Aktivitäts-abhängige Regulation von neuralen Zelloberflächenproteinen und ihre Funktion bei der Synaptogenese: Funktionelle Charakterisierung von CALEB

神经元活动对于大脑的复杂连接是必不可少的。电活动和有助于神经元电路结构重塑的分子之间的联系还没有被很好地理解。因此，在过去的几年里，我们一直在筛选细胞表面蛋白，这些蛋白建立了电活动之间的联系，并可能在电路形成中发挥作用。我们鉴定了两种细胞表面蛋白：CAR(柯萨奇病毒腺病毒受体)和CALEB。CAR是免疫球蛋白超家族的一种细胞黏附蛋白，它调节动作电位的频率，调节细胞内游离钙的水平。它主要在发育中的神经系统中表达，在成熟的神经系统中几乎检测不到。Caleb是分化因子EGF家族中含有胰岛素软骨酸盐的成员，可在质膜上进行蛋白水解性转换。这种转化是由电活动促成的。它的缺失导致早期发育阶段的几个大脑结构的突触前分化障碍，以及浦肯野细胞上攀升纤维突触的消除过程中的损害。Caleb仅限于中枢神经系统，主要表达于躯体-树突室。这一后续应用的目的是研究这两种蛋白和另外两种结构上与CAR相关的Ig超家族蛋白(称为BT-IgSF和CLMP)。根据它们在结构上的相似性，我们假设CLMP和BT-IgSF具有与CAR相似的功能。对于CAR和BT-IgSF，我们已经建立了转基因小鼠模型，对于CLMP，我们正在建立基因敲除的过程中。我们打算研究这些突变小鼠的神经元回路的发育。通过电生理学和组织学示踪技术，我们询问这些突变小鼠的神经元回路的形成是否受到损害。此外，我们打算表征与CAR介导的细胞内钙释放有关的信号级联。同样的做法也将适用于BT-IgSF和CLMP。我们推测CAR、BT-IgSF和CLMP调节邻近神经元细胞内游离钙的水平，从而影响神经组织中的钙波。为了进一步研究Caleb在突触前分化过程中的作用，我们将筛选Caleb的突触前受体蛋白。此外，还计划在Caleb突变小鼠的不同脑结构中进行几种电生理特征以及Caleb信号功能的研究。

Neuronal activity is essential for the complex wiring of the brain. The links between electric activity and molecules that contribute to structural remodeling of neuronal circuits are not well understood. In past years we have therefore screened for cell surface proteins which establish a link between electric activity and which might play a role in circuit formation. We characterized two cell surface proteins: CAR (coxsackievirus adenovirus receptor) and CALEB. CAR is a cell adhesion protein of the Ig superfamily which modulates the frequency of action potentials and regulates the level of intracellular free calcium. It is primarily expressed in the developing nervous system and is almost undetectable in the mature nervous system. CALEB is a chondroitinsulfate-containing member of the EGF-family of differentiation factors that becomes proteolytically converted at the plasma membrane. This conversion is facilitated by electric activity. Its absence results in deficits of presynaptic differentiation in several brain structures at early developmental stages and in impairments in the elimination process of the climbing fiber synapse on Purkinje cells. CALEB is restricted to the central nervous system and is predominantly expressed in the somato-dendritic compartment. Aims of this follow-up application are investigations on these two proteins and two additional proteins of the Ig superfamily structurally related to CAR (termed BT-IgSF and clmp). On the basis of their structural similarity we assume that clmp and BT-IgSF subserve a similar function as CAR. For CAR and BT-IgSF we have generated transgenic mouse models and for clmp we are in the process in establishing a knockout. We intend to study the development of neuronal circuits in these mutant mice. By electrophysiological and histological tracing techniques we ask whether the formation of neuronal circuits is impaired in these mutant mice. Furthermore, we intend to characterize a signaling cascade implicated in the CAR-mediated release of calcium from intracellular stores. The same will be done for BT-IgSF and clmp. We speculate that CAR, BT-IgSF and clmp modulate the level of intracellular free calcium in neighboring neurons and thereby influence calcium waves in neural tissues. To characterize the role of CALEB in the process of presynaptic differentiation further we will screen for a pre-synaptic receptor protein of CALEB. In addition several electrophysiological characterizations in different brain structures in CALEB mutant mice as wells as investigations on the signaling function of CALEB are planned.