Using cryo-electron tomography (cryo-ET) to study the molecular architecture of synapses

Neurons communicate at specialized terminals called synapses, where hundreds of molecules work together in harmony to ensure proper and efficient signal transmission, which is crucial for normal brain functions. Aberrant synaptic signalling is involved in many neurodevelopmental and neurodegenerative diseases such as autism and Parkinson’s disease. This highlights the need to understand the organization and interactions of molecular complexes at the synapse. Cryo-electron tomography (cryo-ET) offers a close-to-native preservation of biological specimen and the unique possibility to capture both the cellular context and the molecular details, and thus can be utilized to advance structural understanding of the synapse. However, it is challenging to access sufficiently thin synapses for high-resolution cryo-ET. We address this problem by employing cryo-focused-ion beam milling to produce electron transparent slices containing intact synapses in cultured neurons. We obtained high-quality cryo-tomograms of synapses, showing unprecedented molecular details. In addition, we also apply correlative light and electron microscopy and preliminary results show that fluorescence labelling is useful in discerning synaptic elements in the crowded cryo-electron micrographs and 3D-tomograms. These technical developments pave the way for studies on the molecular architecture of synapses, such as the structural basis of synaptic plasticity.