Chloë Trippaers

Pentameric glycine receptors (GlyRs) are key modulators of inhibitory neurotransmission, yet visualization of their activity across neuronal compartments has remained a challenge. Current methods that employ intracellularly tagged genetically encoded fluorescent proteins are prone to artefacts, as the tags can disrupt protein interactions that regulate receptor trafficking and positioning within the cell. We developed a novel, genetically encoded GlyRα2 activity reporter by fusing a chloride-sensitive fluorescent protein, mClYFP, to the extracellular N-terminus of GlyRα2. This chimeric receptor allows real-time nanoscopic visualization of the receptor and glycine-induced chloride concentration changes using total internal reflection fluorescence microscopy and ratio image analysis. Simultaneous electrophysiological and fluorescence measurements validated the functionality of both the ion channel and mClYFP components of our GlyRα2 activity reporter. The GlyRα2 ion channel characteristics are preserved, and the extracellular mClYFP tag reports chloride concentration changes in the physiological range. Therefore, mClYFP-GlyRα2 allowed us to detect receptor activity of chloride-permeable ionotropic receptors. In addition, we demonstrate that mClYFP-GlyRα2 can be effectively expressed in physiologically relevant striatal neurons. We present an extracellularly located, receptor-specific sensor that enables surface-accessible tracking of chloride ion dynamics in live cells. Our approach enables spatially resolved, non-invasive monitoring of chloride permeable receptor signaling, offering a powerful tool to investigate pentameric receptor function at the nanoscale.

Synaptic dysfunction is a hallmark of neurodevelopmental disorders (NDDs), often linked to genes involved in cytoskeletal regulation. While the role of these genes has been extensively studied in neurons, microglial functions such as phagocytosis are also dependent on cytoskeletal dynamics. We demonstrate that disturbance of actin cytoskeletal regulation in microglia, modeled by genetically impairing the scaffold protein Disrupted-in-Schizophrenia 1 (DISC1), which integrates actin-binding proteins, causes a shift in actin regulatory balance favoring filopodial versus lamellipodial actin organization. The resulting microglia-specific dysregulation of actin dynamics leads to excessive uptake of synaptic proteins. Genetically engineered DISC1-deficient mice show diminished hippocampal excitatory transmission and associated spatial memory deficits. Reintroducing wild-type microglia-like cells via bone marrow transplantation in adult DISC1-deficient mice restores the synaptic function of neurons and rescues cognitive performance. These findings reveal a pivotal role for microglial actin cytoskeletal remodeling in preserving synaptic integrity and cognitive health. Targeting microglial cytoskeletal dynamics may effectively address cognitive impairments associated with NDDs, even in adulthood.

Microglia activity can drive excessive synaptic loss during the prodromal phase of Alzheimer’s disease (AD) and is associated with lowered cyclic adenosine monophosphate (cAMP) due to cAMP phosphodiesterase 4B (PDE4B). This study aimed to investigate whether long-term inhibition of PDE4B by A33 (3 mg/kg/day) can prevent synapse loss and its associated cognitive decline in APPswe/PS1dE9 mice. This model is characterized by a chimeric mouse/human APP with the Swedish mutation and human PSEN1 lacking exon 9 (dE9), both under the control of the mouse prion protein promoter. The effects on cognitive function of prolonged A33 treatment from 20 days to 4 months of age, was assessed at 7–8 months. PDE4B inhibition significantly improved both the working and spatial memory of APPswe/PSdE9 mice after treatment ended. At the cellular level, in vitro inhibition of PDE4B induced microglial filopodia formation, suggesting that regulation of PDE4B activity can counteract microglia activation. Further research is needed to investigate if this could prevent microglia from adopting their ‘disease-associated microglia (DAM)’ phenotype in vivo. These findings support the possibility that PDE4B is a potential target in combating AD pathology and that early intervention using A33 may be a promising treatment strategy for AD.

Recent studies suggested that microglia, the primary brain immune cells, can affect circuit connectivity and neuronal function^1,2. Microglia infiltrate the neuroepithelium early in embryonic development and are maintained in the brain throughout adulthood^3,4. Several maternal environmental factors—such as an aberrant microbiome, immune activation and poor nutrition—can influence prenatal brain development^5,6. Nevertheless, it is unknown how changes in the prenatal environment instruct the developmental trajectory of infiltrating microglia, which in turn affect brain development and function. Here we show that, after maternal immune activation (MIA) in mice, microglia from the offspring have a long-lived decrease in immune reactivity (blunting) across the developmental trajectory. The blunted immune response was accompanied by changes in chromatin accessibility and reduced transcription factor occupancy of the open chromatin. Single-cell RNA-sequencing analysis revealed that MIA does not induce a distinct subpopulation but, rather, decreases the contribution to inflammatory microglia states. Prenatal replacement of microglia from MIA offspring with physiological infiltration of naive microglia ameliorated the immune blunting and restored a decrease in presynaptic vesicle release probability onto dopamine receptor type-two medium spiny neurons, indicating that aberrantly formed microglia due to an adverse prenatal environment affect the long-term microglia reactivity and proper striatal circuit development.