After 2 years, come to meet us in person at Cell Bio 2022!
Booth #1636: December 4 – 6, 2022, Washington D.C.
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Learn about our micropatterning, microfabrication, or hydrogel structuration process for a wide range of applications: cell force measurement, disease modeling, cell migration, microfluidics, organ-on-a-chip, spheroïd formation, 3D cell culture, cryo-ET …
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Poster presentation at Cell Bio 2022 on Tuesday December 6th
POSTER # B415/P2801: ENGINEERED IN VITRO MODELS BASED ON SELECTIVE NEURONAL ADHESION TO STUDY SYNAPSE FORMATION, N. PIETTE, P. STRALE, M. MUNIER, M. SAINLOS, I. CHAMMA, O. THOUMINE, V. STUDER; IINS-CNRS BORDEAUX, FRANCE
Information transfer in the brain is ensured by a complex network of specialized interconnected neurons responding to environmental cues. The proper functioning of this network relies on the establishment and maintenance of synaptic connections, nanoscale organization of their molecular components, and plasticity of their architecture. It is well known that these mechanisms are regulated by specific transsynaptic adhesion proteins linked to membrane receptors or scaffolding proteins.
We present here two cell based in vitro models that we have specifically engineered to study the biophysical properties of trans-synaptic adhesion proteins and their role in the nanoscale organization of synapses during development. These models rely on micropatterns of synaptic proteins printed on glass coverslips. They were fabricated with a commercially available contactless and maskless UV projection system which allows us to reproducibly control the shape and the protein density of the microcontact areas.
In the first model, recombinant Fc-tagged neuronal protein (e.g. Neurexin1β) are anchored to 5µm wide patterned lines and presented to COS-7 cells expressing fluorescent tagged paired protein (e.g. Neuroligin1) providing highly selective recognition. By using fluorescence recovery after photobleaching and single particle tracking we are able to measure the diffusion coefficient and the interaction rate of trans-synaptic adhesion complexes, respectively. We also show how to use this model to measure the interaction between trans-membrane synaptic proteins and cytosolic partners (e.g. PSD95).
The second model consists in surfaces micropatterned with arrays of small dots (5µm diameter) coated with Fc-tagged neuronal ligands. When cultured on these substrates, primary neurons form thousands of standardized hemi-synapses after 14 days in vitro. We show that we can precisely locate patches of pre or post synaptic markers as well as the cytoskeleton within micropatterned areas. By averaging the super-resolved STORM (Stochastic Optical Reconstruction Microscopy) image of hundreds of these standardized hemi synapses for each developmental stage, we hope to get a new insight on the sequence of events leading from the creation of an initial contact to the formation of stable synapses.
Together, these two in vitro models will allow us to screen and characterize the role of synaptic proteins in the context of synaptogenesis. We also hope to get a better understanding of the role of synaptic adhesion proteins in the nanoscale organization of the synapse during development.