Optogenetic Interrogation of Functional Synapse Formation by Corticospinal Tract Axons in the Injured Spinal Cord
Optogenetic Interrogation of Functional Synapse Formation by Corticospinal Tract Axons in the Injured Spinal Cord
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DOI：
10.1523/jneurosci.4203-15.2016
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发表时间：
2016-05-25
影响因子：
5.3
通讯作者：
Blackmore, Murray G.
中科院分区：
文献类型：
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作者：
Jayaprakash, Naveen;Wang, Zimei;Blackmore, Murray G.
为了恢复中枢神经系统损伤后的功能，必须刺激轴突延伸到失神经区域，关键的是，必须形成具有适当目标的功能性突触。我们之前的研究表明，脊髓损伤后，转录因子Sox11的强迫过表达会增加皮质脊髓束（CST）神经元的轴突生长。然而，行为结果并没有得到改善，这就提出了一个问题，即新发芽的轴突是否能够形成功能性突触。在这里，我们开发了一种光遗传学策略，结合单单位细胞外记录，来评估sox11刺激的CST轴突在颈脊髓回路中的功能整合能力。初始时间过程实验确定了病毒表达的通道视紫红质（ChR2）在颈脊髓CST细胞体和轴突终末的表达和功能。在成年小鼠中进行锥体切除术以剥夺左侧脊髓的CST输入，并使用腺相关病毒(AAV)-Sox11或AAV- ebfp对照以及AAV- chr2治疗右侧CST。正如预期的那样，Sox11治疗导致CST轴突强劲的中线交叉进入先前失神经的左脊髓。CST末端的光遗传激活导致了清晰的突触后反应，表明sox11刺激的轴突能够形成功能性突触。cst诱发的脊髓活动的分布图揭示了完整和新神经支配的脊髓组织之间的总体相似性。这些数据表明，sox11刺激的CST轴突形成的功能性突触没有显著的行为益处，这表明新突触可能是错误定位的，或者在协调功能输出的能力方面受到损害。
To restore function after injury to the CNS, axons must be stimulated to extend into denervated territory and, critically, must form functional synapses with appropriate targets. We showed previously that forced overexpression of the transcription factor Sox11 increases axon growth by corticospinal tract (CST) neurons after spinal injury. However, behavioral outcomes were not improved, raising the question of whether the newly sprouted axons are able to form functional synapses. Here we developed an optogenetic strategy, paired with single-unit extracellular recordings, to assess the ability of Sox11-stimulated CST axons to functionally integrate in the circuitry of the cervical spinal cord. Initial time course experiments established the expression and function of virally expressed Channelrhodopsin (ChR2) in CST cell bodies and in axon terminals in cervical spinal cord. Pyramidotomies were performed in adult mice to deprive the left side of the spinal cord of CST input, and the right CST was treated with adeno-associated virus (AAV)-Sox11 orAAV-EBFPcontrol, along with AAV-ChR2. As expected, Sox11 treatment caused robust midline crossing of CST axons into previously denervated left spinal cord. Clear postsynaptic responses resulted from optogenetic activation of CST terminals, demonstrating the ability of Sox11-stimulated axons to form functional synapses. Mapping of the distribution of CST-evoked spinal activity revealed overall similarity between intact and newly innervated spinal tissue. These data demonstrate the formation of functional synapses by Sox11-stimulated CST axons without significant behavioral benefit, suggesting that new synapses may be mistargeted or otherwise impaired in the ability to coordinate functional output.