Rapid and Sensitive Mapping of Long-Range Connections In Vitro Using Flavoprotein Autofluorescence Imaging Combined With Laser Photostimulation
Rapid and Sensitive Mapping of Long-Range Connections In Vitro Using Flavoprotein Autofluorescence Imaging Combined With Laser Photostimulation
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DOI：
10.1152/jn.91291.2008
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发表时间：
2009-06-01
影响因子：
2.5
通讯作者：
Issa, N. P.
中科院分区：
文献类型：
--
作者：
Llano, D. A.;Theyel, B. B.;Issa, N. P.
拉诺 DA、泰塞尔 BB、马利克 AK、谢尔曼 SM、伊萨 NP。使用黄素蛋白自发荧光成像与激光光刺激相结合，快速、灵敏地绘制体外长程连接图。 《神经生理学杂志》101：3325-3340，2009 年。首次发表于 2009 年 3 月 25 日； doi：10.1152/jn.91291.2008。我们研究了使用黄素蛋白自发荧光 (FA) 作为绘制长程神经连接的工具，并将 FA 与谷氨酸的激光解笼结合起来，以促进体外快速长程绘制。使用体感丘脑皮质切片，我们确定 FA 的空间分辨率 >= 100-200 μm，并且检测丘脑皮质突触活动的灵敏度接近全细胞记录的灵敏度。用 DNQX 和 AP5 阻断离子型谷氨酸受体，消除了皮质对丘脑电刺激的反应。 FA 与使用笼状谷氨酸的光刺激相结合，揭示了强大的长距离连接模式，可以在包含丘脑皮质、皮质丘脑或皮质皮质连接的体感、听觉和视觉系统的切片中轻松评估这些模式。我们绘制了从丘脑腹侧后核（VPM）到初级体感皮层桶区的投影，并确认了之前使用更费力的方法描述的地形。我们还制作了从 VPM 到丘脑网状核的投影的新颖图，显示了沿背腹轴的精确地形。重要的是，生成连接图（六个刺激位置）只需要大约 30 秒。这些数据表明 FA 是探索和测量连通性的敏感工具，当与谷氨酸光刺激结合时，可以快速绘制单个动物的远程投影图。
Llano DA, Theyel BB, Mallik AK, Sherman SM, Issa NP. Rapid and sensitive mapping of long-range connections in vitro using flavoprotein autofluorescence imaging combined with laser photostimulation. J Neurophysiol 101: 3325-3340, 2009. First published March 25, 2009; doi:10.1152/jn.91291.2008. We investigated the use of flavoprotein autofluorescence (FA) as a tool to map long-range neural connections and combined FA with laser-uncaging of glutamate to facilitate rapid long-range mapping in vitro. Using the somatosensory thalamocortical slice, we determined that the spatial resolution of FA is >= 100-200 mu m and that the sensitivity for detecting thalamocortical synaptic activity approximates that of whole cell recording. Blockade of ionotropic glutamate receptors with DNQX and AP5 abolished cortical responses to electrical thalamic stimulation. The combination of FA with photostimulation using caged glutamate revealed robust long-distance connectivity patterns that could be readily assessed in slices from the somatosensory, auditory, and visual systems that contained thalamocortical, corticothalamic, or corticocortical connections. We mapped the projection from the ventral posterior nucleus of thalamus (VPM) to the primary somatosensory cortex-barrel field and confirmed topography that had been previously described using more laborious methods. We also produced a novel map of the projections from the VPM to the thalamic reticular nucleus, showing precise topography along the dorsoventral axis. Importantly, only about 30 s were needed to generate the connectivity map (six stimulus locations). These data suggest that FA is a sensitive tool for exploring and measuring connectivity and, when coupled with glutamate photostimulation, can rapidly map long-range projections in a single animal.