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: 8520408
• 负责人: Paul James Mathews
• 金额: $ 4.63万
• 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 2014-06-27
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8520408
• 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; optogenetics; 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.

描述（由申请人提供）：针对小脑的实验病变和钝性创伤导致行为异常，表明该大脑区域在控制平滑的配位运动和运动记忆中起着重要作用（Fine，Ionita，＆Lohr，＆Lohr，2002）。具体而言，研究人员认为，小脑会评估意图和动作之间的差异，然后调整电动机输出以纠正这些差异，以产生所需的平滑运动行为。实验表明，这些校正是由小脑皮层和深核中突触连接强度的动态变化产生的。此外，这些变化很可能是由从两种特定途径的信号的关联或重合检测驱动的，一种是通过苔藓纤维（MF，携带感官信息），另一个是通过攀爬纤维（CF，表明电动机命令中的差异或错误）来驱动的。起源于下橄榄，攀岩纤维提供了独特而强大的输入，该输入在小脑皮层的唯一输出中产生了“复杂的尖峰”，Purkinje细胞（PC）。当与平行纤维（PF；苔藓纤维继电器）配对时，该输入会降低PC的体感感应场（Jvrntell＆Ekerot，2002）。接受场的这种变化是由于PF-PC突触的一部分抑制，这种机制被认为消除了产生不希望的运动行为的感觉信号。类似的实验还表明，CFS驱动了突触到PC的分子层抑制性中间神经元（MLI）的接收场的关联变化。然而，CF-MLI连接的性质尚不清楚，驱动促进联想可塑性的机制也不清楚，从而导致人们已知的接收场变化。小脑知识当前状态的这种缺乏是无法可靠刺激CF的结果，而无需激活其他神经元类型的相邻轴突。为了克服这一技术挑战，已经开发了一种新型的光遗传学方法，以允许对孤立的CFS进行强有力的刺激。该提案的第一个目的将进一步确认初步结果，证明了通过系统地探索鲁棒CF刺激所需的光学刺激和病毒注射参数来表达ChannelRhodopsin 2的光刺激CF的可靠性和特异性。我建议使用这种技术来描述CF-MLI传输的性质以及管理平行纤维和MLI之间CF驱动的关联可塑性的机制和规则。这将通过在选择性CF光刺激期间从MLIS中的全细胞贴片夹记录来完成。这些实验将是其中第一个说明光遗传技术在探索小脑皮层中的有效性。最后，这些实验的结果将可以更好地预测小脑皮层如何评估和纠正意图和动作之间的差异。