Colocalization of synapse marker proteins evaluated by STED-microscopy reveals patterns of neuronal synapse distribution in vitro.

摘要: Abstract Background Quantification of synapses and their morphological analysis are extensively used in network development connectivity studies, drug screening other areas neuroscience. Thus, a number quantitative approaches were introduced so far. However, most the available methods highly tailored to specific applications have limitations for widespread use. New method We present new plugin open-source software ImageJ provide modifiable, high-throughput easy use synaptic puncta analysis. Our approach is based on colocalization pre- postsynaptic protein markers. Structurally completed glutamatergic GABAergic identified by VGLUT1-PSD95 VGAT-gephyrin colocalization, respectively. By combining conventional confocal microscopy with stimulated emission depletion (STED) imaging, we propose quantify scaffolding clusters, recruited single density. Results In proof-of-concept study, reveal differential distribution synapse density reference perineuronal net (PNN) expression. Using super-resolution STED demonstrate that composed significantly more compared uncompleted synapses. Comparison existing Synapse Counter offers rapid unbiased research tool broad spectrum neuroscientists. The proposed clusters quantification exploits imaging comprehensive composition. Conclusions results strongly substantiate benefits colocalization-based detection.