Imagerie optique de la plasticité synaptique
|Advisor:||De Koninck, Paul; Méthot, Mario|
|Abstract:||Classical measurements of synaptic plasticity have involved electrophysiological methods which provide high sensitivity for detecting small changes in synaptic strength. However, this approach does not provide much information about the location of the synapses that undergo plastic changes. Because synaptic plasticity can be synapse-specific, having the ability to monitor changes in synaptic strength at individual synapses is important in order to enable simultaneously monitoring of local molecular mechanisms associated with the plasticity. New fluorescent tools developed in the last decades allow to directly visualize synaptic activity, signaling, and remodeling at individual synapses. During my Master studies, I used optical imaging of a genetically-encoded calcium (Ca2+) sensor, GCaMP6f, to record miniature synaptic Ca2+ transients (MSCTs) in cultured rat hippocampal neurons. For these experiments, I performed video-microscopy on neurons perfused with external solution lacking Mg2+ and containing Tetrodotoxin (0Mg2+/TTX). I have observed highly localized and transient increases of intracellular Ca2+ in dendritic compartments and spines. To test whether these MSCTs can be potentiated, I have measured them before and after a 5 min stimulation known to induce plasticity in cultured neurons (0Mg2+/Glycine/Bicuculline, cLTP). A lasting increase in the frequency and amplitude of MSCTs, for at least an hour, arose from this stimulation protocol. I have thus investigated the molecular mechanisms of this plasticity. The MSCTs are mostly mediated by NMDA receptors, since they are almost totally blocked by the selective antagonist to the receptor, AP5. Moreover, addition of AP5 only during the cLTP stimulation blocks the MSCT plasticity. It thus appears that both the MSCTs and their plasticity are NMDA receptor-dependent. Interestingly, the MSCTs and their plasticity are not blocked by the AMPA receptor antagonists NBQX, pointing to possible changes in NMDA receptor content, postsynaptic Ca2+ signaling, or presynaptic neurotransmitter release. Also, while we found that CaMKII signaling is non-essential for the induction of the plasticity, preliminary data are showing a plausible PKA-dependency of the plasticity. To test these hypotheses, I have also tried to combine Ca2+ imaging with imaging of other pre and postsynaptic components, to identify the molecular mechanisms responsible for the MSCT plasticity. Overall, this new approach presented in this thesis might provide new knowledge on the diversity of molecular processes that support synaptic potentiation.|
|Document Type:||Mémoire de maîtrise|
|Open Access Date:||24 April 2018|
|Collection:||Thèses et mémoires|
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