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首页> 外文期刊>The European Journal of Neuroscience >Viral transduction of the neonatal brain delivers controllable genetic mosaicism for visualising and manipulating neuronal circuits in vivo
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Viral transduction of the neonatal brain delivers controllable genetic mosaicism for visualising and manipulating neuronal circuits in vivo

机译：新生儿大脑的病毒转导提供可控制的遗传镶嵌，以可视化和操纵体内神经元回路

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摘要

The neonatal intraventricular injection of adeno-associated virus has been shown to transduce neurons widely throughout the brain, but its full potential for experimental neuroscience has not been adequately explored. We report a detailed analysis of the method's versatility with an emphasis on experimental applications where tools for genetic manipulation are currently lacking. Viral injection into the neonatal mouse brain is fast, easy, and accesses regions of the brain including the cerebellum and brainstem that have been difficult to target with other techniques such as electroporation. We show that viral transduction produces an inherently mosaic expression pattern that can be exploited by varying the titer to transduce isolated neurons or densely-packed populations. We demonstrate that the expression of virally-encoded proteins is active much sooner than previously believed, allowing genetic perturbation during critical periods of neuronal plasticity, but is also long-lasting and stable, allowing chronic studies of aging. We harness these features to visualise and manipulate neurons in the hindbrain that have been recalcitrant to approaches commonly applied in the cortex. We show that viral labeling aids the analysis of postnatal dendritic maturation in cerebellar Purkinje neurons by allowing individual cells to be readily distinguished, and then demonstrate that the same sparse labeling allows live in vivo imaging of mature Purkinje neurons at a resolution sufficient for complete analytical reconstruction. Given the rising availability of viral constructs, packaging services, and genetically modified animals, these techniques should facilitate a wide range of experiments into brain development, function, and degeneration.
机译：新生儿脑室内注射腺伴随病毒已被证明可在整个大脑中广泛地传导神经元，但尚未充分探索其在实验神经科学中的全部潜力。我们报告了该方法的多功能性的详细分析，重点是目前缺乏基因操作工具的实验应用。将病毒注射到新生鼠脑中是快速，容易的，并且可以进入脑部区域，包括小脑和脑干，这些区域很难用其他技术（例如电穿孔）来靶向。我们显示病毒转导产生固有的镶嵌表达模式，可以通过改变滴度来转导分离的神经元或密集的种群来利用。我们证明，病毒编码蛋白的表达比以前认为的要活跃得多，可以在神经元可塑性的关键时期发生基因扰动，而且可以持久稳定，可以进行衰老的长期研究。我们利用这些功能来可视化和操纵后脑中的神经元，这些神经元对通常在皮质中使用的方法具有顽固性。我们显示病毒标记通过允许容易区分单个细胞，有助于小脑浦肯野神经元的产后树突状成熟的分析，然后证明相同的稀疏标记允许成熟浦肯野神经元的活体内成像以足以进行完整分析重建的分辨率。鉴于病毒构建物，包装服务和转基因动物的可用性不断提高，这些技术应有助于大脑发育，功能和变性的广泛实验。

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