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t-GRASP，一种用于评估神经元连接的靶向 GRASP 方法。

t-GRASP, a targeted GRASP for assessing neuronal connectivity.

机构信息

Department of Cell Biology and Neuroscience, Montana State University Bozeman, MT 59717, United States.

Department of Cell Biology and Neuroscience, Montana State University Bozeman, MT 59717, United States.

出版信息

J Neurosci Methods. 2018 Aug 1;306:94-102. doi: 10.1016/j.jneumeth.2018.05.014. Epub 2018 May 21.

DOI:10.1016/j.jneumeth.2018.05.014
PMID:29792886
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC6689385/
Abstract

BACKGROUND

Understanding how behaviors are generated by neural circuits requires knowledge of the synaptic connections between the composite neurons. Methods for mapping synaptic connections, such as electron microscopy and paired recordings, are labor intensive and alternative methods are thus desirable.

NEW METHOD

Development of a targeted GFP Reconstitution Across Synaptic Partners(GRASP) method, t-GRASP, for assessing neural connectivity is described.

RESULTS

Numerous different pre-synaptic and post-synaptic/dendritic proteins were tested for enhancing the specificity of GRASP signal to synaptic regions. Pairing of both targeted pre- and post-t-GRASP constructs resulted in strong preferential GRASP signal in synaptic regions in Drosophila larval sensory neurons, larval neuromuscular junctions, and adult photoreceptor neurons with minimal false-positive signal.

COMPARISON WITH EXISTING METHODS

Activity-independent t-GRASP exhibits an enhancement of GRASP signal specificity for synaptic contact sites as compared to existing Drosophila GRASP methods. Fly strains were developed for expression of both pre- and post-t-GRASP with each of the three Drosophila binary transcription systems, thus enabling GRASP assays to be performed between any two driver pairs of any transcription system in either direction, an option not available for existing Drosophila GRASP methods.

CONCLUSIONS

t-GRASP is a novel targeted GRASP method for assessing synaptic connectivity between Drosophila neurons. Its flexibility of use with all three Drosophila binary transcription systems significantly expands the potential use of GRASP in Drosophila.

摘要

背景

了解神经网络中的行为是如何产生的，需要了解复合神经元之间的突触连接。映射突触连接的方法，如电子显微镜和配对记录，劳动强度大，因此需要替代方法。

新方法

描述了一种用于评估神经连接的靶向 GFP 重组跨越突触伙伴（GRASP）方法 t-GRASP 的开发。

结果

测试了许多不同的突触前和突触后/树突蛋白，以增强 GRASP 信号对突触区域的特异性。靶向前和后 t-GRASP 构建体的配对导致在果蝇幼虫感觉神经元、幼虫神经肌肉接头和成年光感受器神经元中的突触区域中出现强烈的优先 GRASP 信号，而假阳性信号最小。

与现有方法的比较

与现有的果蝇 GRASP 方法相比，无活性的 t-GRASP 表现出对突触接触位点的 GRASP 信号特异性的增强。开发了用于表达前和后 t-GRASP 的双等位基因菌株，每个菌株都有三种果蝇双转录系统之一，从而使 GRASP 测定能够在任何两个驱动对之间以任何方向进行，这是现有果蝇 GRASP 方法所没有的选项。

结论

t-GRASP 是一种用于评估果蝇神经元之间突触连接的新型靶向 GRASP 方法。它与所有三种果蝇双转录系统的灵活使用显著扩展了 GRASP 在果蝇中的潜在用途。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e1a5/6689385/3098da873e22/nihms-1041056-f0006.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e1a5/6689385/b6e26809584b/nihms-1041056-f0001.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e1a5/6689385/f55ee60bf1b1/nihms-1041056-f0002.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e1a5/6689385/1289e0a38df0/nihms-1041056-f0003.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e1a5/6689385/c3a993b293df/nihms-1041056-f0004.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e1a5/6689385/48e2597d87b2/nihms-1041056-f0005.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e1a5/6689385/3098da873e22/nihms-1041056-f0006.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e1a5/6689385/b6e26809584b/nihms-1041056-f0001.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e1a5/6689385/f55ee60bf1b1/nihms-1041056-f0002.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e1a5/6689385/1289e0a38df0/nihms-1041056-f0003.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e1a5/6689385/c3a993b293df/nihms-1041056-f0004.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e1a5/6689385/48e2597d87b2/nihms-1041056-f0005.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e1a5/6689385/3098da873e22/nihms-1041056-f0006.jpg

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