Role of neuronal and microglial identity in complement-dependent synaptic refinement – COMPLEMENT

Microglia are the resident macrophages of the brain. They fulfill multiple functions across development and adulthood and under disease conditions. Emerging evidence indicates that microglia display differences in their functions that are not exclusively driven by their milieu, rather by the unique properties these cells possess. Yet, the intrinsic heterogeneity of microglial cells has been largely overlooked. Recent studies suggest that distinct microglial “subtypes” perform unique functions, but the functional implications of the different microglial subtypes in the developing brain is still largely elusive. Microglial cells have a critical role in brain development. In particular, the complement system, a component of the innate immune system, remodels brain circuits in the developing thalamus by fostering the elimination of excess synapses by microglia, a process termed synaptic pruning. The role of the complement system in synaptic pruning in the cortex is controversial. The 6-layered neocortex of mammals contains several different types of pyramidal neurons, each type populating a specific layer. We have observed that the complement system eliminates synapses in layer 3 (L3) pyramidal neurons, but not in layer 5 (L5) pyramidal neurons of frontal cortex during early postnatal maturation. These intriguing results suggest that the complement-dependent refinement of cortical networks is neuron type-specific. Recent studies have demonstrated that cortical microglial cells interact with specific neuron subtypes via distinct ligand/receptor pairs, but the function of these interactions remains unknown. Here, we hypothesize that specific microglia subtypes target specific pyramidal neurons for complement-dependent synaptic pruning.
The project has three main goals. Firstly, we will seek to demonstrate that neuronal identity determines complement-dependent synaptic pruning. To achieve this aim, we will use in utero electroporation to respecify the identity of L3 pyramidal neurons and endow them with molecular and morphofunctional characteristics of L5 pyramidal neurons. Secondly, we will expand existing scRNA seq and spatial omics datasets of cortical microglia cells to identify ligand/receptor pairs which enable the interactions of distinct subtypes of neurons and microglial cells. Thirdly, we will alter the expression of distinct ligand/receptor pairs in neurons and microglial cells to identify microglia subtypes that interact specifically with L3, but not L5 pyramidal neurons, for synaptic pruning. For this we will use in utero electroporation to target specific neuronal cell populations, and novel tools including lipid nanoparticles and adeno-associated viral vectors to target microglial cells.