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Rôle des astrocytes dans la spasticité après traumatisme de la moelle épinière

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"Les astrocytes, ça sert à rien !" Si vous n'êtes pas d'accord avec ce dicton désuet, ce projet est fait pour vous!
Dans ce projet, il s'agira de mieux comprendre leur modification morpho-fonctionnelle dans un contexte pathologique de lésion de la moelle épinière chez la souris et de déterminer comment ses modifications post-lésionnelles influencent l'excitabilité des motoneurones spinaux. Nous nous focaliserons sur deux grandes fonctions astrocytaires : (1) le maintien de l'homéostasie potassique et glutamatergique et (2) la libération de gliotransmitteurs (ATP) modulant l'excitabilité neuronale. Ce projet permettra de mieux comprendre les mécanismes impliqués dans la spasticité qui est un trouble moteur très fréquent chez les patients médullo-lésés.

Description

State of the art: Spasticity is a common motor impairment after spinal cord injury (SCI). One of the main clinical manifestation is a chronic hypertonia associated with hyper-reflexia. Current treatments are limited mostly to supportive measures and there is no satisfactory cure. Thus, there is a critical need for a better understanding of cell interaction mechanisms to develop appropriately targeted repair strategies. Successive works from our group demonstrated that spasticity results from a excitatory/inhibitory imbalance of motoneurons (MN) in part due to an increase of the persistent Na+ current (Brocard et al. Nat. Med. 2016), concomitant with a disinhibition (Boulenguez et al. Nat. Med. 2010) both driven by calpain-I proteolytic activity (Plantier et al. Elife, 2019). However, the upstream initiators of this calpain-mediated excitatory/inhibitory imbalance are still not unknown. Although most of the proposed mechanisms focused on neuronal elements, it is now well appreciated that astrocytes actively modulate neuronal circuit excitability after CNS injury (Nimmerjahn and Bergles, 2015). Astrocytes are thus considered as active partners in neural information processing: (1) Reactive astrocytes interact with neighbouring neurons by releasing neuroactive substances such as D-serine and ATP (Koizumi et al. 2005). ATP release dynamically inhibits glutamatergic synaptic transmission (Witts and Miles, 2017) but display opposite effects in injured CNS (Coull et al. 2005). (2) Astrocytes also modulate neuronal excitability by taking up extrasynaptic glutamate (Gluo) and potassium (K+o) mainly via the glutamate transporter GLT-1 and the K+ channel Kir4.1, respectively (Sofroniew and Vinters, 2010). Although it has been shown that these two astrocytic proteins focally decrease after SCI in the surrounding sites of the lesion (Olsen et al. 2010), it is totally unknown if this decrease also occurs in the motor pool of MN and whether astrocytic glutamate and ATP release vary after SCI.

Objectives: The aim of this project will consist in determining to what extent the spinal astrocytes modulate the MN firing properties and contribute to the development of spasticity in SCI mice. Aim 1 (cellular level, spinal slice): Determine the astrocyte-MN crosstalk mechanisms responsible of MN hyperexcitability (impaired K+/glutamate homeostasis and increased gliotransmitters release). Aim 2 (microcircuit level, whole-mount spinal cord): Image and modulate the reactive astrocytic activity and gliotransmitters spillover surrounding the MN pool from SCI mice. Aim 3 (systems level, in vivo): Recording hindlimb muscles activity after targeting astrocytes with gene therapy to alleviate spasticity.

Methods: Multi-scaled approaches based on two−photon calcium imaging (astrocytic activity), electrophysiology (patch-clamp of motoneurons/astrocytes, extracellular ventral root recordings, EMGs), single-cell RT-PCR (astrocyte), opto-genetics (ChR2-H134R, eArchT3.0) and AAVs injection from transgenic mice lines (ALDH1L1-eGFP, floxed Gcamp6f, ALDH1L1-CreERT2, Chat-Cre) will be used.

Expected results: We hypothesize that synergistic action of (1) astrocytic impaired K+ and glutamate clearance with (2) increased release of glutamate/ATP by reactive astrocytes contribute to the excitatory/inhibitory imbalance of extensor MNs following SCI.

Profil recherché

Les candidat.e.s motivé.e.s, curieux/curieuses, rigoureux/ses seront privilégié.e.s. Un intérêt sur la thématique de la physiologie spinale et du dialogue neurone-astrocytaire est fortement recommandé. Une expérience préalable avec les techniques d'électrophysiologie (type patch-clamp) sera considérée comme un plus.

Établissement d'accueil

http://www.int.univ-amu.fr/

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