Synaptic integration and intrinsic firing properties of basal ganglia neurons

我们实验室的工作重点是位于中脑的多巴胺释放神经元的整合和兴奋性的细胞和亚细胞原理。在最近的一个项目中，我们研究了位于多巴胺神经元树突上的脊髓突触的特性，并比较了脊突和干突触对兴奋性的相对贡献。在这项研究之前，关于树突棘存在的证据是混合的，还没有研究中脑多巴胺神经元树突棘中的电和钙信号的功能研究。在表达荧光标记的PSD95的小鼠系中，使用双光子谷氨酸注入成像来定位谷氨酸能突触，我们发现多巴胺神经元表达功能性脊椎，突触EPSP的大小与PSD-95的存在呈正相关。最后，我们发现，多巴胺神经元特有的缓慢起搏放电与由于颈椎狭窄而引起的脊髓电位的增加相结合，产生了一种新的脊髓钙离子增强，这种增强发生在从棘波周期的中后期的一个窗口中。这项研究的结果于2015年发表在《eLife》杂志上。未来的工作将测试这种起搏器诱发的钙增强的离子机制，并探索其对突触可塑性的影响。 该实验室的其他主要兴趣包括1)识别功能和生化上独特的中脑多巴胺神经元亚群，以及2)了解这些神经元如何融入基底节回路。为此，一个项目比较了投射到伏隔核和背侧纹状体的多巴胺神经元亚群对诱发和突触产生的GABA能抑制的反应。中脑多巴胺神经元暂停其放电，以回应奖赏遗漏或厌恶刺激。我们的结果表明，刺激GABA能输入或模拟抑制的超极化电流注射所诱发的多巴胺神经元放电暂停，可被在低于峰阈值的电压下招募的一类钾电导增强。重要的是，这些记录在中伏隔神经元的A型钾电流表现出相当慢的失活动力学，与第二电导、超极化激活电导或Ih的较弱表达相结合，相对于黑质纹状体神经元，超极化诱导的峰电位延迟延长。鉴于最近的解剖学研究发现，多巴胺神经元亚群共享很大程度上重叠的输入，这些结果表明，这些输入的整合在不同的多巴胺神经元中不同，相对于黑质纹状体神经元，这一特征可能对厌恶信号具有重要意义。 第二个项目研究了黑质内多巴胺神经元的功能异质性。众所周知，在帕金森氏病中易受细胞死亡影响的SNC内，存在不同的脆弱和弹性神经元亚群，可以根据它们对钙结合蛋白Calbindin的表达来区分它们。我们发现，脆弱的钙结合蛋白缺乏的神经元和弹性的钙结合蛋白阳性的多巴胺能神经元在生理、钙信号、树突分支和兴奋性突触传递方面存在本质上的差异。有趣的是，我们发现钙结合蛋白缺乏的神经元表现出低阈值的去极化，并伴随着树突状钙的大量增加。我们发现这种钙离子是通过电压门控的一类称为T-型钙离子的钙通道进入的，我们目前正在探索这一通道对钙结合蛋白缺乏细胞的生理作用。 最后，我们参加了今年的一个合作项目，产生了两个出版物。其中一个项目是与哈佛医学院的Bruce Bean博士合作，研究了导致黑质多巴胺能神经元动作电位复极化的钾通道。该项目于2015年发表在《神经科学杂志》上。与Ellen Sidransky博士的实验室的第二次合作研究了葡萄糖脑苷酶如何影响帕金森症。这项研究的一篇手稿最近作为一篇文章发表在2016年的《神经科学杂志》上。

The work in our laboratory focuses on cellular and subcellular principles of integration and excitability in dopamine-releasing neurons located in the midbrain. In a recent project, we examined the properties of spine synapses located on the dendrites of dopamine neurons, and compared the relative contribution of spine and shaft synapses to excitability. Prior to this study, evidence for the presence of dendritic spines had been mixed and there had been no functional study examining electrical and Ca2+ signaling in dendritic spines of midbrain dopamine neurons. Using two-photon uncaging of glutamate with imaging in a mouse line that expresses fluorescently tagged-PSD95 to locate glutamatergic synapses, we found that dopamine neurons express functional spines and that the size of synaptic EPSPs correlated positively with the presence of PSD-95. Lastly, we found that the characteristic slow pacemaker firing of dopamine neurons combines with boosting of spine potentials due to the narrow spine neck to produce a novel enhancement of spine Ca2+ that occurs periodically in a window from the middle to the late phase of the spike cycle. The results of this study were published in Elife in 2015. Future work will test the ionic mechanism of this pacemaker-evoked Ca enhancement and explore its implications for synaptic plasticity. Other major interests of the lab include 1) identifying functionally and biochemically unique subpopulations of midbrain dopamine neurons and 2) understanding how these neurons fit into the basal ganglia circuit. To this end, one project compared the responses to evoked and synaptically-generated GABAergic inhibition of dopamine neurons subpopulations projecting to nucleus accumbens and dorsal striatum. Midbrain dopamine neurons pause their firing in response to reward omission or aversive stimuli. Our results show that pauses in dopamine neuron firing, evoked either by stimulation of GABAergic inputs or by hyperpolarizing current injections mimicking inhibition, were enhanced by a subclass of potassium conductances recruited at voltages below spike threshold. Importantly, these A-type potassium currents recorded in mesoaccumbal neurons displayed substantially slower inactivation kinetics which, combined with weaker expression of a second conductance, hyperpolarization-activated conductance or Ih, lengthened hyperpolarization-induced delays in spiking relative to nigrostriatal neurons. Given recent anatomical studies that find that dopamine neuron subpopulations share largely overlapping inputs, these results suggest that integration of these inputs differs among dopamine neurons favoring higher sensitivity to inhibition in mesoaccumbal relative to nigrostriatal neurons, a feature that may be important for aversive signaling. A second project examined functional heterogeneity of dopamine neurons within the substantia nigra. It is known that within the SNc, which is susceptible to cell death in Parkinsons Disease, there are distinct subpopulations of vulnerable and resilient neurons that can be distinguished according to their expression of the calcium (Ca2+) binding protein, calbindin. We found that that vulnerable calbindin-lacking neurons and resilient calbindin-positive dopaminergic neurons differ substantially in their physiology, calcium signaling, dendritic branching, and excitatory synaptic transmission. Interestingly, we found that calbindin-lacking neurons display low-threshold depolarizations that were accompanied by a large increase in dendritic calcium. We found that this Ca2+enters through a subclass of voltage-gated Ca channels called T-type Ca2+ and we are currently exploring the contribution of this channel to physiology of calbindin-lacking cells. Lastly, we participated in a collaborative projects this year which resulted in two publications. One project, a collaboration with Dr. Bruce Bean at Harvard Medical School, examined the potassium channels that contribute to repolarization of action potentials in SNc dopaminergic neurons. This project was published in the Journal of Neuroscience in 2015. A second collaboration with the laboratory of Dr. Ellen Sidransky examined how glucocerebrosidase impacts parkinsonism. A manuscript of this study was recently published as an article in the Journal of Neuroscience in 2016.