Animal Physiology Group, Department of Biology, University of Kaiserslautern, Erwin Schrödinger-Straße 13, D 67663 Kaiserslautern, Germany.
Physiology of Neuronal Networks Group, Department of Biology, University of Kaiserslautern, Erwin Schrödinger-Straße 13, D 67663 Kaiserslautern, Germany.
Int J Mol Sci. 2019 Jun 10;20(11):2821. doi: 10.3390/ijms20112821.
Anisotropy of tracer-coupled networks is a hallmark in many brain regions. In the past, the topography of these networks was analyzed using various approaches, which focused on different aspects, e.g., position, tracer signal, or direction of coupled cells. Here, we developed a vector-based method to analyze the extent and preferential direction of tracer spreading. As a model region, we chose the lateral superior olive-a nucleus that exhibits specialized network topography. In acute slices, sulforhodamine 101-positive astrocytes were patch-clamped and dialyzed with the GJ-permeable tracer neurobiotin, which was subsequently labeled with avidin alexa fluor 488. A predetermined threshold was used to differentiate between tracer-coupled and tracer-uncoupled cells. Tracer extent was calculated from the vector means of tracer-coupled cells in four 90° sectors. We then computed the preferential direction using a rotating coordinate system and post hoc fitting of these results with a sinusoidal function. The new method allows for an objective analysis of tracer spreading that provides information about shape and orientation of GJ networks. We expect this approach to become a vital tool for the analysis of coupling anisotropy in many brain regions.
示踪剂偶联网络的各向异性是许多脑区的一个标志。过去，这些网络的拓扑结构使用各种方法进行了分析，这些方法侧重于不同的方面，例如，偶联细胞的位置、示踪信号或方向。在这里，我们开发了一种基于向量的方法来分析示踪剂扩散的程度和优先方向。作为一个模型区域，我们选择了外侧上橄榄核，这是一个表现出特殊网络拓扑结构的核团。在急性切片中，用 GJ 通透性示踪剂神经生物素对 Sulforhodamine 101 阳性星形胶质细胞进行了膜片钳记录和灌流，随后用亲和素 Alexa Fluor 488 进行了标记。使用预定的阈值将示踪剂偶联细胞与未偶联细胞区分开来。示踪剂范围是通过在四个 90°扇区中对示踪剂偶联细胞的向量平均值计算得出的。然后，我们使用旋转坐标系计算优先方向，并对这些结果进行事后拟合正弦函数。这种新方法可以对示踪剂扩散进行客观分析，提供关于 GJ 网络形状和方向的信息。我们预计这种方法将成为分析许多脑区偶联各向异性的重要工具。
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