Distinct nanoscale calcium channel and synaptic vesicle topographies contribute to the diversity of synaptic function

The nanoscale topographical arrangement of voltage-gated calcium channels (VGCC) and synaptic vesicles (SVs) determines synaptic strength and plasticity, but whether distinct spatial distributions underpin diversity of synaptic function is unknown. We performed single bouton Ca2+ imaging, Ca2+ chelator competition, immunogold electron microscopic (EM) localization of VGCCs and the active zone (AZ) protein Munc13-1, at two cerebellar synapses. Unexpectedly, we found that weak synapses exhibited 3-fold more VGCCs than strong synapses, while the coupling distance was 5-fold longer. Reaction-diffusion modelling could explain both functional and structural data with two strikingly different nanotopographical motifs: strong synapses are composed of SVs that are tightly coupled (~10 nm) to VGCC clusters, whereas at weak synapses VGCCs were excluded from the vicinity (~50 nm) of docked vesicles. The distinct VGCC-SV topographical motifs also confer differential sensitivity to neuromodulation. Thus VGCC-SV arrangements are not canonical and their diversity could underlie functional heterogeneity across CNS synapses.电压门控钙通道（voltage-gated calcium channels, VGCC）与突触囊泡（synaptic vesicles, SVs）的纳米级拓扑排布决定突触强度与可塑性，但目前尚不清楚独特的空间分布模式是否支撑了突触功能的多样性。我们在两类小脑突触中开展了单突触扣结钙离子成像、钙离子螯合剂竞争实验，以及电压门控钙通道与活性区（active zone, AZ）蛋白Munc13-1的免疫金电子显微镜（electron microscopic, EM）定位。出乎意料的是，我们发现弱突触的电压门控钙通道数量为强突触的3倍，而耦合距离则是强突触的5倍。反应扩散建模可通过两种截然不同的纳米拓扑模式阐释功能与结构数据：强突触由与电压门控钙通道簇紧密耦合（约10 nm）的突触囊泡构成，而弱突触的电压门控钙通道则被排除在锚定突触囊泡的邻近区域（约50 nm）之外。这种独特的电压门控钙通道-突触囊泡拓扑模式还会对神经调控产生差异化敏感性。综上，电压门控钙通道与突触囊泡的排布并非经典规范模式，其多样性或为中枢神经系统（central nervous system, CNS）突触间功能异质性的核心成因。