An optogenetic approach to exploring climbing fiber connections in the cerebellum

探索小脑攀爬纤维连接的光遗传学方法

• 批准号: 8125240
• 负责人: Paul James Mathews
• 金额: $ 5.13万
• 依托单位: UNIVERSITY OF CALIFORNIA LOS ANGELES
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-08-28 至 2014-08-27
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8125240
• 关键词: Acute; Automobile Driving; Axon; Behavior; Behavioral; Brain; Brain region; Cell Nucleus; Cells; Cerebellar Diseases; Cerebellar cortex structure; Cerebellum; Complement; Complex; Detection; Disease; Educational process of instructing; Effectiveness; Fiber; Glutamates; Goals; In Vitro; Individual; Inferior; Injection of therapeutic agent; Intention; Interneurons; Ion Channel; Knowledge; Lead; Learning; Lesion; Light; Location; Long-Term Depression; Memory; Mental Depression; Methods; Motor; Motor output; Movement; Myoepithelial cell; Nature; Nervous system structure; Neurons; Olives - dietary; Optics; Output; Pathway interactions; Play; Property; Proteins; Purkinje Cells; Research Personnel; Role; Sensory; Shapes; Signal Pathway; Signal Transduction; Slice; Specificity; Spottings; Structure of molecular layer of cerebellar cortex; Symptoms; Synapses; Synaptic plasticity; Techniques; Testing; Trauma; Viral; Work; age effect; cell type; improved; in vivo; knowledge base; mossy fiber; motor control; motor learning; novel; patch clamp; receptive field; research study; response; somatosensory; stellate cell; therapy design; transmission process

DESCRIPTION (provided by applicant): Experimental lesions and blunt force traumas to the cerebellum result in behavioral abnormalities that indicate this brain region plays an important role in controlling smooth coordinated movement and motor memory (Fine, Ionita, & Lohr, 2002). Specifically, researchers believe the cerebellum evaluates the disparities between intention and action, and then adjusts the motor output to correct for these disparities in order to generate a desired, smooth-motor behavior. Experiments suggest these corrections arise from dynamic changes in the strength of synaptic connections in both the cerebellar cortex and deep nuclei. In addition, these changes are likely driven by the association or coincident detection of signals from two specific pathways, one by way of the mossy fibers (MF, carrying sensory information) and the other by way of the climbing fibers (CF, indicating a disparity or error in the motor command). Originating in the inferior olive the climbing fiber delivers a unique and powerful input that generates a "complex spike" in the sole output of the cerebellar cortex, the Purkinje cells (PCs). This input, when paired with parallel fiber (PF; mossy fiber relay) activation decreases the somatosensory receptive fields of PCs (Jvrntell & Ekerot, 2002). This change in receptive field is due to the depression of a subset of PF-PC synapses, a mechanism believed to remove sensory signals producing undesired motor behaviors. Similar experiments also demonstrate CFs drive associative changes in the receptive fields of molecular layer inhibitory interneurons (MLI) that synapse onto PCs. However, the nature of the CF-MLI connection remains unclear, nor are the mechanisms driving the associative plasticity that result in receptive field changes known. This deficiency in the current state of cerebellar knowledge is the result of an inability to reliably stimulate CFs without activating neighboring axons from other neuron types. To overcome this technical challenge, a novel optogenetic approach has been developed to allow robust stimulation of isolated CFs. The first aim of this proposal will further confirm preliminary results demonstrating the reliability and specificity of photostimulating CFs expressing Channelrhodopsin 2 by systematically exploring the optical stimulation and viral injection parameters necessary for robust CF stimulation. Using this technique, I propose to describe both the nature of CF-MLI transmission as well as the mechanisms and rules governing the CF- driven associative plasticity between parallel fibers and MLIs. This will be accomplished through whole-cell patch clamp recordings from MLIs in acute slices during selective CF photostimulation. These experiments will be the first of their kind to illustrate the effectiveness of optogenetic techniques in exploring the cerebellar cortex. In the end results from these experiments will allow for better predictions of how the cerebellar cortex evaluates and corrects for disparities between intention and action. PUBLIC HEALTH RELEVANCE: Cerebellar dysfunction, which usually results from disease, blunt force trauma or the effects of aging, often presents itself as deficits in motor control and/or motor memory. The goal of this proposal is to examine the basic cellular properties and mechanisms governing motor control and memory in the cerebellar cortex using a novel technique to stimulate neurons in the brain. With a more informed knowledge base of cerebellar function at the cellular level researchers will be better prepared to design therapies to cure or alleviate the symptoms of cerebellar dysfunctions.

描述（由申请人提供）：小脑的实验性损伤和钝力创伤导致行为异常，表明该脑区在控制平稳协调运动和运动记忆中起着重要作用（Fine，Ionita和Lohr，2002）。具体来说，研究人员认为小脑评估意图和行动之间的差异，然后调整运动输出以纠正这些差异，从而产生所需的平滑运动行为。实验表明，这些校正来自小脑皮层和深层核中突触连接强度的动态变化。此外，这些变化可能是由来自两个特定通路的信号的关联或同时检测驱动的，一个通过苔藓纤维（MF，携带感觉信息），另一个通过攀爬纤维（CF，指示运动命令中的差异或错误）。起源于下橄榄的攀爬纤维提供了一个独特而强大的输入，在小脑皮层的唯一输出中产生一个“复杂的尖峰”，浦肯野细胞（PC）。这种输入，当与平行纤维（PF;苔藓纤维中继）激活配对时，会降低PC的体感感受野（Jvrntell & Ekerot，2002）。感受野的这种变化是由于PF-PC突触子集的抑制，这是一种被认为可以消除产生不期望的运动行为的感觉信号的机制。类似的实验还表明，CF驱动突触到PC上的分子层抑制性中间神经元（MLI）的感受野的关联变化。然而，CF-MLI连接的性质仍然不清楚，也不是已知的机制驱动的关联可塑性，导致感受野的变化。目前小脑知识的这种缺陷是由于无法在不激活其他神经元类型的邻近轴突的情况下可靠地刺激CF。为了克服这一技术挑战，已经开发了一种新的光遗传学方法，以允许对分离的CF进行稳健刺激。该提案的第一个目的将通过系统地探索稳健CF刺激所需的光学刺激和病毒注射参数来进一步证实初步结果，所述初步结果证明表达视紫红质2的光刺激CF的可靠性和特异性。使用这种技术，我建议描述的性质CF-MLI传输以及机制和规则的CF驱动的关联可塑性之间的平行纤维和MLI。这将通过选择性CF光刺激期间急性切片中MLI的全细胞膜片钳记录来实现。这些实验将是第一个说明光遗传学技术在探索小脑皮层方面的有效性的实验。最终，这些实验的结果将有助于更好地预测小脑皮层如何评估和纠正意图与行动之间的差异。 公共卫生关系：小脑功能障碍通常由疾病、钝力创伤或衰老的影响引起，通常表现为运动控制和/或运动记忆的缺陷。这项计划的目的是研究小脑皮质中控制运动控制和记忆的基本细胞特性和机制，使用一种新的技术来刺激大脑中的神经元。随着小脑功能在细胞水平上的知识基础更加丰富，研究人员将更好地准备设计治疗或缓解小脑功能障碍症状的疗法。