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Etude en microscopie de fluorescence à deux photons in vivo de l'intégration multi vibrissale chez le rat

Julien Bertherat 1
1 Dynamique corticale et mécanismes de codage - équipe de l'IBENS
IBENS - Institut de biologie de l'ENS Paris (UMR 8197/1024)
Abstract : Rats possess an array of long whiskers, they can actively move and which enable them to explore space, localize objects and discriminate textures. Sensory information coming from follicles' mecano receptors where whiskers are inserted goes through mesencephalic and thalamic relays and reaches discrete units, called "barrels", in layer IV of the primary somato-sensory cortex. Then, sensory information spreads to layer II/III of the primary somato-sensory cortex where horizontal connections enable integration of information coming from several whiskers. My work tackles the question of multi whisker integration in the somato-sensory cortex using in vivo two photon microscopy and covers mainly two chapters. I first deal with all the optical developments in two photon microscopy that were realized in order to optimize in vivo recordings of the neuronal activity. Then, I consider the question of multi whisker integration using this technique. I developed an original two photon microscopy set-up which is able to record neuronal activity in layer II/III of the cortex while giving access to the recorded neuronal network architecture. That microscope allows high speed detection of single calcium transiants while requiring low power from the excitation laser. Thus, both photo bleaching and photo toxicity are limited. Furthermore, I identified noise sources during in vivo recordings and implemented appropriate solutions (holding system, image movement correction algorithm...) to reduce or correct these noises in order to be limited only by shot noise. Finally, I evaluated individual cells recordings quality excluding all risk of important contamination by neuropil signal. Using that two photon microscope, I have studied the neuronal responses in layer II/III to different stimulations. First, applying multi-whisker stimulations showing either no or full correlations between the whiskers, I have shown that neurons are spatially segregated depending on their response to these multi-whisker stimuli, this segregation being related to the below barrel organization in layer IV. Then, I have looked at neuronal responses induced by single whisker deflections as a function of two parameters: the stimulus "phase" in the 2D space of preferred stimuli and the stimulus direction in the 2D physical space. I have shown the absence of strong correlation between "phase" and direction tunings.
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Julien Bertherat. Etude en microscopie de fluorescence à deux photons in vivo de l'intégration multi vibrissale chez le rat. Neurosciences. Ecole Normale Supérieure de Paris - ENS Paris, 2012. Français. ⟨tel-00789937⟩

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