Our Research
Non-neuronal cells called glia make up 40-70% of cells in the CNS. Yet, they were ignored for almost a century, as research focused on electrically excitable neurons that assemble into circuits via synaptic connections. New findings show that glial cells can regulate neural development, synapse function, and electrical properties of neurons, suggesting that they critically inform learning, memory, and behavior. Glia are also exquisitely sensitive to CNS perturbations and many disease risk genes are predominantly expressed by glia, indicating that these cells are likely the most promising therapeutic targets for a wide array of CNS pathologies. Understanding how glial-neuron interactions shape brain function is one of the biggest knowledge gaps in the field of neuroscience. Our research focuses primarily on microglia, the brain’s immune cells. Microglia are ubiquitous, highly dynamic, and possess a surprising repertoire of mechanisms for shaping neuronal function. An overarching goal of our research program is to yield transformative insight into basic brain function and to promote innovative approaches to treating CNS circuit dysfunction and disease through study of these cells.
Lysosome status as a critical driver of microglial aging.
Changes in microglial function during aging may critically shape which neurons are most vulnerable to declining synaptic function and neurodegenerative disease. Our recent study suggested important links between the status of microglial lysosomes and proliferative and inflammatory responses of these cells during aging (Moca et al. 2022 J Neurosci). To explore this connection further, in a follow-up study we analyzed autofluorescent, protein-lipid aggregates (lipofuscin) that accumulate within microglia during aging. We found that these aggregates start accumulating in microglial lysosomes in middle age and that this accumulation is more severe in the ventral tegmental area (VTA), where microglia exhibit early inflammation during aging.
Knockout of a key microglial chemokine receptor (CX3CR1) worsens lipofuscin accumulation, lysosome swelling, and proliferation in VTA microglia, supporting mechanistic links between these phenomena. RNAseq of microglia with high- and low- lipofuscin burdens revealed that lipofuscin-high microglia have 1000+ differentially expressed genes compared to low-lipofuscin microglia, highlighting how profoundly this degradative burden impacts overall cell state. Strikingly, lipo-high and lipo-low microglia from striatum and midbrain were highly divergent in their patterns of gene expression, showing that microglial response to aging associated degradative burden is brain region specific. We developed a novel protocol to purify microglial lipofuscin and generated the first cell-specific proteomic analysis of this material, revealing a surprising contribution of histones and cytoskeletal proteins to aggregate accumulation. We are collaborating with Dr. Rosa Paolicelli (University of Lausanne) to treat cultured microglia with lipofuscin purified via our protocol to determine if lipofuscin lysosome overload is sufficient to cause microglial proliferation and inflammatory or trophic factor production in vitro. We are collaborating with Dr. Carol Barnes (U Arizona), to analyze microglial lipofuscin accumulation in brain tissue from aged, cognitively characterized macaques.
To test causal links between lysosome status and phenotypic changes observed in microglia during aging, we are using transgenic mice to genetically induce lysosome overload independently of aging (Cln3-/- mice, collaboration with Dr. Andrea Ballabio, Baylor). Using these mice, we found that genetically-driven lysosome challenge is sufficient to cause microglial proliferation and lysosome enlargement that is more severe in VTA microglia compared to nucleus accumbens (NAc) microglia, similar to what we observe during aging. Bulk RNAseq of microglia from Cln3-/- and control mice again highlights prominent regional differences in the microglial response to this lysosomal challenge.
Long-term outlook: Lysosomes are complex organelles containing over 60 lytic enzymes and a similar number of membrane transporters and signaling proteins. Through proteomics of microglial lipofuscin, RNAseq of lipofuscin-burdened microglia from aging mice, and RNAseq of microglia with genetically-driven lysosome overload we have identified candidate lysosome components that most directly contribute to lysosome overload and regulation of synapse-relevant microglial attributes. Our future research in this area promises to identify new therapeutic targets for manipulating microglial aging to preserve synaptic health and cognition.
**We are grateful to support from The Glen Foundation for Medical Research / American Federation on Aging Research and NIH/NIA which has made this work possible.**
Mitochondria as central regulators of microglial phenotype.
Almost nothing is known about the mechanisms that allow microglia to integrate diverse “input” signals from the microenvironment and link them with “output” changes in microglial attributes and function. We believe that mitochondria, as organelles, may possess this capability to integrate multiple input signals and coordinately regulate numerous cell attributes to alter overall microglial phenotype. In addition to producing ATP, mitochondria mediate numerous intracellular signaling functions and, in macrophages (microglial-like cells), mitochondria regulate multiple cellular properties, including phagocytic and inflammatory profiles. We generated transgenic crosses to GFP-tag mitochondria specifically in microglia (flox-stop-mitoEGFP; CX3CR1CreER/+ mice). We mapped the mitochondrial landscape within microglia for the first time using fixed tissue imaging, FACS-based approaches with mitochondrial dyes, multiphoton imaging of mitochondrial motility, and RNAseq of microglia with distinct mitochondrial membrane potentials throughout the lifespan (Espinoza and Schaler et al. BioRxIV 2025). Consistent with the idea that these organelles coordinately regulate cellular attributes, we find that changes in microglial phenotype – across brain regions, after inflammatory challenge, during postnatal development, and during aging – are all accompanied by and even preceded by significant changes in the status of mitochondria. We also generated transgenic crosses to selectively manipulate mitochondrial function within microglia (TFAMfl/fl; CX3CR1CreER/+ mice) and found that altered mitochondrial status increased microglial morphological complexity and altered expression of multiple homeostatic and inflammatory genes in a brain-region specific manner.
