Influence des facteurs sécrétés par les oligodendrocytes sur la physiologie des neurones GABAergiques

Les interactions entre les neurones et les oligodendrocytes assurent de nombreuses fonctions essentielles à la maturation des circuits corticaux. A travers la formation de myéline, les oligodendrocytes augmente considérablement la vitesse et la fiabilité de propagation des potentiels d’action. Ce phénomène repose sur l’agrégation des canaux sodiques voltage-dépendants aux nœuds de Ranvier. Bien que certains travaux aient attribué un rôle clé aux contacts entre les oligodendrocytes et l’axone dans la formation des nœuds de Ranvier, notre laboratoire a démontré que des facteurs sécrétés par les oligodendrocytes permettent l’agrégation de protéines nodales le long des axones GABAergiques de l’hippocampe avant la myélinisation, formant ainsi des prénoeuds. Au cours de ma thèse, nous avons identifié les facteurs agrégeants en un complexe multimoléculaire constitué de la Contactine-1, en association avec Tenascine-R et Phosphacan. Afin d’identifier les cibles neuronales des oligodendrocytes, nous avons ensuite étudié l’influence des facteurs sécrétés par les oligodendrocytes sur la physiologie des neurones GABAergiques. La combinaison d’analyses électrophysiologiques et transcriptomiques suggère que les facteurs sécrétés par les oligodendrocytes tendent à atténuer les effets causés par l’élimination des cellules gliales des cultures. L’absence de cellules gliales diminue la fréquence de décharge des potentiels d’action et le nombre d’événements excitateurs reçus par les neurones GABAergiques. Des changements spécifiques d’expression génique sont également observés et partiellement restaurés par l’ajout de milieu conditionné d’oligodendrocytes.Oligodendrocyte interactions with neurons fulfill distinct functions essential for cortical circuit maturation, albeit not completely understood. Through myelin formation, oligodendrocytes greatly increase speed and reliability of action potential propagation. This process requires high densities of voltage-gated sodium channels at nodes of Ranvier. Mechanisms underlying their assembly remain poorly understood. While evidences have attributed a key role to contacts between myelinating oligodendrocytes and the axon, our laboratory suggested that nodes of Ranvier formation may differ between neuronal subtypes. Oligodendrocyte secreted factors are indeed sufficient to cluster nodal proteins along hippocampal GABAergic axons prior to myelination, thereby forming node-like clusters or prenodes. Yet, the nature of the factors involved remained elusive. During my thesis, we identified the oligodendrocyte-derived clustering factors as a multimolecular complex, consisting of Contactin-1 combined with Phosphacan or Tenascin-R. To identify neuronal targets of oligodendrocytes, we investigated the influence of oligodendrocyte secreted factors on hippocampal GABAergic neuron physiology. Combination of electrophysiological and transcriptomic analyses suggested that oligodendrocyte secreted cues tend to reverse the observed changes in GABAergic neuron physiology caused by removing glial cells from cultures. Absence of glial cells decreases action potential discharge and excitatory events received by GABAergic neurons. Specific changes in transcripts for ion channels, transporters and synaptic markers were also induced and partly restored upon addition of oligodendrocyte secreted factors

Neuron–Oligodendrocyte Communication in Myelination of Cortical GABAergic Cells

Axonal myelination by oligodendrocytes increases the speed and reliability of action potential propagation, and so plays a pivotal role in cortical information processing. The extent and profile of myelination vary between different cortical layers and groups of neurons. Two subtypes of cortical GABAergic neurons are myelinated: fast-spiking parvalbumin-expressing cells and somatostatin-containing cells. The expression of pre-nodes on the axon of these inhibitory cells before myelination illuminates communication between oligodendrocytes and neurons. We explore the consequences of myelination for action potential propagation, for patterns of neuronal connectivity and for the expression of behavioral plasticity

Influence des facteurs sécrétés par les oligodendrocytes sur la physiologie des neurones GABAergiques

Oligodendrocyte interactions with neurons fulfill distinct functions essential for cortical circuit maturation, albeit not completely understood. Through myelin formation, oligodendrocytes greatly increase speed and reliability of action potential propagation. This process requires high densities of voltage-gated sodium channels at nodes of Ranvier. Mechanisms underlying their assembly remain poorly understood. While evidences have attributed a key role to contacts between myelinating oligodendrocytes and the axon, our laboratory suggested that nodes of Ranvier formation may differ between neuronal subtypes. Oligodendrocyte secreted factors are indeed sufficient to cluster nodal proteins along hippocampal GABAergic axons prior to myelination, thereby forming node-like clusters or prenodes. Yet, the nature of the factors involved remained elusive. During my thesis, we identified the oligodendrocyte-derived clustering factors as a multimolecular complex, consisting of Contactin-1 combined with Phosphacan or Tenascin-R. To identify neuronal targets of oligodendrocytes, we investigated the influence of oligodendrocyte secreted factors on hippocampal GABAergic neuron physiology. Combination of electrophysiological and transcriptomic analyses suggested that oligodendrocyte secreted cues tend to reverse the observed changes in GABAergic neuron physiology caused by removing glial cells from cultures. Absence of glial cells decreases action potential discharge and excitatory events received by GABAergic neurons. Specific changes in transcripts for ion channels, transporters and synaptic markers were also induced and partly restored upon addition of oligodendrocyte secreted factors.Les interactions entre les neurones et les oligodendrocytes assurent de nombreuses fonctions essentielles à la maturation des circuits corticaux. A travers la formation de myéline, les oligodendrocytes augmente considérablement la vitesse et la fiabilité de propagation des potentiels d’action. Ce phénomène repose sur l’agrégation des canaux sodiques voltage-dépendants aux nœuds de Ranvier. Bien que certains travaux aient attribué un rôle clé aux contacts entre les oligodendrocytes et l’axone dans la formation des nœuds de Ranvier, notre laboratoire a démontré que des facteurs sécrétés par les oligodendrocytes permettent l’agrégation de protéines nodales le long des axones GABAergiques de l’hippocampe avant la myélinisation, formant ainsi des prénoeuds. Au cours de ma thèse, nous avons identifié les facteurs agrégeants en un complexe multimoléculaire constitué de la Contactine-1, en association avec Tenascine-R et Phosphacan. Afin d’identifier les cibles neuronales des oligodendrocytes, nous avons ensuite étudié l’influence des facteurs sécrétés par les oligodendrocytes sur la physiologie des neurones GABAergiques. La combinaison d’analyses électrophysiologiques et transcriptomiques suggère que les facteurs sécrétés par les oligodendrocytes tendent à atténuer les effets causés par l’élimination des cellules gliales des cultures. L’absence de cellules gliales diminue la fréquence de décharge des potentiels d’action et le nombre d’événements excitateurs reçus par les neurones GABAergiques. Des changements spécifiques d’expression génique sont également observés et partiellement restaurés par l’ajout de milieu conditionné d’oligodendrocytes

