Microglia-dependent presynaptic disruption in an organotypic hippocampal slice culture model of neuroinflammation

Background: Systemic inflammation, such as occurs during sepsis, bone fracture, infections or post-operative trauma, has been linked to synapse loss and cognitive decline in human patients and animal models. Organotypic hippocampal slice cultures (OHSCs) represent an underused tool in neuroinflammation; retaining much of the neuronal architecture, synaptic connections and diversity of cell types present in the hippocampus in vivo whilst providing convenient access to manipulate and sample the culture medium and observe cellular reactions as in other in vitro methods. Here we report the development of an OHSC model of synaptic disruption after aseptic inflammation and investigate the underlying mechanism. Methods: OHSCs were generated from P6-P9 C57BL/6, the APP transgenic TgCRND8 model, or wild-type littermate mice according to the interface method. Aseptic inflammation was induced via addition of lipopolysaccharide (LPS) and cultures were analysed for changes in synaptic proteins via western blot. qPCR and ELISA analysis of the slice tissue and culture medium respectively determined changes in gene expression and protein secretion. Microglia were selectively depleted using the toxin clodronate and the effect of IL1β was assessed using a specific neutralising monoclonal antibody. Results: Addition of LPS caused a loss of the presynaptic protein synaptophysin via a mechanism dependent on microglia and involving IL1β. Washout of LPS via medium exchange allows for partial recovery of synaptic protein levels after 2 weeks. TgCRND8 OHSCs do not show alterations in IL1β expression at a timepoint where they exhibit spontaneous synaptophysin loss, and LPS does not alter levels of APP or Aβ in wild-type OHSCs. This indicates that although synaptophysin loss is seen in both systems, there is likely to be distinct underlying pathogenic mechanisms between the neuroinflammatory and amyloid models. Conclusions: We report the development of an OHSC model of LPS-induced synaptophysin loss and demonstrate a key role for microglia and involvement of IL1β. We propose that distinct molecular mechanisms lead to synaptophysin protein loss in LPS- exposed versus TgCRND8 OHSCs and provide a new experimental paradigm for assessing chronic changes in synaptic proteins, and synaptic plasticity, following acute inflammatory insults.