Mannal Nadja, Kleiner Katharina, Fauler Michael, Dougalis Antonios, Poetschke Christina, Liss Birgit
Institute of Applied Physiology, University of Ulm, Ulm, Germany.
Linacre and New College, University of Oxford, Oxford, United Kingdom.
Front Synaptic Neurosci. 2021 Feb 26;13:635050. doi: 10.3389/fnsyn.2021.635050. eCollection 2021.
Dopaminergic (DA) midbrain neurons within the substantia nigra (SN) display an autonomous pacemaker activity that is crucial for dopamine release and voluntary movement control. Their progressive degeneration is a hallmark of Parkinson's disease. Their metabolically demanding activity-mode affects Ca homeostasis, elevates metabolic stress, and renders SN DA neurons particularly vulnerable to degenerative stressors. Accordingly, their activity is regulated by complex mechanisms, notably by dopamine itself, via inhibitory D2-autoreceptors and the neuroprotective neuronal Ca sensor NCS-1. Analyzing regulation of SN DA neuron activity-pattern is complicated by their high vulnerability. We studied this activity and its control by dopamine, NCS-1, and glucose with extracellular multi-electrode array (MEA) recordings from midbrain slices of juvenile and adult mice. Our tailored MEA- and spike sorting-protocols allowed high throughput and long recording times. According to individual dopamine-responses, we identified two distinct SN cell-types, in similar frequency: dopamine-inhibited and dopamine-excited neurons. Dopamine-excited neurons were either silent in the absence of dopamine, or they displayed pacemaker-activities, similar to that of dopamine-inhibited neurons. Inhibition of pacemaker-activity by dopamine is typical for SN DA neurons, and it can undergo prominent desensitization. We show for adult mice, that the number of SN DA neurons with desensitized dopamine-inhibition was increased (60-100%) by a knockout of NCS-1, or by prevention of NCS-1 binding to D2-autoreceptors, while time-course and degrees of desensitization were not altered. The number of neurons with desensitized D2-responses was also higher (65%) at high glucose-levels (25 mM), compared to lower glucose (2.5 mM), while again desensitization-kinetics were unaltered. However, spontaneous firing-rates were significantly higher at high glucose-levels (~20%). Moreover, transient glucose-deprivation (1 mM) induced a fast and fully-reversible pacemaker frequency reduction. To directly address and quantify glucose-sensing properties of SN DA neurons, we continuously monitored their electrical activity, while altering extracellular glucose concentrations stepwise from 0.5 mM up to 25 mM. SN DA neurons were excited by glucose, with EC values ranging from 0.35 to 2.3 mM. In conclusion, we identified a novel, common subtype of dopamine-excited SN neurons, and a complex, joint regulation of dopamine-inhibited neurons by dopamine and glucose, within the range of physiological brain glucose-levels.
黑质（SN）内的多巴胺能（DA）中脑神经元表现出自主起搏器活动，这对于多巴胺释放和自主运动控制至关重要。它们的渐进性退化是帕金森病的一个标志。它们对代谢要求很高的活动模式会影响钙稳态，增加代谢应激，并使SN DA神经元特别容易受到退化应激源的影响。因此，它们的活动受到复杂机制的调节，特别是多巴胺本身通过抑制性D2自受体和神经保护性神经元钙传感器NCS-1进行调节。由于SN DA神经元高度脆弱，分析其活动模式的调节变得复杂。我们通过对幼年和成年小鼠中脑切片进行细胞外多电极阵列（MEA）记录，研究了这种活动及其受多巴胺、NCS-1和葡萄糖的控制。我们定制的MEA和尖峰分类方案实现了高通量和长时间记录。根据个体对多巴胺的反应，我们以相似的频率识别出两种不同的SN细胞类型：多巴胺抑制型和多巴胺兴奋型神经元。多巴胺兴奋型神经元在没有多巴胺时要么沉默，要么表现出起搏器活动，类似于多巴胺抑制型神经元。多巴胺对起搏器活动的抑制是SN DA神经元的典型特征，并且可能会发生显著的脱敏。我们发现，对于成年小鼠，通过敲除NCS-1或阻止NCS-1与D2自受体结合，多巴胺抑制脱敏的SN DA神经元数量增加了约60%-100%，而脱敏的时间进程和程度没有改变。与较低葡萄糖水平（2.5 mM）相比，在高葡萄糖水平（25 mM）下，D2反应脱敏的神经元数量也更高（约65%），而脱敏动力学同样未改变。然而，在高葡萄糖水平下，自发放电率显著更高（约20%）。此外，短暂的葡萄糖剥夺（1 mM）会导致起搏器频率快速且完全可逆地降低。为了直接研究和量化SN DA神经元的葡萄糖感应特性，我们连续监测它们的电活动，将细胞外葡萄糖浓度从0.5 mM逐步提高到25 mM。SN DA神经元对葡萄糖有兴奋反应，其EC值范围为0.35至2.3 mM。总之，我们在生理性脑葡萄糖水平范围内，识别出一种新的、常见的多巴胺兴奋型SN神经元亚型，以及多巴胺和葡萄糖对多巴胺抑制型神经元的复杂联合调节。
