少突胶质细胞,中枢神经系统(CNS)的产生髓鞘的神经胶质细胞,对髓鞘形成和回路功能至关重要。越来越多的证据表明,少突胶质细胞中的细胞内钙(Ca2)动力学介导了活性依赖性和非依赖性髓鞘形成。解开髓鞘少突胶质细胞如何协调和整合Ca2+信号,特别是关于轴突放电,对于深入了解它们在中枢神经系统发育和功能中的作用至关重要,无论是健康还是疾病。在这个框架中,我们使用重组腺相关病毒(rAAV)/Olig001衣壳变体表达遗传编码的钙(Ca2)指示剂jGCaMP8s,在髓磷脂碱性蛋白(MBP)启动子的控制下。在我们的研究中,该工具对髓鞘化和成熟少突胶质细胞表现出优异的向性和选择性,它允许监测髓鞘形成细胞中的Ca2+活性,在分离的原代培养物和器官型脊髓外植体中。通过对器官培养物中少突胶质细胞中的髓鞘Ca2事件进行实时成像,我们观察到在不同的体外发育阶段,Ca2事件的幅度和持续时间迅速下降。活跃的髓鞘重塑和生长通过Ca2+信号在髓鞘轴突界面水平上被调节,在器官培养的早期髓鞘形成过程中,这个阶段是通过发射轴突动作电位来微调的。在髓鞘形成的后期,成熟少突胶质细胞中的Ca2+事件不再显示这样的调节,强调复杂的Ca2+信号参与中枢神经系统髓鞘形成。意义声明确定成熟少突胶质细胞中驱动Ca2+事件的来源和机制,通常通过限制性转基因品系进行研究,事实证明是具有挑战性的。为了解决这个问题,我们使用rAAV/Olig001选择性表达jGCaMP8s,在MBP启动子的转录控制下,在体外和离体监测髓鞘化和成熟少突胶质细胞中的Ca2+活性。我们的发现表明,Ca2+动力学经历成熟依赖性调制,并且神经元活动可以在各个发育阶段对Ca2活性产生不同的影响。我们的研究引入了一种有价值的遗传工具,用于监测髓鞘形成细胞中的Ca2信号传导,以研究Ca2调节如何影响少突胶质细胞功能以及与轴突的动态相互作用。
Oligodendrocytes, the myelin-producing glial cells of the central nervous system (CNS), crucially contribute to myelination and circuit function. An increasing amount of evidence suggests that intracellular calcium (Ca2+) dynamics in oligodendrocytes mediates activity-dependent and activity-independent myelination. Unraveling how myelinating oligodendrocytes orchestrate and integrate Ca2+ signals, particularly in relation to axonal firing, is crucial for gaining insights into their role in the CNS development and function, both in health and disease. In this framework, we used the recombinant adeno-associated virus/Olig001 capsid variant to express the genetically encoded Ca2+ indicator jGCaMP8s, under the control of the myelin basic protein promoter. In our study, this tool exhibits excellent tropism and selectivity for myelinating and mature oligodendrocytes, and it allows monitoring Ca2+ activity in myelin-forming cells, both in isolated primary cultures and organotypic spinal cord explants. By live imaging of myelin Ca2+ events in oligodendrocytes within organ cultures, we observed a rapid decline in the amplitude and duration of Ca2+ events across different in vitro developmental stages. Active myelin sheath remodeling and growth are modulated at the level of myelin-axon interface through Ca2+ signaling, and, during early myelination in organ cultures, this phase is finely tuned by the firing of axon action potentials. In the later stages of myelination, Ca2+ events in mature oligodendrocytes no longer display such a modulation, underscoring the involvement of complex Ca2+ signaling in CNS myelination.