Unique Luminal Localization of VGAT-C Terminus Allows for Selective Labeling of Active Cortical GABAergic Synapses
Unique Luminal Localization of VGAT-C Terminus Allows for Selective Labeling of Active Cortical GABAergic Synapses
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DOI：
10.1523/jneurosci.3887-08.2008
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发表时间：
2008-12-03
影响因子：
5.3
通讯作者：
Haertig, Wolfgang
中科院分区：
文献类型：
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作者：
Martens, Henrik;Weston, Matthew C.;Haertig, Wolfgang
神经递质进入突触小泡是由囊泡神经递质转运体介导的。虽然这些转运蛋白属于不同的家族，但它们都被认为与偶数个跨膜结构域共享一个共同的整体拓扑结构。利用抗原表位特异性抗体和质谱仪，我们发现囊泡型GABA转运体(VGAT)具有数量不等的跨膜区，N端面向细胞质，C端位于突触囊泡腔内。免疫细胞化学和膜片钳电生理证实，识别VGAT C末端的抗体(anti-VGAT-C)选择性地标记活培养的海马神经元和纹状体神经元的GABA能神经末梢。将荧光显色的抗VGAT-C注射到小鼠的海马区，可在体内对GABA能突触进行特异性标记。总体而言，我们的数据为研究新的GABA释放位点，表征抑制性囊泡运输，并确定它们在已识别的GABA能突触中对抑制性神经传递的贡献提供了可能性。
Neurotransmitter uptake into synaptic vesicles is mediated by vesicular neurotransmitter transporters. Although these transporters belong to different families, they all are thought to share a common overall topology with an even number of transmembrane domains. Using epitope-specific antibodies and mass spectrometry we show that the vesicular GABA transporter ( VGAT) possesses an uneven number of transmembrane domains, with the N terminus facing the cytoplasm and the C terminus residing in the synaptic vesicle lumen. Antibodies recognizing the C terminus of VGAT(anti-VGAT-C) selectively label GABAergic nerve terminals of live cultured hippocampal and striatal neurons as confirmed by immunocytochemistry and patch-clamp electrophysiology. Injection of fluorochromated anti-VGAT-C into the hippocampus of mice results in specific labeling of GABAergic synapses in vivo. Overall, our data open the possibility of studying novel GABA release sites, characterizing inhibitory vesicle trafficking, and establishing their contribution to inhibitory neurotransmission at identified GABAergic synapses.