NOUVEAUX MÉCANISMES DE L’IMPLICATION DES OLIGODENDROCYTES DANS LA CONDUCTION DE L’INFLUX NERVEUX

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Neuron–Oligodendrocyte Communication in Myelination of Cortical GABAergic Cells

International audienceAxonal myelination by oligodendrocytes increases the speed and reliability of action potential propagation, and so plays a pivotal role in cortical information processing. The extent and profile of myelination vary between different cortical layers and groups of neurons. Two subtypes of cortical GABAergic neurons are myelinated: fast-spiking parvalbumin-expressing cells and somatostatin-containing cells. The expression of pre-nodes on the axon of these inhibitory cells before myelination illuminates communication between oligodendrocytes and neurons. We explore the consequences of myelination for action potential propagation, for patterns of neuronal connectivity and for the expression of behavioral plasticity

Generation of Oligodendrocytes and Oligodendrocyte-Conditioned Medium for Co-Culture Experiments

International audienceIn the central nervous system, oligodendrocytes are well-known for their role in axon myelination, that accelerates the propagation of action potentials through saltatory conduction. Moreover, an increasing number of reports suggest that oligodendrocytes interact with neurons beyond myelination, notably through the secretion of soluble factors. Here, we present a detailed protocol allowing purification of oligodendroglial lineage cells from glial cell cultures also containing astrocytes and microglial cells. The method relies on overnight shaking at 37 °C, which allows selective detachment of the overlying oligodendroglial cells and microglial cells, and the elimination of microglia by differential adhesion. We then describe the culture of oligodendrocytes and production of oligodendrocyte-conditioned medium (OCM). We also provide the kinetics of OCM treatment or oligodendrocytes addition to purified hippocampal neurons in co-culture experiments, studying oligodendrocyte-neuron interactions

Oligodendrocyte secreted factors shape hippocampal GABAergic neuron transcriptome and physiology

International audienceOligodendrocytes form myelin for central nervous system axons and release factors which signal to neurons during myelination. Here, we ask how oligodendroglial factors influence hippocampal GABAergic neuron physiology. In mixed hippocampal cultures, GABAergic neurons fired action potentials (APs) of short duration and received high frequencies of excitatory synaptic events. In purified neuronal cultures without glial cells, GABAergic neuron excitability increased and the frequency of synaptic events decreased. These effects were largely reversed by adding oligodendrocyte conditioned medium (OCM). We compared the transcriptomic signature with the electrophysiological phenotype of single neurons in these three culture conditions. Genes expressed by single pyramidal or GABAergic neurons largely conformed to expected cell-type specific patterns. Multiple genes of GABAergic neurons were significantly downregulated by the transition from mixed cultures containing glial cells to purified neuronal cultures. Levels of these genes were restored by the addition of OCM to purified cultures. Clustering genes with similar changes in expression between different culture conditions revealed processes affected by oligodendroglial factors. Enriched genes are linked to roles in synapse assembly, AP generation, and transmembrane ion transport, including of zinc. These results provide new insight into the molecular targets by which oligodendrocytes influence neuron excitability and synaptic function

Role of a Contactin multi‐molecular complex secreted by oligodendrocytes in nodal protein clustering in the CNS

International audienceThe fast and reliable propagation of action potentials along myelinated fibers relies on the clustering of voltage-gated sodium channels at nodes of Ranvier. Axo-glial communication is required for assembly of nodal proteins in the central nervous system, yet the underlying mechanisms remain poorly understood. Oligodendrocytes are known to support node of Ranvier assembly through paranodal junction formation. In addition, the formation of early nodal protein clusters (or prenodes) along axons prior to myelination has been reported, and can be induced by oligodendrocyte conditioned medium (OCM). Our recent work on cultured hippocampal neurons showed that OCM-induced prenodes are associated with an increased conduction velocity (Freeman et al., 2015). We here unravel the nature of the oligodendroglial secreted factors. Mass spectrometry analysis of OCM identified several candidate proteins (i.e., Contactin-1, ChL1, NrCAM, Noelin2, RPTP/Phosphacan, and Tenascin-R). We show that Contactin-1 combined with RPTP/Phosphacan or Tenascin-R induces clusters of nodal proteins along hippocampal GABAergic axons. Furthermore, Contactin-1-immunodepleted OCM or OCM from Cntn1-null mice display significantly reduced clustering activity, that is restored by addition of soluble Contactin-1. Altogether, our results identify Contactin-1 secreted by oligodendrocytes as a novel factor that may influence early steps of nodal sodium channel cluster formation along specific axon populations