Synaptic integration and intrinsic firing properties of basal ganglia neurons

基底节神经元的突触整合和内在放电特性

• 批准号: 8557101
• 负责人: ZAYD M KHALIQ
• 金额: $ 121.28万
• 依托单位: NATIONAL INSTITUTE OF NEUROLOGICAL DISORDERS AND STROKE
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/8557101
• 关键词: Action Potentials; Address; Back; Basal Ganglia; Behavior; Brain; Brain Diseases; Brain Stem; Breeding; Calcium; Calcium Oscillations; Calcium Signaling; Cell Nucleus; Dendrites; Dependence; Depressed mood; Distal; Dopamine; Fire - disasters; Frequencies; Functional disorder; Gated Ion Channel; Glutamates; Goals; Image; Imaging Techniques; Immunohistochemistry; Laboratories; Laser Scanning Microscopy; Learning; Lesion; Light; Link; Measurable; Membrane; Mental Depression; Midbrain structure; Motor; Movement; Mus; N-Methyl-D-Aspartate Receptors; Neurons; Output; Parkinson Disease; Patch-Clamp Techniques; Pattern; Play; Population; Positioning Attribute; Postdoctoral Fellow; Principal Investigator; Property; Research; Rewards; Role; Schizophrenia; Slice; Somatic Cell; Source; Stimulus; Structure of subthalamic nucleus; Substantia nigra structure; Synapses; Synaptic Transmission; Synaptic plasticity; System; Testing; Time; Training; Transgenic Mice; addiction; base; cholinergic; cholinergic neuron; dopaminergic neuron; member; neuronal cell body; pars compacta; patch clamp; posters; research study; transmission process; two-photon; voltage

Dopamine neurons are key players in the reward and motor systems. They fire tonically at low rates of 2-5 Hz and fire bursts of action potentials at 15-20 Hz during reward-relevant behaviors. Bursting is not an intrinsic property of dopamine neurons but depends critically on synaptic inputs. How excitatory and modulatory inputs facilitate burst firing in dopamine neurons is unknown. We are testing the hypothesis that burst firing depends on both fast synaptic transmission and slow, metabotropic glutamatergic transmission coming from the subthalamic nucleus as well cholinergic inputs from the brainstem pedunculopontine nucleus. Because the lab is newly established, we have only started to make progress on the proposed aims. We have made good progress in setting up and staffing the laboratory. There are currently four (4) members of the lab including the Principal Investigator (one postdoctoral fellow, a graduate fellow, and a Post-Baccalaureate IRTA). We have constructed three patch-clamp setups which we use daily for brain slice recordings. We also have a separate setup dedicated to two-photon laser scanning microscopy (2PLSM) which we use for simultaneous calcium-imaging of the dendrites and patch-clamp recordings from dopamine neurons. Our initial studies test the contribution of excitatory input from the subthalamic nucleus (STN) to synaptically evoked high-frequency firing in substantia nigra pars compacta (SNc) dopamine neurons. To start, we are examining short-term synaptic plasticity of the STN to SNc synapse. STN neurons fire repetitively at high tonic rates (20-30 Hz) but fire bursts of spikes up to 200 Hz for short periods of time (100-200 ms). We discovered that there is little short-term plasticity across a broad range of frequencies (2-100 Hz), with synapses depressing only slightly (max 35%) for short bursts of 5 stimuli. We have also shown that slightly more depression (about 40% on average) occurs during long trains of 60 stimuli at 20 Hz. Despite this initial depression, however, synaptic release is reasonably maintained at tonic rates, suggesting that tonic, sustained input from STN neurons can possibly have a significant impact on the excitability of SNc dopamine neurons. The postbacc fellow involved with this project presented a poster of this material at the Summer Intern Poster Day 2012 in Natcher Auditorium. The brainstem pendunculopontine nucleus (PPN) provides the sole source of cholinergic input to SNc and activation this nucleus stimulates burst firing in dopamine neurons. Likewise, lesioning the PPN limits bursting in dopamine neurons suggesting that cholinergic input is necessary for burst firing in SNc dopamine neurons. Therefore, it is possible that cholinergic input enhances the impact of excitatory input from other sources such as the STN, for example, through depolarization and relief of Mg2+ block of NMDA receptors. To test this hypothesis, we have obtained transgenic mice that express the light-activated molecule, channelrhodopsin, only in cholinergic neurons (ChAT-ChR2). We are current breeding the mice and these studies are just starting to get underway. Dopamine neurons are capable of releasing dopamine not only from terminals but also from dendrites into the neighboring substantia nigra pars reticulata (SNr), the main output nucleus of the basal ganglia. As such, dendrites of dopamine neurons are positioned to have a major impact on basal ganglia function. Somatodendritic release of dopamine occurs in a calcium-dependent manner and calcium influx is likely determined by the interaction of synaptic inputs and dendritic excitability. We are beginning to examine calcium signals in the dendrites dopamine neurons using two-photon imaging techniques. So far, we have probed calcium signals in proximal to distal dendrites during spontaneous action potential firing, which we record simultaneously from whole-cell somatic recordings. In our limited number of preliminary experiments, we have consistently found measurable calcium oscillations linked to somatic firing throughout the proximal as well distal dendrites at distances up to 500 um away from the soma. This suggests that there is reliable back-propagation of action potential even throughout the distal dendrites, thus maximizing the influence of somatodendritically released dopamine in the SNr. Since high-frequency burst firing in dopamine neurons is thought to enhance dopamine released from terminals, we have also begun to examine the activity-dependence of calcium signals in the dendrites of dopamine neurons during high-frequency firing.

