Blaum Natalie, Ghelani Tina, Götz Torsten W B, Chronister Keagan S, Bengochea Mercedes, Ceresnova Livia, Christensen Christian F, Moulin Thiago C, Kern Hanna, Thomas Ulrich, Heine Martin, Sigrist Stephan J, Walter Alexander M
Molecular and Theoretical Neuroscience, Department of Neuroscience, Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen 2200, Denmark.
Molecular and Theoretical Neuroscience, Leibniz-Forschungsinstitut für Molekulare Pharmakologie, Berlin 13125, Germany.
Proc Natl Acad Sci U S A. 2025 Aug 26;122(34):e2514151122. doi: 10.1073/pnas.2514151122. Epub 2025 Aug 20.
Neuromodulators control mood, arousal, and behavior by inducing synaptic plasticity via G-protein-coupled receptors. While long-term presynaptic potentiation requires structural changes, mechanisms enabling potentiation within minutes remain unclear. Using the neuromuscular junction, we show that octopamine, the invertebrate analog of norepinephrine, potentiates evoked neurotransmitter release on the timescale of one minute via a G-protein-coupled pathway involving presynaptic OAMB receptors and phospholipase C. This fast potentiation correlates with elevated signals of the release factor Unc13A and the scaffolding protein Bruchpilot. Live, single-molecule imaging of endogenously tagged Unc13 revealed its instantly reduced motility and increased concentration in synaptic nanoclusters with potentiation. Presynaptic knockdown of Unc13A fully blocked fast potentiation. Moreover, deleting its N-terminal localization sequence mislocalized the protein fragment to the cytosol, but still allowed for rapid plasma membrane recruitment by diacylglycerol (DAG) analog phorbol esters and octopamine, implicating a role of more C-terminal domains. A point mutation of endogenous Unc13 in its DAG-binding C1 domain blocked plasticity-induced nanoscopic enrichment and synaptic potentiation. The mutation increased basal neurotransmission but reduced Unc13 levels, revealing a gain of function and potential homeostatic compensation. The mutation also blocked phorbol ester-induced potentiation, decreased the calcium sensitivity of neurotransmission, and caused short-term synaptic depression. Homeostatic potentiation induced by postsynaptic receptor block mirrored octopamine-induced Unc13 recruitment and required presynaptic OAMB receptors, indicating overlapping machinery. Thus, rapid Unc13 immobilization and nanoscale compaction are salient features of fast presynaptic potentiation.
神经调质通过G蛋白偶联受体诱导突触可塑性来控制情绪、觉醒和行为。虽然长期的突触前增强需要结构变化，但在几分钟内实现增强的机制仍不清楚。利用神经肌肉接头，我们发现章鱼胺（去甲肾上腺素的无脊椎动物类似物）通过涉及突触前OAMB受体和磷脂酶C的G蛋白偶联途径，在一分钟的时间尺度上增强诱发的神经递质释放。这种快速增强与释放因子Unc13A和支架蛋白Bruchpilot的信号升高相关。对内源标记的Unc13进行实时单分子成像显示，随着增强，其运动性立即降低，在突触纳米簇中的浓度增加。Unc13A的突触前敲低完全阻断了快速增强。此外，删除其N端定位序列会使蛋白片段错误定位于细胞质，但仍允许二酰基甘油（DAG）类似物佛波酯和章鱼胺快速招募到质膜，这暗示了更多C端结构域的作用。内源性Unc13在其DAG结合C1结构域中的点突变阻断了可塑性诱导的纳米级富集和突触增强。该突变增加了基础神经传递，但降低了Unc13水平，揭示了功能获得和潜在的稳态补偿。该突变还阻断了佛波酯诱导的增强，降低了神经传递的钙敏感性，并导致短期突触抑制。突触后受体阻断诱导的稳态增强反映了章鱼胺诱导的Unc13招募，并且需要突触前OAMB受体，表明机制重叠。因此，快速的Unc13固定和纳米级压实是快速突触前增强的显著特征。
