Abstract
Interleukin-1 beta (IL-1β) is a pro-inflammatory cytokine involved in the initiation of inflammation. In immune cells, this cytokine is synthesized as an immature precursor (pro-IL-1β) and requires enzymatic cleavage to become biologically active. For decades, it was thought that the role of IL-1β was limited to the regulation of inflammatory processes. However, a growing number of evidence supports the idea that, under homeostatic conditions, non-immune cells of the central nervous system (CNS) can express and/or be reactive to IL-1β. Among other things, this cytokine can modulate synaptic activity and network plasticity following sustained neuronal activity. These effects are mediated by IL-1 receptors, expressed at dendritic spines, which can physically and functionally interact with N-methyl-D-aspartate (NMDA) receptors. Based on these observations, we sought 1/ to characterize the cell type capable of producing IL-1β under homeostatic conditions in the CNS and 2/ to decipher the cellular mechanisms involved in the maturation and release of this cytokine.In this study, IL-1β expression was analyzed in mouse hippocampus using RNAscope, an in-situ hybridization technique. IL-1β mRNA was detected in hippocampal pyramidal neurons, specifically in the dentate gyrus and Cornu Ammonis 1 (CA1) areas. Under homeostatic conditions, approximately 40% of CA1 neurons express on average 0.7 molecules of mRNA encoding IL-1β. We also used a BRET biosensor to track IL-1β maturation in time and space. Expressed in cultured hippocampal excitatory neurons, this tool allowed us to identify that sustained neuronal activity leads to the maturation and release of this cytokine from neurons. Even if our data also show that neuronal IL-1β is preferentially localized in dendritic spines, the detection of BRET signals by microscopy shows that IL-1β cleavage can also occur indifferently in the cell body, dendrites or dendritic spines. IL-1β maturation is governed by NMDA receptor activation and involves the GluN2A and GluN2B subunits. Finally, by coupling biotinylation proximity labeling technology using the APEX2 enzyme and mass spectrometry analysis, we identified IL-1β-associated proteins in neurons in basal condition and after NMDA receptor activation. Our results identified a protease potentially responsible for neuronal IL-1β maturation and show an enrichment in proteins involved in vesicular transport and exocytosis mechanisms following NMDA stimulation.These results show that hippocampal excitatory neurons can synthesize, mature and release IL-1β under non-inflammatory conditions in the CNS. All the tools developed during this thesis will allow to decipher the cellular mechanisms associated with the functions of IL-1β under physiological conditions in the CNS but can also be used to better characterize the involvement of this cytokine in various physio-pathological processes throughout the organism.