General method for targeting genes to specific neuronal subtypes in mammals

描述（由申请人提供）：本项目的目标是开发一种广泛适用的方法，用于在大鼠、灵长类动物和其他哺乳动物脑中方便地进行转基因（如光遗传学试剂）的细胞类型特异性表达。大脑中的回路由不同的神经元亚型组成，这些亚型通常由特定基因的选择性表达来定义。例如，由D1R和D2R多巴胺受体的表达定义的神经元代表在强化学习和成瘾中发挥不同作用的不同神经元亚群。将越来越多的分子工具（包括荧光团和光遗传学试剂）的表达靶向于这种遗传学定义的神经元亚型的能力提供了用于解剖神经回路的有力方法。然而，目前用于实现细胞类型特异性转基因表达的方法通常依赖于转基因小鼠品系。细胞特异性转基因表达因此不能容易地实现在其他模式生物，如大鼠或猴子，这可能提供实验优势。此外，即使当小鼠是合适的模型系统时，新的转基因小鼠品系的产生和维持也是昂贵的、劳动密集型的和缓慢的。因此，实现转基因的特定细胞类型表达的一般方法将具有许多潜在的研究和治疗应用。我们建议开发一种新的策略，在特定的神经元亚型的报告基因的表达。我们的策略是基于Cre或Flp重组酶在特定神经元细胞类型中的特异性表达。然后，转基因的表达可以被限制在表达重组酶的神经元中。然而，与先前通过产生转基因敲入小鼠实现细胞类型特异性重组酶表达的策略不同，我们的方法可以通过重组病毒递送重组酶和转基因。因此，我们的方法可以应用于生物体中的遗传操作的潜力是有限的。我们的方法将有助于理解成瘾的大脑回路，并可能最终允许在治疗人类神经精神疾病的神经元子回路的目标。

DESCRIPTION (provided by applicant): The goal of this project is to develop a broadly applicable method for convenient cell-type specific expression of transgenes, such as optogenetic reagents, in rat, primate and other mammalian brains. Circuits in the brain consist of diverse neuronal subtypes which are often defined by the selective expression of specific genes. For example, neurons defined by the expression of the D1R and D2R dopamine receptors represent distinct neuronal subpopulations which play distinct roles in reinforcement learning and addiction. The ability to target the expression of a growing number of molecular tools, including flourophores and optogenetic reagents, to such genetically-defined neuronal subtypes provides a powerful method for dissecting neural circuits. However, current approaches for achieving cell-type specific transgene expression typically rely on transgenic mouse lines. Cell-specific transgene expression cannot therefore be easily achieved in other model organisms, such as rats or monkeys, which may offer experimental advantages. Moreover, even when the mouse is a suitable model system, the generation and maintenance of new transgenic mouse lines is expensive, labor-intensive and slow. A general method for achieving specific cell-type expression of transgenes would therefore have many potential applications for research and therapy. We propose to develop a novel strategy for the expression of reporter genes in specific neuronal subtypes. Our strategy is based on the specific expression of either Cre or Flp recombinase in specific neuronal cell-types. The expression of transgenes can then be restricted to neurons that express the recombinase. However, unlike previous strategies in which cell-type specific recombinase expression is achieved through the generation of transgenic knock-in mice, our method can deliver both the recombinase and the transgene via recombinant viruses. Thus our method can be applied in organisms in which the potential for genetic manipulation is limited. Our approach will contribute to the understanding of the brain circuitry underlying addiction, and may eventually allow targeting of neuronal sub circuits in the treatment of human neuropsychiatric disorders.