Biology subjects

Chastagnier, Y.

Publications and source records attributed to Chastagnier, Y..

3 recordsLinked to original sources

bioRxiv2024

Optical sectioning for reflection interference microscopy

Reflection Interference Contrast Microscopy (RICM, also known as interference reflection microscopy) and related techniques have become of wide interest to the biophysical, soft matter and biochemistry communities owing to their exquisite sensitivity for characterising thin films or individual nanoscopic objects adsorbed onto surfaces, or for monitoring cell-substrate interactions. Over the recent years, striking progresses have been made to improve the sensitivity and the quantitative analysis of RICM. Its use in more complex environments, with spurious reflections stemming from a variety of structures in the sample, remains however challenging. In this paper, we demonstrate two optical sectioning methods that effectively reduce such background and can be readily implemented in a conventional RICM setup: line confocal detection, and structured illumination microscopy. We characterise experimentally the benefits to image quality and demonstrate the use of the methods for quantitative imaging of complex biological and biomimetic samples: cellular membranes, thin organic films, surface biofunctionalization. We then discuss the benefits of each method and provide guidelines to arbitrate between sectioning and signal-to-noise ratio. Finally, we provide a detailed description of our experimental setup and a home-written image acquisition and processing software that should allow the interested reader to duplicate such a setup on a home-built or commercial microscope.

biophysics↗

bioRxiv2023

Voltage tunes mGlu5 receptor function, impacting synaptic transmission

Voltage sensitivity is a common feature of many membrane proteins, including some G-protein coupled receptors (GPCRs). However, the functional consequences of voltage sensitivity in GPCRs are not well understood. In this study, we investigated the voltage sensitivity of the post-synaptic metabotropic glutamate receptor mGlu5 and its impact on synaptic transmission. Using biosensors and electrophysiological recordings in non-excitable HEK293T cells or neurons, we found that mGlu5 receptor function is optimal at resting membrane potentials. We observed that membrane depolarization significantly reduced mGlu5 receptor activation, Gq-PLC/PKC stimulation, Ca2+ release, and mGlu5 receptor-gated currents through TRPC6 channels or NMDA receptors. Notably, we report a previously unknown activity of the NMDA receptor at the resting potential of neurons, enabled by mGlu5. Our findings suggest that mGlu5 receptor activity is directly regulated by membrane voltage which may have a significant impact on synaptic processes and pathophysiological functions.

neuroscience↗

bioRxiv2020

Restoring Glutamate receptosome dynamics at synapses rescues Autism-like deficits in Shank3-deficient mice.

Shank3 monogenic mutations lead to Autism Spectrum Disorders (ASD). Shank3 is part of the glutamate receptosome that physically links ionotropic NMDA receptors to metabotropic mGlu5 receptors through interactions with scaffolding proteins PSD95-GKAP-Shank3-Homer. A main physiological function of the glutamate receptosome is to control NMDA synaptic function that is required for plasticity induction. Intact glutamate receptosome supports glutamate receptors activation and plasticity induction, while glutamate receptosome disruption blocks receptors activity, preventing the induction of subsequent plasticity. Despite possible impact on metaplasticity and cognitive behaviors, scaffold interaction dynamics and their consequences are poorly defined. Here we used mGlu5-Homer interaction as a biosensor of glutamate receptosome integrity to report changes of NMDA synaptic function. Combining BRET imaging and electrophysiology, we show that a transient neuronal depolarization inducing NMDA-dependent plasticity disrupts glutamate receptosome in a long-lasting manner at synapses and induces signaling required for the expression of the initiated neuronal plasticity such as ERK and mTOR pathways. Glutamate receptosome disruption also decreases NMDA/AMPA currents ratio, freezing the sensitivity of the synapse to subsequent changes of neuronal activity. These data show the importance of a fine-tuning of protein-protein interactions within glutamate receptosome, driven by changes of neuronal activity, to control plasticity. In a mouse model of ASD, a truncated mutant form of Shank3 prevents the integrity of the glutamate receptosome. These mice display altered plasticity, anxiety-like and stereotyped behaviors. Interestingly, repairing the integrity of glutamate receptosome and its sensitivity to the neuronal activity rescued synaptic transmission, plasticity and some behavioral traits of Shank3{triangleup}C mice. Altogether, our findings characterize mechanisms by which Shank3 mutations cause ASD and highlight scaffold dynamics as new therapeutic target.

neuroscience↗