多巴胺神经元是奖励和运动系统的关键角色。在与奖励相关的行为期间，它们以 2-5 Hz 的低频率发射，并以 15-20 Hz 的频率发射动作电位。爆发不是多巴胺神经元的固有特性，而是很大程度上取决于突触输入。兴奋性和调节性输入如何促进多巴胺神经元的爆发性放电尚不清楚。我们正在测试这样的假设：突发放电取决于来自丘脑底核的快速突触传递和缓慢的代谢性谷氨酸能传递以及来自脑干脚桥核的胆碱能输入。由于实验室是新成立的，我们才刚刚开始在拟议的目标上取得进展。 我们在实验室的设立和人员配备方面取得了良好进展。该实验室目前有四 (4) 名成员，包括首席研究员（一名博士后研究员、一名研究生和一名学士后 IRTA）。我们构建了三个膜片钳装置，每天用于脑切片记录。我们还有一个专门用于双光子激光扫描显微镜（2PLSM）的单独装置，我们用它来同时对树突进行钙成像，并进行多巴胺神经元的膜片钳记录。 我们的初步研究测试了丘脑底核（STN）的兴奋性输入对黑质致密部（SNc）多巴胺神经元突触诱发高频放电的贡献。首先，我们正在研究 STN 到 SNc 突触的短期突触可塑性。 STN 神经元以高紧张率 (20-30 Hz) 重复放电，但会在短时间内 (100-200 ms) 爆发高达 200 Hz 的尖峰。我们发现，在较宽的频率范围（2-100 Hz）内几乎没有短期可塑性，对于 5 次刺激的短脉冲，突触仅轻微抑制（最大 35%）。我们还发现，在 20 Hz 的 60 次刺激的长序列中，抑郁症的发生率略高（平均约为 40%）。然而，尽管最初存在这种抑制，但突触释放仍以强直速率合理维持，这表明来自 STN 神经元的强直、持续输入可能对 SNc 多巴胺神经元的兴奋性产生重大影响。 参与该项目的 postbacc 研究员在 Natcher 礼堂举行的 2012 年暑期实习生海报日上展示了该材料的海报。 脑干桥桥核 (PPN) 为 SNc 提供胆碱能输入的唯一来源，激活该核会刺激多巴胺神经元的爆发放电。同样，损伤 PPN 会限制多巴胺神经元的爆发，这表明胆碱能输入对于 SNc 多巴胺神经元的爆发是必要的。因此，胆碱能输入可能会增强来自其他来源（例如 STN）的兴奋性输入的影响，例如通过去极化和缓解 NMDA 受体的 Mg2+ 阻断。 为了验证这一假设，我们获得了仅在胆碱能神经元（ChAT-ChR2）中表达光激活分子视紫红质的转基因小鼠。我们目前正在培育小鼠，这些研究才刚刚开始。 多巴胺神经元不仅能够从末端释放多巴胺，还能够从树突释放多巴胺到邻近的黑质网状部（SNr），即基底神经节的主要输出核。因此，多巴胺神经元的树突对基底神经节功能产生重大影响。多巴胺的体细胞树突释放以钙依赖性方式发生，钙流入可能由突触输入和树突兴奋性的相互作用决定。我们开始使用双光子成像技术检查树突多巴胺神经元中的钙信号。到目前为止，我们已经探测了自发动作电位放电过程中近端到远端树突的钙信号，我们从全细胞体细胞记录中同时记录了这些信号。在我们有限数量的初步实验中，我们一致发现可测量的钙振荡与体细胞放电相关，遍及近端和远端树突，距体体距离可达 500 微米。这表明即使在整个远端树突中，动作电位也存在可靠的反向传播，从而使 SNr 中体细胞树突释放的多巴胺的影响最大化。由于多巴胺神经元的高频突发放电被认为可以增强末梢释放的多巴胺，因此我们也开始研究高频放电期间多巴胺神经元树突中钙信号的活动依赖性。