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Abstract Microglia are sexually dimorphic, yet, this critical aspect is often overlooked in neuroscientific studies. Decades of research have revealed the dynamic nature of microglial-neuronal interactions, but seldom consider how this dynamism varies with microglial sex differences, leaving a significant gap in our knowledge. This study focuses on P2RY12, a highly expressed microglial signature gene that mediates microglial-neuronal interactions, we show that adult females have a significantly higher expression of the receptor than adult male microglia. We further demonstrate that a genetic deletion of P2RY12 induces sex-specific cellular perturbations with microglia and neurons in females more significantly affected. Correspondingly, female mice lacking P2RY12 exhibit unique behavioral anomalies not observed in male counterparts. These findings underscore the critical, sex-specific roles of P2RY12 in microglial-neuronal interactions, offering new insights into basal interactions and potential implications for CNS disease mechanisms. Graphical Abstract

Abstract Microglia, the primary immune cells of the central nervous system (CNS), are derived from the yolk sac and populate the brain during development. Once microglia migrate to the CNS, they are self-renewing and require CSF1R signaling for their maintenance. Pexidartinib (PLX3397, PLX), a small molecule inhibitor of the CSF1R, has been shown to effectively deplete microglia since microglial maintenance is CSF1R-dependent. There have, however, been several conflicting reports that have shown the potential off-target effects of PLX on peripheral immune cells particularly those of lymphoid origin. Given this controversy in the use of the PLX family of drugs, it has become important to ascertain to what extent PLX affects the peripheral immune profile in lymphoid (spleen, and bone marrow) and non-lymphoid (kidney, lungs, and heart) organs. PLX3397 chow treatment at 660 mg/kg for 7 days significantly reduced CD45+ macrophages, CX3CR1-GFP cells, CD11b+CD45intermediate cells, and P2RY12 expression in the brain. However, there were minimal effects on peripheral immune cells from both lymphoid and non-lymphoid organs except in the heart where there was a significant decrease in CD3+ cells, inflammatory and patrolling monocytes, and CD11b+Ly6G+ neutrophils. We then stimulated the immune system with 1 mg/kg of LPS which resulted in a significant reduction in the number of innate immune cells. In this context, PLX did not alter the cytokine profile in the serum and the brain of naïve mice but did so in the LPS-stimulated group resulting in a significant reduction in TNFα, IL-1α, IFN-γ and IL-1β. Furthermore, PLX did not alter locomotor activity in the open field test suggesting that microglia do not contribute to LPS-induced sickness behavior. Our results provide an assessment of immune cell populations with PLX3397 treatment on brain, lymphoid and non-lymphoid organs without and during LPS treatment that can serve as a resource for understanding consequences of such approaches.

Abstract Microglia are highly dynamic immune cells of the central nervous system (CNS). Microglial processes interact with neuronal elements constantly on the order of minutes. The functional significance of this acute microglia-neuron interaction and its potential role in the context of pain is still largely unknown. Here, we found that spinal microglia increased their process motility and electrophysiological reactivity within an hour after the insult in a mouse model of formalin-induced acute, sustained, inflammatory pain. Using an ablation strategy to specifically deplete resident microglia in the CNS, we demonstrate that microglia participate in formalin-induced acute sustained pain behaviors by amplifying neuronal activity in the spinal dorsal horn. Moreover, we identified that the P2Y12 receptor, which is specifically expressed in microglia in the CNS, was required for microglial function in formalin-induced pain. Taken together, our study provides a novel insight into the contribution of microglia and the P2Y12 receptor in inflammatory pain that could be used for potential therapeutic strategies.

Disruption of the blood brain barrier can occur in several diseases. Here the authors show that targeted ablation of astrocytes results in a plasticity mechanism in nearby cells to maintain cerebrovascular coverage, but that this mechanism is impaired in older animals.

Microglia are involved in debris clearance and synaptic pruning, among other processes. However, their direct interaction with the brain vasculature is less clear. Here, the authors show that capillary-associated microglia (CAMs) regulate vascular tone via PANX1-P2RY12 signalling.

Summary: Although an imbalance between neuronal excitation and inhibition underlies seizures, clinical approaches that target these mechanisms are insufficient in containing seizures in patients with epilepsy, raising the need for alternative approaches. Brain-resident microglia contribute to the development and stability of neuronal structure and functional networks that are perturbed during seizures. However, the extent of microglial contributions in response to seizures in vivo remain to be elucidated. Using two-photon in vivo imaging to visualize microglial dynamics, we show that severe seizures induce formation of microglial process pouches that target but rarely engulf beaded neuronal dendrites. Microglial process pouches are stable for hours, although they often shrink in size. We further find that microglial process pouches are associated with a better structural resolution of beaded dendrites. These findings provide evidence for the structural resolution of injured dendrites by microglia as a form of neuroprotection.

Microglia are unique brain-resident, myeloid cells. They have received growing interest for their implication in an increasing number of neurodevelopmental, acute injury, and neurodegenerative disorders of the central nervous system (CNS). Fate-mapping studies establish microglial ontogeny from the periphery during development, while recent transcriptomic studies highlight microglial identity as distinct from other CNS cells and peripheral myeloid cells. This evidence for a unique microglial ontogeny and identity raises questions regarding their identity and functions across species. This review will examine the available evidence for microglia in invertebrate and vertebrate species to clarify similarities and differences in microglial identity, ontogeny, and physiology across species. This discussion highlights conserved and divergent microglial properties through evolution. Finally, we suggest several interesting research directions from an evolutionary perspective to adequately understand the significance of microglia emergence. A proper appreciation of microglia from this perspective could inform the development of specific therapies geared at targeting microglia in various pathologies.

