Synaptic integration and intrinsic firing properties of basal ganglia neurons

黑质多巴胺（SNc）神经元以低速率紧张性地放电，导致动作电位反向传播。在我们实验室最近完成的一项研究中，我们发现背景紧张性放电的速率对树突定位的电导和突触驱动的放电反应产生了强大的影响。使用双光子谷氨酸释放和突触刺激，我们表明，快速紧张性放电（4 - 5 Hz）加强突触驱动的突发放电输出在NMDA受体依赖的方式。此外，我们发现，从1 Hz到6 Hz的强直率的增加显着增强了平均树突状细胞的钙信号，与一些细胞表现出高达五倍的线性预测的钙值增加。与SNc神经元相比，腹侧被盖区（VTA）多巴胺神经元显示出更多的线性增加的Ca与速率表明SNc神经元中的游离Ca的更高的负载。该研究已提交并已发表（Hage和Khaliq，2015）。 继续研究中脑多巴胺神经元之间的功能异质性的兴趣，我们有两个项目接近完成，确定重要的差异，在阈下电导形状的放电和树突兴奋性的多巴胺神经元亚群。在第一项研究中，我们比较了投射到前额叶皮层、中脑核和背侧纹状体的逆行标记多巴胺神经元。通过比较SNc亚群（黑质纹状体神经元），我们发现，腹侧被盖区亚群（mesocortical和mesoporbal神经元）表现出更强的敏感性和更长的延迟，以尖峰发作后超极化刺激。这种不同的敏感性是由于存在一个缓慢衰减的钾电流，更强烈地表达在腹侧被盖区多巴胺神经元。我们的计划是测试这一发现对多巴胺神经元亚群之间的尖峰时间和同步性的影响。在第二项研究中，我们研究了SNc多巴胺神经元的树突兴奋性。在过去的电生理学研究中，SNc神经元被认为是比VTA神经元更同质的群体。然而，研究这些细胞的树突钙信号，我们发现SNc神经元之间阈下反应的强度和强度存在显着差异。特别是，我们发现，SNc多巴胺神经元的主要部分表现出非线性去极化突发样反应诱发和突触输入。这些非线性响应由树突状T型Ca通道驱动，并且令人惊讶的是，倾向于几乎完全存在于钙结合蛋白阴性的SNc多巴胺能神经元中。这一发现具有潜在的临床相关性，因为帕金森病患者中钙结合蛋白阴性的多巴胺神经元往往会选择性死亡。沿着我们最近发表的结果（Hage和Khaliq，2015），这项研究支持了Ca电导在塑造SNc多巴胺能神经元的生理学方面发挥更主导作用的观点。 在我们对多巴胺神经元的突触生理学感兴趣之后，我们一直在研究多巴胺神经元上树突棘的特性。树突棘存在的解剖学证据与一些观察到棘状突起存在的研究相混合，而其他研究则得出多巴胺神经元主要是刺状的结论。我们发现，多巴胺神经元明确表达棘在幼年和成年小鼠。我们已经证实，棘存在于快速灌注的组织制备以及高尔基染色的组织。此外，我们已经测试了功能使用的组合钙成像在脊柱头部和谷氨酸解开。这些实验表明，它们确实表达突触受体，并可能形成功能性突触。我们研究了棘长度对棘功能的影响，目前正在比较棘与树突轴突触的整合。该项目已接近完成，我们计划在不久的将来提交。最后，我们参与了与Ellen Sidransky博士实验室的合作项目。最近提交了这项研究的手稿，目前正在审查中。

Substantia nigra dopamine (SNc) neurons fire tonically at low rates resulting in action potential backpropagation. In a recently completed study from our lab, we found that the rate of background tonic firing exerts a powerful influence over dendritically-located conductances and synaptically-driven firing responses. Using two-photon glutamate-uncaging and synaptic stimulation, we show that fast tonic firing (4-5 Hz) intensifies synaptically-driven burst firing output in an NMDA-receptor dependent manner. Furthermore, we found that increasing the tonic rate from 1 to 6 Hz dramatically enhanced the average dendritic Ca signals, with some cells showing up to five-fold increase over linearly predicted Ca values. By contrast to SNc neurons, ventral tegmental area (VTA) dopamine neurons showed much more linear increases in Ca with rate suggesting a higher load of free Ca in SNc neurons. This study was submitted and has been published (Hage and Khaliq, 2015). Continuing our interest in examining functional heterogeneity among midbrain dopamine neurons, we have two projects nearing completion that identify important differences in subthreshold conductances that shape firing and dendritic excitability in subpopulations of dopamine neurons. In the first study, we compared retrogradely-labeled dopamine neurons that project to prefrontal cortex, nucleus accumbens and dorsal striatum. By comparison to SNc subpopulations (nigrostriatal neurons), we find that VTA subpopulations (mesocortical and mesoaccumbal neurons) show stronger sensitivity and longer delays to spike onset following hyperpolarizing stimuli. This differential sensitivity is due to the presence of a slowly decaying potassium current that is more strongly expressed in VTA dopamine neurons. Our plan is to test the implications of this finding on spike timing and synchrony among dopamine neuron subpopulations. In a second study, we examine dendritic excitability in SNc dopamine neurons. In past electrophysiological studies, SNc neurons were considered a much more homogeneous population than VTA neurons. Studying dendritic Ca signals of these cells, however, we have discovered dramatic differences in the strength and intensity of subthreshold responses in among SNc neurons. In particular, we find that a major fraction of SNc dopamine neurons exhibits non-linear depolarizing burst-like responses to evoked and synaptic inputs. These non-linear responses are driven by dendritic T-type Ca channels, and surprisingly, tend to be present almost exclusively in calbindin-negative SNc dopaminergic neurons. This finding has potential clinical relevance because calbindin-negative dopamine neurons tend to selectively die in Parkinsons patients. Along with our recently published results (Hage and Khaliq, 2015), this study supports the idea that Ca conductances play a much more dominant role in shaping the physiology of SNc dopaminergic neurons. Following our interest in the synaptic physiology of dopamine neurons, we have been examining the properties of dendritic spines on dopamine neurons. Anatomical evidence for the presence of dendritic spines has been mixed with some studies observing the presence of spine-like processes while others conclude that dopamine neurons are largely aspiny. We find that dopamine neurons clearly express spines in both juvenile and adult mice. We have confirmed that spines are present in fast perfused tissue preparations as well as golgi-stained tissue. Furthermore, we have tested the functionality using a combination of Ca imaging in the spine head and glutamate uncaging. These experiments show that they indeed express synaptic receptors and likely form functional synapses. We examined the effect of spine length on the function of spines and are currently comparing integration onto spines versus onto dendritic shaft synapses. This project is nearly completed and we plan to submit in the near future. Lastly, we participated in a collaborative project with the laboratory of Dr. Ellen Sidransky. A manuscript of this study was recently submitted and is under review.