2 research outputs found
Introducing Diinamic, a flexible and robust method for clustering analysis in single-molecule localization microscopy
International audienceAbstract Super-resolution microscopy allowed major improvements in our capacity to describe and explain biological organization at the nanoscale. Single-molecule localization microscopy (SMLM) uses the positions of molecules to create super-resolved images, but it can also provide new insights into the organization of molecules through appropriate pointillistic analyses that fully exploit the sparse nature of SMLM data. However, the main drawback of SMLM is the lack of analytical tools easily applicable to the diverse types of data that can arise from biological samples. Typically, a cloud of detections may be a cluster of molecules or not depending on the local density of detections, but also on the size of molecules themselves, the labeling technique, the photo-physics of the fluorophore, and the imaging conditions. We aimed to set an easy-to-use clustering analysis protocol adaptable to different types of data. Here, we introduce Diinamic, which combines different density-based analyses and optional thresholding to facilitate the detection of clusters. On simulated or real SMLM data, Diinamic correctly identified clusters of different sizes and densities, being performant even in noisy datasets with multiple detections per fluorophore. It also detected subdomains (“nanodomains”) in clusters with non-homogeneous distribution of detections
Conformational state-dependent regulation of GABAA receptor diffusion and subsynaptic domains
International audienceThe efficacy of GABAergic synapses relies on the number of postsynaptic GABAA receptors (GABAARs), which is regulated by a diffusion capture mechanism. Here, we report that the conformational state of GABAARs influences their membrane dynamics. Indeed, pharmacological and mutational manipulations of receptor favoring active or desensitized states altered GABAAR diffusion leading to the disorganization of GABAAR subsynaptic domains and gephyrin scaffold, as detected by super-resolution microscopy. Active and desensitized receptors were confined to perisynaptic endocytic zones, and some of them were further internalized. We propose that following their activation or desensitization, synaptic receptors rapidly diffuse at the periphery of the synapse where they remain confined until they switch back to a resting state or are internalized. We speculate that this allows a renewal of activatable receptors at the synapse, contributing to maintain the efficacy of the synaptic transmission, in particular on sustained GABA transmission