Abstract Microglia have been shown to be of critical importance to the progression of temporal lobe epilepsy. However, the broad transcriptional changes that these cells undergo following seizure induction is not well understood. As such, we utilized RNAseq analysis upon microglia isolated from the hippocampus to determine expression pattern alterations following kainic acid induced seizure. We determined that microglia undergo dramatic changes to their expression patterns, particularly with regard to mitochondrial activity and metabolism. We also observed that microglia initiate immunological activity, specifically increasing interferon beta responsiveness. Our results provide novel insights into microglia transcriptional regulation following acute seizures and suggest potential therapeutic targets specifically in microglia for the treatment of seizures and epilepsy.

Summary: Microglia are an exquisitely tiled and self-contained population in the CNS that do not receive contributions from circulating monocytes in the periphery. While microglia are long-lived cells, the extent to which their cell bodies are fixed and the molecular mechanisms by which the microglial landscape is regulated have not been determined. Using chronic in vivo two-photon imaging to follow the microglial population in young adult mice, we document a daily rearrangement of the microglial landscape. Furthermore, we show that the microglial landscape can be modulated by severe seizures, acute injury, and sensory deprivation. Finally, we demonstrate a critical role for microglial P2Y12Rs in regulating the microglial landscape through cellular translocation independent of proliferation. These findings suggest that microglial patrol the CNS through both process motility and soma translocation. : Using a chronic in vivo imaging approach, Eyo et al. show that the physical positions of brain microglia change daily and that these changes increase following certain experimental manipulations. The mechanism underlying these changes involves cell translocation controlled by microglial-specific P2Y12 receptors. Keywords: microglia, P2Y12, seizures, epilepsy, whisker trimming, microglial landscape, two photon chronic imaging

Summary: Spinal long-term potentiation (LTP) at C-fiber synapses is hypothesized to underlie chronic pain. However, a causal link between spinal LTP and chronic pain is still lacking. Here, we report that high-frequency stimulation (HFS; 100 Hz, 10 V) of the mouse sciatic nerve reliably induces spinal LTP without causing nerve injury. LTP-inducible stimulation triggers chronic pain lasting for more than 35 days and increases the number of calcitonin gene-related peptide (CGRP) terminals in the spinal dorsal horn. The behavioral and morphological changes can be prevented by blocking NMDA receptors, ablating spinal microglia, or conditionally deleting microglial brain-derived neurotrophic factor (BDNF). HFS-induced spinal LTP, microglial activation, and upregulation of BDNF are inhibited by antibodies against colony-stimulating factor 1 (CSF-1). Together, our results show that microglial CSF1 and BDNF signaling are indispensable for spinal LTP and chronic pain. The microglia-dependent transition of synaptic potentiation to structural alterations in pain pathways may underlie pain chronicity. : Zhou et al. characterize chronic pain behaviors triggered by LTP-inducible HFS without nerve injury. They identify that HFS-induced LTP is accompanied by an increase in CGRP terminals in the spinal dorsal horn. Activation of neuronal CSF1-microglial BDNF signaling is indispensable for the synaptic and structural plasticity underlying HFS-induced chronic pain. Keywords: long-term potentiation, chronic pain, calcitonin gene-related peptide, microglia, high-frequency stimulation, colony-stimulating factor 1, brain-derived neurotrophic factor

Peripheral nerve injury causes neuropathic pain accompanied by remarkable microgliosis in the spinal cord dorsal horn. However, it is still debated whether infiltrated monocytes contribute to injury-induced expansion of the microglial population. Here, we found that spinal microgliosis predominantly results from local proliferation of resident microglia but not from infiltrating monocytes after spinal nerve transection (SNT) by using two genetic mouse models (CCR2RFP/+:CX3CR1GFP/+ and CX3CR1creER/+:R26tdTomato/+ mice) as well as specific staining of microglia and macrophages. Pharmacological inhibition of SNT-induced microglial proliferation correlated with attenuated neuropathic pain hypersensitivities. Microglial proliferation is partially controlled by purinergic and fractalkine signaling, as CX3CR1−/− and P2Y12−/− mice show reduced spinal microglial proliferation and neuropathic pain. These results suggest that local microglial proliferation is the sole source of spinal microgliosis, which represents a potential therapeutic target for neuropathic pain management.

Unlike other resident neural cells that are of neuroectodermal origin, microglia are resident neural cells of mesodermal origin. Traditionally recognized for their immune functions during disease, new roles are being attributed to these cells in the development and maintenance of the central nervous system (CNS) including specific communication with neurons. In this review, we highlight some of the recent findings on the bidirectional interaction between neurons and microglia. We discuss these interactions along two lines. First, we review data that suggest that microglial activity is modulated by neuronal signals, focusing on evidence that (i) neurons are capable of regulating microglial activation state and influence basal microglial activities; (ii) classic neurotransmitters affect microglial behavior; (iii) chemotactic signals attract microglia during acute neuronal injury. Next, we discuss some of the recent data on how microglia signal to neurons. Signaling mechanisms include (i) direct physical contact of microglial processes with neuronal elements; (ii) microglial regulation of neuronal synapse and circuit by fractalkine, complement, and DAP12 signaling. In addition, we discuss the use of microglial depletion strategies in studying the role of microglia in neuronal development and synaptic physiology. Deciphering the mechanisms of bidirectional microglial-neuronal communication provides novel insights in understanding microglial function in both the healthy and diseased brain.

Microglia and monocytes contribute to neuropathic pain states, but the precise role of the two cell types is not clear. Here Peng et al.use temporally controlled ablation of monocytes and microglia in mice to show that these cells work together to initiate neuropathic-pain like behaviour, but are less important in the maintenance phase.