Howard Hughes Medical Institute, Oregon Health and Science University, Portland, United States.
Vollum Institute, Oregon Health and Science University, Portland, United States.
Elife. 2024 Nov 4;13:RP98458. doi: 10.7554/eLife.98458.
Chemical synapses are the major sites of communication between neurons in the nervous system and mediate either excitatory or inhibitory signaling. At excitatory synapses, glutamate is the primary neurotransmitter and upon release from presynaptic vesicles, is detected by postsynaptic glutamate receptors, which include ionotropic AMPA and NMDA receptors. Here, we have developed methods to identify glutamatergic synapses in brain tissue slices, label AMPA receptors with small gold nanoparticles (AuNPs), and prepare lamella for cryo-electron tomography studies. The targeted imaging of glutamatergic synapses in the lamella is facilitated by fluorescent pre- and postsynaptic signatures, and the subsequent tomograms allow for the identification of key features of chemical synapses, including synaptic vesicles, the synaptic cleft, and AuNP-labeled AMPA receptors. These methods pave the way for imaging brain regions at high resolution, using unstained, unfixed samples preserved under near-native conditions.
化学突触是神经系统中神经元之间主要的通讯位点，介导兴奋性或抑制性信号传递。在兴奋性突触中，谷氨酸是主要的神经递质，在从突触前囊泡释放后，被突触后谷氨酸受体检测到，包括离子型 AMPA 和 NMDA 受体。在这里，我们开发了在脑组织切片中识别谷氨酸能突触的方法，用小金纳米颗粒（AuNPs）标记 AMPA 受体，并准备薄片进行冷冻电子断层扫描研究。通过荧光前突触和后突触标记物，靶向成像薄片中的谷氨酸能突触，随后的断层扫描允许识别化学突触的关键特征，包括突触小泡、突触间隙和 AuNP 标记的 AMPA 受体。这些方法为使用未经染色、未经固定的、在接近天然状态下保存的样本，以高分辨率成像脑区铺平了道路。
