Genetic and phenotypic analysis of tippy mutant mice

描述（由申请人提供）：在人类中，先天性小脑共济失调是相当常见的大脑出生缺陷，在儿科人群中引起显著的发病率和死亡率。尽管它们的临床患病率，这些疾病的遗传和发育基础了解甚少。在模式生物中进行的大量研究表明，浦肯野细胞具有精细的、广泛分支的、典型的树突状乔木，对正常的小脑功能至关重要。此外，浦肯野细胞的功能与其细胞形态直接相关。然而，对浦肯野细胞形态发育的内在因素和机制知之甚少。自发的神经系统小鼠突变体tippy具有巨大的潜力，可以为深入了解浦肯野细胞树突状发育程序提供见解。初步的表型分析表明，tipppy突变小鼠的浦肯野细胞在树突发生和脊柱发生方面存在缺陷，这很可能不是外部因素的结果。在这个提议中，一个定位克隆策略将被用于鉴定tipppy突变小鼠的分子损伤和鉴定致病tippy基因（Aim I）。为了获得对小脑表型的详细了解，将进行大量定量和定性免疫组织化学分析（目标II和III）。具体来说，在Aim II中，将使用高尔基染色的浦肯野细胞的透明成像图来量化突变体浦肯野细胞分支和脊柱缺陷。Aim III将进行各种免疫组织化学和顺行标记实验，以更充分地表征浦肯野细胞突触及其传入神经的分布和组成。一旦tippy基因被确定，其导致浦肯野细胞表型的机制将被研究，更广泛的目标是使用tippy小鼠作为模型来解剖调节浦肯野细胞树突发育的内在发育信号。迫切需要的浦肯野细胞树突状发育程序的内在阐明将可能提供对许多先天性共济失调的分子和生化基础的更全面的理解。

DESCRIPTION (provided by applicant): In humans, congenital cerebellar ataxias are fairly common birth defects of the brain that cause significant morbidity and mortality in pediatric populations. Despite their clinical prevalance, the genetic and developmental basis of these diseases are poorly understood. Extensive work in model organisms has demonstrated that Purkinje cells, with their elaborate, extensively branched, stereotypical dendritic arbors are essential to normal cerebellar function. Furthermore, Purkinje cell function is directly related to its cell morphology. However, very little is known about the intrinsic factors and mechanisms driving Purkinje cell morphological development. The spontaneous neurological mouse mutant, tippy, holds great potential to provide insight into the intrinsic Purkinje cell dendritic developmental program. Preliminary phenotypic analysis demonstrates that Purkinje cells of tippy mutant mice have defects in dendritogenesis and spinogenesis that are most likely not a consequence of extrinsic factors. In this proposal, a positional cloning strategy will be used to identify the molecular lesion in tippy mutant mice and to identify the causative tippy gene (Aim I). To obtain a detailed understanding of the cerebellar phenotype, a number of quantitative and qualitative immunohistochemical analyses will be performed (Aims II and III). Specifically, the tippy mutant Purkinje cell branching and spine defect will be quantified using camera lucida drawings of Golgi-stained Purkinje cells in Aim II. Various immunohistochemical and anterograde labeling experiments will be performed in Aim III to more fully characterize the distribution and composition of tippy Purkinje cell synapses with their afferents. Once the tippy gene is identified, the mechanism by which it causes the Purkinje cell phenotype will be investigated with the broader aim of using the tippy mouse as a model to dissect out the intrinsic developmental signals that regulate Purkinje cell dendritic development. The much needed illumination of the intrinsic Purkinje cell dendritic developmental program will likely provide a more comprehensive understanding of the molecular and biochemical basis of numerous congenital ataxias.