Anterograde monosynaptic transneuronal tracers derived from herpes simplex virus 1 strain H129.
Anterograde monosynaptic transneuronal tracers derived from herpes simplex virus 1 strain H129.
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源自单纯疱疹病毒 1 株 H129 的顺行单突触跨神经元示踪剂
DOI：
10.1186/s13024-017-0179-7
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发表时间：
2017-05-12
影响因子：
15.1
通讯作者：
Luo MH
中科院分区：
文献类型：
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作者：
Zeng WB;Jiang HF;Gang YD;Song YG;Shen ZZ;Yang H;Dong X;Tian YL;Ni RJ;Liu Y;Tang N;Li X;Jiang X;Gao D;Androulakis M;He XB;Xia HM;Ming YZ;Lu Y;Zhou JN;Zhang C;Xia XS;Shu Y;Zeng SQ;Xu F;Zhao F;Luo MH
单纯疱疹病毒1型129株(H129)代表了一种很有前途的顺行神经回路示踪工具，它是对现有逆行示踪剂的补充。然而，目前的H129衍生示踪剂是多突触的，既不足以标记神经元的细节，也不能确定直接投射靶标为单突触示踪剂。其中，H129-G4是通过在H129基因组中插入双链连接的绿色荧光蛋白盒而获得的；H129-ΔTK-TdT是通过删除胸苷激酶(TK)基因并在H129基因组中添加tdTomato编码基因而获得的。结果H129-G4能够通过多个突触传递，用绿色荧光蛋白标记神经元，并显示标记神经元的形态细节。H129-ΔTK-TdT既不在神经元中复制也不扩散，而是在辅助病毒互补表达的TK存在的情况下，向突触后神经元传递tdTomato并标记。H129-ΔTK-TdT还能够在Cre转基因小鼠的特定脑区定位特定神经元类型的直接投影组。结论在互补表达TK的辅助病毒的辅助下，H129-ΔTK-TdT在最初感染的神经元中复制，通过一个突触顺序性传递，并用td番茄标记突触后神经元。H129-ΔTK-TDT顺行单突触示踪系统为神经元回路的直接输出标测提供了一个有用的工具。H129-G4是一种顺行多突触示踪剂，具有足够强的标记信号，足以显示神经元形态的细节。
BackgroundHerpes simplex virus type 1 strain 129 (H129) has represented a promising anterograde neuronal circuit tracing tool, which complements the existing retrograde tracers. However, the current H129 derived tracers are multisynaptic, neither bright enough to label the details of neurons nor capable of determining direct projection targets as monosynaptic tracer.MethodsBased on the bacterial artificial chromosome of H129, we have generated a serial of recombinant viruses for neuronal circuit tracing. Among them, H129-G4 was obtained by inserting binary tandemly connected GFP cassettes into the H129 genome, and H129-ΔTK-tdT was obtained by deleting the thymidine kinase (TK) gene and adding tdTomato coding gene to the H129 genome. Then the obtained viral tracers were tested in vitro and in vivo for the tracing capacity.ResultsH129-G4 is capable of transmitting through multiple synapses, labeling the neurons by green florescent protein, and visualizing the morphological details of the labeled neurons. H129-ΔTK-tdT neither replicates nor spreads in neurons alone, but transmits to and labels the postsynaptic neurons with tdTomato in the presence of complementary expressed TK from a helper virus. H129-ΔTK-tdT is also capable to map the direct projectome of the specific neuron type in the given brain regions in Cre transgenic mice. In the tested brain regions where circuits are well known, the H129-ΔTK-tdT tracing patterns are consistent with the previous results.ConclusionsWith the assistance of the helper virus complimentarily expressing TK, H129-ΔTK-tdT replicates in the initially infected neuron, transmits anterogradely through one synapse, and labeled the postsynaptic neurons with tdTomato. The H129-ΔTK-tdT anterograde monosynaptic tracing system offers a useful tool for mapping the direct output in neuronal circuitry. H129-G4 is an anterograde multisynaptic tracer with a labeling signal strong enough to display the details of neuron morphology.