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EXPERIMENTAL STUDIES IN SENSORY PATHWAYS

感觉通路的实验研究

基本信息

批准号：
6702321
负责人：
Mark D Bevan
金额：
$ 36.93万
依托单位：
NORTHWESTERN UNIVERSITY AT CHICAGO
依托单位国家：
美国
项目类别：
财政年份：
1983
资助国家：
美国
起止时间：
1983-08-01 至 2007-01-31
项目状态：
已结题

项目摘要

Normal basal ganglia function is highly dependent on the activities of dopaminergic (DA) neurons in the SNc and GABAergic neurons in SNr/EP. However, many aspects of the mechanisms controlling SNc and SNr neural activity which influence dopamine and GABA release at their projection targets are still not well understood. Recently there has been a growing interest in converging inputs from the tegmental pedunculopontine (PPN) and subthalamic nucleus (STN) to the SNc DA and SNr GABA neurons. Although the functional role of these inputs is poorly understood, our previous studies suggested that acetylcholine (ACh) inputs from PPN depolarize the resting membrane potential of DA and GABA neurons by a combination of increased cationic influx and decreased K+ efflux, and reduces Ca2+ entry during the pacemaker-like slow depolarization (PLSD), which decreases the amplitude and duration of the subsequent spike after hyperpolarization. We, therefore, hypothesize that (1) the action of ACh increases the tendency for SNc DA and SNr GABAergic neurons to exhibit burst firing in response to glutamatergic (glu) input from the STN and PPN and (2) in addition to glu and ACh excitatory projections from the PPN and STN, but also GABAergic inhibitory projections to SN (causing disinhibition of GABA system by phasic inhibition of the SNr neurons) are modulating SNc DA and SNr GABAergic neuronal firing behavior. To test these hypothesis, we will utilize the newly developed in vitro organotypic culture preparation consisting of PPN, SN, STH and STR to conduct morphological and electrophysiological studies. Morphological studies will involve light and electron microscopy to (1) identify ACh, glu and GABAergic projection patterns and their terminal sites on the SNc DA and SNr GABA neurons, (2a) identify the nuclear origin of ACh and glu projections to SNc DA and SNr GABAergic neurons with a combined biocytin intracellular labeling, and (2b) identify the locations of the intracellularly labeled biocytin terminals making synapses with SNc DA and SNr GABA neurons and (3) identify the location of ACh, glu and GABA receptors on the SNc DA and SNr GABA neurons. Electrophysiological studies will involve use of whole-cell (current) clamp or intracellular sharp electrode recording to delineate the effect of (1) cholinergic receptor activation on Ca2+ entry during the PLSD and AHP,(2) the effect of excitatory synaptic inputs from PPN and STN and GABAergic inhibitory inputs from the STR on firing properties and bursting of SNc DA and SNr GABAergic neurons. Understanding the role of multiple inputs in the control and/or modulation of SNc and SNr cellular activity which affect their target structures (i.e. striatum and thalamus) is of fundamental importance relative to motor and behavioral function as well as clinical entities such as Parkinson's disease.

正常的基底神经节功能高度依赖于SNc中的多巴胺（DA）能神经元和SNr/EP中的GABA能神经元的活动。然而，许多方面的机制控制SNc和SNr神经活动的影响多巴胺和GABA的释放在其投射目标仍然没有得到很好的理解。近年来，研究者们越来越关注从被盖脚桥核（PPN）和丘脑底核（BNZ）到SNc DA和SNr GABA神经元的会聚输入。虽然这些输入的功能作用知之甚少，我们以前的研究表明，乙酰胆碱（ACh）的输入PPN通过增加阳离子内流和减少K+流出的组合降低DA和GABA神经元的静息膜电位，并减少在起搏器样缓慢去极化（PLSD），这降低了随后的超极化后尖峰的幅度和持续时间的Ca 2+内流。因此，我们假设：（1）ACh的作用增加了SNc DA和SNr GABA能神经元对来自PPN和PPN的谷氨酸能（glu）输入的反应，以及（2）除了来自PPN和PPN的glu和ACh兴奋性投射外，而且GABA能抑制向SN的投射（通过SNr神经元的阶段性抑制引起GABA系统的去抑制）调节SNc DA和SNr GABA能神经元放电行为。为了验证这些假设，我们将利用新开发的体外器官型培养制剂组成的PPN，SN，STH和STR进行形态学和电生理研究。形态学研究将涉及光学和电子显微镜，以（1）鉴定ACh、Glu和GABA能投射模式及其在SNc DA和SNr GABA能神经元上的终末位点，（2a）用联合生物胞素细胞内标记鉴定ACh和Glu投射到SNc DA和SNr GABA能神经元的核起源，和（2b）鉴定与SNc DA和SNr GABA神经元形成突触的细胞内标记的生物胞素终末的位置和（3）鉴定ACh的位置，Glu和GABA受体的SNc DA和SNr GABA神经元。电生理学研究将涉及使用全细胞（电流）钳或细胞内锐电极记录来描述（1）在PLSD和AHP期间胆碱能受体激活对Ca 2+内流的影响，（2）来自PPN的兴奋性突触输入和来自STR的GABA能抑制性输入对SNc DA和SNr GABA能神经元的放电特性和爆发的影响。了解多个输入在控制和/或调节影响其靶结构（即纹状体和丘脑）的SNc和SNr细胞活性中的作用对于运动和行为功能以及临床实体如帕金森病具有根本重要性。