Plasticity in deep cerebellar nuclei as a function of classical conditioning

描述(申请人提供)：为了帮助确定学习和记忆的生物学基础，我们的研究试图了解小脑的可塑性如何调节眨眼条件作用。为了追求这一目标，我们在体外记录了兔小脑皮质蒲肯野细胞在眼皮条件化后学习特异性突触和细胞膜的变化。这种方法有两个局限性。首先，许多不同的小脑小叶参与眼皮条件反射。其次，无法确认检测到变化的细胞是否参与了行为。现在，这些问题有了一个潜在的解决方案。首先，所有小脑小叶都投射到小脑深层核团，越来越多的证据表明，由于眨眼条件反射，间位前核内的神经活动、突触数量和基因表达发生了变化。其次，有逆行神经元追踪技术能够识别与眨眼有关的小脑深层神经元。然而，尽管可以看到这些细胞，但在成人小脑深核周围有一个致密的神经周网，阻止了从这些细胞进行记录。为了克服所有这些限制，我们正在开发一种大鼠小脑片制备方法，在这种方法中，我们可以在眼皮内注射荧光逆行示踪剂后，可视化并记录前间位中的眨眼相关神经元。有了这个准备，我们将达到两个特定的目标：(1)识别和表征与大鼠眨眼有关的前间置神经元；(2)确定由于眨眼条件作用而发生的已识别的前间置神经元突触和固有膜特性的变化。这项工作之所以重要，是因为它将对理解学习和记忆背后的突触和内在膜变化产生影响，这是一个众所周知的行为模型。这项拟议的工作具有创新性，因为它开发了一种切片制备方法，首次提供了涉及学习和记忆的已识别神经元角色的数据。它结合了一种新的方法-视觉可识别的反应相关神经元在前间置核与先进的体外电生理记录技术在一个众所周知的学习和记忆-大鼠眨眼条件作用的模型中。与公共卫生相关：有证据表明，动物和人类小脑深层核团的损伤对运动和非运动形式的学习和记忆都有重大影响。因此，检测大鼠小脑核细胞突触和固有膜特性的变化作为眨眼条件作用的功能，可能为学习和记忆的生物学机制提供深入的见解。通过识别这些变化，拟议的工作有可能帮助理解学习和记忆失败的发病机制，并开发针对这些失败的干预策略。

DESCRIPTION (provided by applicant): To help determine the biological basis of learning and memory, our research seeks to understand how plasticity in the cerebellum mediates eyeblink conditioning. To pursue this goal, we have recorded in vitro learning-specific synaptic and membrane changes in Purkinje cells of the rabbit cerebellar cortex following eyelid conditioning. There are two limitations to this approach. First, a number of different cerebellar lobules are involved in eyelid conditioning. Second, it is not possible to confirm whether cells in which changes are detected were involved in the behavior. There is now a potential solution to these problems. First, all cerebellar lobules project to deep cerebellar nuclei and there is growing evidence that changes in neural activity, synapse number and gene expression occur within the anterior interpositus nucleus as a result of eyeblink conditioning. Second, there are retrograde neuronal track tracing techniques able to identify deep cerebellar neurons involved in the eyeblink. However, although these cells can be visualized, there is a dense perineuronal net surrounding adult deep cerebellar nuclei that prevents recording from these cells. To overcome all of these limitations, we are developing a rat pup cerebellar slice preparation in which we can visualize and record from eyeblink-related neurons in the anterior interpositus following injection of a fluorescent retrograde tracer into the eyelid. With this preparation in hand, we will address two specific aims: (1) identify and characterize anterior interpositus neurons involved in the rat eyeblink; and (2) determine the changes in synaptic and intrinsic membrane properties of identified anterior interpositus neurons that occur as a result of eyeblink conditioning. This work is important because of the impact it will have on understanding the synaptic and intrinsic membrane changes underlying learning and memory in a well-understood behavioral model. The proposed work is innovative because it develops a slice preparation that for the first time provides data on the role of identified neurons involved in learning and memory. It combines a novel approach - visually identifiable response-related neurons in the anterior interpositus nucleus with advanced in vitro electrophysiological recording techniques in a well-understood model of learning and memory - rat eyeblink conditioning. PUBLIC HEALTH RELEVANCE: The evidence is clear that damage to the deep cerebellar nuclei in animals and humans has a major impact on both motor and non-motor forms of learning and memory. Consequently, examining changes in the synaptic and intrinsic membrane properties of cells in the cerebellar nuclei as a function of eyeblink conditioning in rats may provide insights into the biological mechanisms of learning and memory. By identifying these changes, the proposed work has the potential to help understand the pathogenesis of failures in learning and memory and to develop intervention strategies for those failures.