Viral transduction of the neonatal brain delivers controllable genetic mosaicism for visualising and manipulating neuronal circuits in vivo.
Viral transduction of the neonatal brain delivers controllable genetic mosaicism for visualising and manipulating neuronal circuits in vivo.
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DOI：
10.1111/ejn.12126
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发表时间：
2013-04
期刊：
影响因子：
--
通讯作者：
Jankowsky JL
中科院分区：
文献类型：
--
作者：
Kim JY;Ash RT;Ceballos-Diaz C;Levites Y;Golde TE;Smirnakis SM;Jankowsky JL
新生儿脑室内注射腺相关病毒已被证明能广泛地感染整个大脑的神经元，但其在实验神经科学中的全部潜力尚未得到充分的探索。我们报告了一个详细的分析方法的多功能性，重点是实验应用的遗传操作的工具，目前缺乏。将病毒注射到新生小鼠脑中是快速、容易的，并且进入用其他技术（例如电穿孔）难以靶向的脑区域，包括小脑和脑干。我们表明，病毒转导产生一个固有的马赛克表达模式，可以利用不同的滴度，使分离的神经元或密集包装的人口。我们证明，病毒编码的蛋白质的表达比以前认为的要快得多，允许在神经元可塑性的关键时期进行遗传扰动，但也是持久和稳定的，允许对衰老进行长期研究。我们利用这些特征来可视化和操纵后脑中的神经元，这些神经元已经被普遍应用于皮层的方法所拒绝。我们表明，病毒标记有助于分析小脑浦肯野神经元的出生后树突状成熟，使个别细胞容易区分，然后证明，相同的稀疏标记允许活在体内成像的成熟浦肯野神经元的分辨率足以完成分析重建。鉴于病毒构建、包装服务和转基因动物的可用性不断提高，这些技术应该有助于对大脑发育、功能和退化进行广泛的实验。
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 cerebellum and brain stem 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 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 visualize 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 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.
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影响因子：
5.4
作者：
Akache, Bassel;Grimm, Dirk;Kay, Mark A.
通讯作者：
Kay, Mark A.
影响因子：
14.8
作者：
Holtmaat, Anthony;Bonhoeffer, Tobias;Chow, David K.;Chuckowree, Jyoti;De Paola, Vincenzo;Hofer, Sonja B.;Huebener, Mark;Keck, Tara;Knott, Graham;Lee, Wei-Chung A.;Mostany, Ricardo;Mrsic-Flogel, Tom D.;Nedivi, Elly;Portera-Cailliau, Carlos;Svoboda, Karel;Trachtenberg, Joshua T.;Wilbrecht, Linda
通讯作者：
Wilbrecht, Linda
DOI：
10.1038/mt.2008.166
发表时间：
2008-10
期刊：
Molecular therapy : the journal of the American Society of Gene Therapy
影响因子：
--
作者：
通讯作者：
--
影响因子：
25
作者：
Dombeck, Daniel A.;Harvey, Christopher D.;Tian, Lin;Looger, Loren L.;Tank, David W.
通讯作者：
Tank, David W.
影响因子：
1.5
作者：
Guo, CY;Yang, WY;Lobe, CG
通讯作者：
Lobe, CG