清楚。我们通过结合小鼠 Y 型神经节细胞的全细胞记录和在其树突和整个内丛状层表达的谷氨酸传感器（iGluSnFR）的双光子荧光成像，研究了亚基非线性的突触基础。为评估 iGluSnFR 的动态范围而设计的对照实验表明，Y 细胞树突的荧光反应与同时记录的兴奋性电流成正比增加。空间分辨率足以轻易分辨出混合的 ON 和 OFF 双极末端的独立释放。iGluSnFR 在 Y 细胞树突上的反应表现出强烈的环绕抑制，反映了突触前释放位点的感受野特性。对位于空间模式的反应将 Y 细胞非线性的起源定位在空间整合后的双极细胞输出。OFF 和 ON 通路的潜在机制不同：OFF 突触表现出短暂释放和强烈的整流，而 ON 突触表现出相对持续的释放和较弱的整流。在 ON 突触上，快速释放起始与较慢释放结束的组合解释了突触后神经节细胞的非线性反应。通过对整个内丛状层的成像，我们发现在最中心水平有短暂、整流的释放，而在边界附近释放越来越持续。通过实时可视化谷氨酸释放，iGluSnFR 为在完整神经回路中表征谷氨酸突触提供了一种强大的工具。
