GENE THERAPY OF BASAL FOREBRAIN CHOLINERGIC LESIONS

描述：(摘自申请者的摘要)胆碱能损伤 使用免疫毒素的动物的基底前脑(CBF)系统产生健壮的 行为缺陷和模仿阿尔茨海默病(AD)的某些方面。这 模型对于测试乙酰胆碱在认知中的作用是有用的 过程和新的治疗策略，如基因疗法。有能力 控制基因表达是这项技术的重要要求 临床应用。本提案中正在探索的载体系统可能是 外来管制的。这项提案将调查恢复 CBF免疫毒素损伤动物的神经递质功能 通过补充胆碱促进移植后ACh的释放，2) 能抑制或诱导胆碱转基因的细胞移植 乙酰转移酶(ChAT)。可阻抑系统(四环素)，以及 诱导系统(蜕皮激素)，在一个快速的， 可逆和高度特定的时尚。 在第一系列实验中，调查人员将确定 不同剂量的外源性胆碱可以增加产量。 和从基因工程成纤维细胞中释放乙酰胆碱 脑血流免疫毒性损伤后大鼠的大脑皮层和海马区。 一旦确定了释放的增加，这一行为的影响 将确定扩大释放。 对于第二和第三系列实验，他们将使用可调节的 给药后诱导或抑制ChAT表达的细胞系 分别为多西环素或木犀草酮A。在它们的免疫毒素损伤中 将证明从这些细胞中释放的乙酰胆碱 新大脑皮层和海马体是行为活动的必要条件和充分条件 恢复。这些可调节基因的效果将在空间上进行测试。 而非空间任务在确定外源性刺激的剂量和持续时间后 诱导或抑制基因表达所需的化合物。如果成功，则 实验将证明ACh的释放可以通过外源控制。 并能显著改善动物的行为。 在嫁接或定向基因之前，必须控制递质的释放 插入实验可以在人体上进行，以便能够 安全地终止基因产品的交付。这些方法不仅将 适用于AD，但其他神经退行性疾病和分娩 其他感兴趣的转基因。

DESCRIPTION: (adapted from applicant's abstract) Lesions of the cholinergic basal forebrain (CBF) system in animals using an immunotoxin produce robust behavioral deficits and mimic some aspects of Alzheimer's disease (AD). This model is useful for testing both the role of acetylcholine in cognitive processes and new therapeutic strategies such as gene therapy. The ability to control gene expression is an important requirement of this technology for clinical application. The vector systems being explored in this proposal may be exogenously regulated. This proposal will investigate mechanisms of restoring neurotransmitter function in animals with CBF immunotoxin lesions by 1) enhancing ACh release postgrafting via choline supplementation, and 2) transplanting cells capable of repressing or inducing the transgene for choline acetyltransferase (ChAT). The repressible system (tetracycline), and the inducible system (eccdysteroid), modulates gene expression of ChAT in a rapid, reversible and highly specific fashion. In the first series of experiments, the investigators will determine whether different doses of exogenously administered choline can augment the production and release of acetylcholine from genetically engineered fibroblasts grafted to the cortex and hippocampus of rats following immunotoxic lesions of the CBF. Once an increase in release has been determined, behavioral effects of this augmented release will be determined. For the second and third series of experiments, they will use the regulatable cell lines that either induce or repress ChAT expression after administration of doxycycline or Muristerone A, respectively. In their immunotoxin lesion they will demonstrate that acetylcholine released from these cells after grafting to the neocortex and hippocampus is both necessary and sufficient for behavioral recovery. The effects of these regulatable genes will be tested in a spatial and non-spatial task after determining the dose and duration of the exogenous compound needed to induce or repress gene expression. If successful, the experiments will demonstrate that release of ACh can be exogenously controlled in animals and results in significant behavioral improvement. The control of transmitter release is necessary before grafting or direct gene insertion experiments can be entertained in humans in order to be able to safely terminate delivery of the gene product. These approaches will not only be applicable for AD, but other neurodegenerative disorders and for delivering other transgenes of interest.