BIOCHEMICAL DIFFERENTIATION OF SOMATIC MOTOR NEURONS

该项目的长期目标是了解 运动神经元表型的发育和成熟，以及如何 外部因素影响着这一过程。为此，我们目前 人们一直致力于制造一组单抗。 用它来识别啮齿动物发育过程中的躯体运动神经元。 一旦在发育过程中这些标记的顺序出现， 确定，运动神经元的抗原谱将作为 准确判断其分化状态的手段。 我们现在已经产生了几种抗体，可以选择性地识别体细胞 运动神经元。其中之一显示了这个类非常严格的专用性 神经元，免疫反应性出现在发育后期。相比之下， 另外两种抗体对相对不成熟的运动神经元进行染色。这些 结果表明运动神经元的分子标志物 分化是存在的，而且单抗技术可以 成功地用来制造针对这种抗原的探针。我们将继续 扩展我们针对这类神经元的标记面板，使用新颖的 免疫方案，以获得完整的抗原谱 运动神经元在整个发育过程中。 利用我们已经产生的抗体，我们的首要目标是 仔细观察免疫活性物质的时空分布 在不同群体的躯体运动神经元中的表位 发育中的脑干和脊髓。然后我们将扰乱传入输入 或目标接触者询问是否获得或维持免疫反应性 取决于这些参数。我们还将在生物化学特征上 并确定运动神经元中存在的免疫反应分子。 最后，我们建议研究运动神经元的分化。 使用脊柱切片制剂进行体外培养。我们会问这些抗原是否 运动神经元的原位表达可以在培养中获得，并且随着 相同的特异度和时间进程。这些实验，结合 原位研究，将提供分子分化的信息 这类重要的胆碱能神经元。

The long term goal of this project is to understand the steps in the development and maturation of the motor neuronal phenotype, and how extrinsic factors influence this process. Toward this end, our current efforts have been focused on generating a panel of monoclonal antibodies with which to identify somatic motor neurons during rodent development. Once the sequential appearance of such markers, during development, is determined, the antigenic profile of a motor neuron will serve as an accurate means of staging its state of differentiation. We have now generated several antibodies that selectively recognize somatic motor neurons. One of these shows a very strict specificity for this class of neurons, and immunoreactivity appears late in development. In contrast, two other antibodies stain relatively immature motor neurons. These results demonstrate that molecular markers of motor neuronal differentiation exists, and that monoclonal antibody techniques can be successfully used to produce probes to such antigens. We will continue to expand our panel of markers for this class of neurons, using novel immunization protocols, in order to obtain a complete antigenic profile of motor neurons throughout development. With the antibodies that we have already generated, our first aim is to examine carefully the spatial and temporal distribution of immunoreactive epitopes, in different populations of somatic motor neurons in the developing brainstem and spinal cord. We will then perturb afferent inputs or target contact to ask if acquisition or maintenance of immunoreactivity is dependent on these parameters. We will also biochemically characterize and identify the immunoreactive molecules present in motor neurons. Finally, we propose to examine the differentiation of motor neurons in vitro using a spinal slice preparation. We will ask whether the antigens expressed by motor neurons in situ can be acquired in culture, and with the same specificity and time course. These experiments, in conjunction with studies in situ, will provide information on the molecular differentiation of this important class of cholinergic neurons.