Periodic Reporting for period 3 - MICRO-COPS (Microglia Control of Physiological Brain States)
Periodo di rendicontazione: 2024-06-01 al 2025-11-30
Microglia (MG) are specialized innate immune cells of the brain. They survey the healthy brain and interact with neurons to modulate brain function. During development, as the brain is built, they remove organelles, cell sub-compartments, and entire cells to shape circuits. In the diseased brain, they attack pathogens, remove debris, and stage inflammatory responses. The Micro-COPS consortium studies the fundamental biology of the neuron-microglia system. The main focus is on how MG execute physiological 'non-immune' functions as modulators of normal neuronal activity, brain circuits, and behaviour, and on how the perturbation of these physiological processes can cause neurological and psychiatric disorders. Our overall objectives are to determine the molecular mechanisms by which reciprocal MG-neuron signalling occurs, to examine how MG affect the function of neurons and their synapses, and to resolve how neuronal activity in specific brain regions shapes the resident MG population for specific functions.
Micro-COPS developed multiple complex new approaches to study MG and neuron biology - e.g. (i) an electrophysiological secretion assay based on light-mediated calcium uncaging and cell capacitance measurements, (ii) a cell lineage tracing assay to assess the origin and fate of MG cells and the effect of neuronal activity, (iii) novel microscopic and imaging methods to study MG biology, (iv) a theoretical framework that links the function of synapses between nerve cells to the number of transmitter receptors, or (v) a novel method to visualize optogenetically stimulated synapses in the EM, which allows to compare the ultrastructure of potentiated synapses to their neighbours on the same dendrite or axon. Further, Micro-COPS generated multiple new genetically modified mouse lines that (i) allow to follow gene expression and transcriptional control in MG, (ii) modulate levels of intracellular second messengers, or (iii) allow to optogenetically induce changes of the intracellular second messenger cAMP in MG. Micro-COPS identified a novel, activity-dependent, neuromodulatory feedback process, where MG sense neuronal activity via neuronal transmitters and in turn suppress neuronal activity. This mechanism protects the brain from excessive activation, may play a major role in maintaining physiological sleep behaviour, and may regulate neuronal responses to sensory inputs. Indeed, this process plays a key role in maintaining neuronal activity-induced changes in blood flow. Micro-COPS also identified a novel MG population in a brain region called the striatum, which controls movements and behaviour in response to the release of the neurotransmitter dopamine. This MG subpopulation expresses the dopamine D1 receptor and plays a critical role in dopamine-induced neuronal function and dopamine-dependent behaviours, such as addiction. Furthermore, Micro-COPS discovered that MG release the messenger TNFα and thereby regulate the function and plasticity of synapses between nerve cells. This process involves massive changes in the modification of cellular proteins via phosphorylation and plays a key role in the control of sleep. In addition, we developed a mouse model that expresses Gq-DREADD under the control of an inducible MG-specific promoter. This model allows to selectively activate MG without affecting other brain cell types. Using these mice, Micro-COPS discovered that specific activation of MG profoundly affects the synapses between nerve cells, specifically their stability, and thereby controls learning and memory. Finally, we found that MG Gq activation protects mice against stroke-related tissue damage and improves behavioural recovery. This finding has major relevance for our understanding of stroke-related brain damage and potential new therapeutic approaches.
Micro-COPS innovated the field by identifying a novel, activity-dependent regulatory feedback of neuromodulation by MG, by which MG can sense neuronal activation and respond to it by suppressing synaptic transmission and neuronal activity. Our findings indicate that this MG-driven negative feedback mechanism operates like inhibitory neurons and plays an essential role in protecting the striatum and other brain regions from excessive neuronal activation. In contrast to pharmacological activation of MG in tissue, our chemogenetic activation strategy - expression of Gq-DREADD under the control of an inducible MG-specific promoter - is truly specific to MG. MG are switched to an activated state in the absence of any danger signals in the tissue. This clearly separates cause and effect, so that all downstream effects on neurons and synapses can be traced back to microglia-neuron communication. We discovered that MG regulate the plasticity of cortical inhibitory synapses through purinergic signalling via P2RX7, mediated by MG-derived TNFα. A key underlying mechanism involves TNFα-mediated modulation of protein phosphorylation during sleep. These findings reveal a fascinating new role of MG in sleep homeostasis. Our approach to combine non-invasive, optogenetic plasticity-induction at defined synapses in intact tissue with electron-dense labels allow to find and analyse the rare potentiated synapses among the many thousands of non-potentiated synapses. This breaks new ground in the field of synaptic plasticity. Likewise, the observation that Gq activation in MG protects neurons against stroke-triggered defects is highly unexpected and potentially medically important - representing a nice example of a tool that had been developed for basic research and now provides new insights into the mechanisms of a human disease.