Long-term outlook: Mitochondria produce ATP and also play central roles in calcium buffering, lipid metabolism, and production and management of reactive oxygen species. We propose that microglial mitochondria, as organelles, may possess the capability to simultaneously regulate multiple synapse- and disease-relevant microglial attributes. This is a new idea that holds great promise for harnessing these cells to improve CNS health. Future work will manipulate distinct functions of microglial mitochondria to continue expanding our understanding of how these organelles shape microglial attributes. Multiple risk genes for Parkinson’s Disease (PD) impact mitochondrial function. We are also using mouse models of PD to determine if risk stems from altered microglial function.
**We are grateful to support from The Parkinson's Foundation which has made this work possible.**
Microglia-extracellular matrix (ECM) interactions and cognitive aging.
The ECM is a meshwork of proteins and sugars that critically influences synapse stability and synaptic plasticity, which is essential for memory. The ECM is also perturbed in many contexts where synaptic dysfunction is evident, including aging, neurodegenerative disease, and stroke. Yet, very little is known about how the ECM itself is regulated, maintained, and remodeled, including potential roles for microglia in regulating these processes. We mined our own and published gene expression data and found that microglia express numerous cell surface receptors for “sensing” the ECM, as well as enzymes capable of synthesizing and degrading ECM components. We developed proteomic workflows to map for the first time changes in ECM composition in multiple brain regions during aging and found key links between status of ECM proteins and status of synaptic proteins. We also found substantial differences across brain region in both ECM composition and ECM remodeling during aging. Via confocal imaging in fixed tissue, we found that hyaluronan scaffolds and microglial interactions with hyaluronan are altered during aging, particularly in the VTA. Finally, we found that depletion of microglia from the CNS or deletion of key microglial receptors that regulate their responses to aging can alter the status of hyaluronan networks and local synapse abundance.
To understand how microglial-ECM interactions relate to cognitive function, we have developed a novel, reward-based spatial memory task in which late middle aged mice show deficits in reward-based memory after longer delays. Immunostaining in fixed tissue from these mice revealed that VTA microglial density is correlated with multiple aspects of cognitive performance in aging but not young adult mice. (Gray et al. BioRxIV 2024.01.04.574215). We are now using a combination of tissue proteomics and spatial transcriptomics in behaviorally characterized young and old mice to define how ECM composition, synaptic components, and microglial ECM-sensing receptors and degradative enzymes relate to cognitive preservation or cognitive decline.
Long-term outlook: Revealing that glial cells and factors extrinsic to synapses are key determinants of synaptic decline would represent a paradigm shift for the cognitive aging field, which has focused largely on neuron-intrinsic factors shaping synapse function. Our proteomic and spatial transcriptomic map of microglial-ECM interactions and associated cognitive performance during aging promises to reveal specific molecules most closely aligned with synapse integrity. Future research will manipulate these targets with the aim of modulating microglial-ECM interactions to preserve learning and memory.
**We are grateful to support from The McKnight Brain Research Foundation / American Federation on Aging Research has made this work possible.**
Microglial regulation of circuit maturation and the impact of early life stress (ELS).
A key factor contributing to elevated risk for psychiatric sequelae following ELS is perturbed maturation of the brain’s reward circuitry. Yet, we know relatively little about the cellular and molecular mechanisms that support proper reward circuit development and how such mechanisms are impacted by ELS. Microglia support key aspects of circuit maturation and stress causes dramatic changes in microglial attributes. We found that microglia in the nucleus accumbens (NAc), a key reward circuit nucleus, are dramatically overproduced during a discrete postnatal developmental window in mice (Hope et al. 2020 Eur J Neurosci). This NAc microglial overproduction exceeds that observed in other brain regions and is tightly aligned with local synaptogenesis (De Biase lab, unpublished). In addition, in collaboration with Dr. Laura DeNardo (UCLA), we found that depletion of microglia perturbs NAc glutamatergic synapse maturation and adult NAc-dependent behaviors (Gongwer and Etienne et al. BioRxIV 2025.01.15.633068), revealing a key role for these cells in supporting reward circuit development. Continuing this collaborative work with the DeNardo lab, we are using a mouse model of ELS (limited nesting model) to determine whether ELS similarly perturbs microglial support of NAc circuit maturation. We are using RNAseq of NAc microglia and NAc medium spiny neurons, as well as proteomics of NAc tissue, from ELS and control mice to reveal the impact of ELS on microglial functional state and potential microglial-derived synaptogenic cues.
Long-term outlook: These studies will define for the first time the role that microglial-synapse interactions play in NAc circuit maturation and how they are impacted by ELS. Future research will include comprehensive behavioral analysis after manipulating microglial ability to respond to stress and after manipulating putative microglial synaptogenic cues identified via RNAseq and proteomics. The overall aim is to identify strategies for preserving homeostatic microglial-synapse interactions to prevent long term psychiatric sequelae in individuals who experience early life and adolescent stress.
**We are grateful to support from The Brain and Behavior Research Foundation (NARSAD), The Brain Research Foundation, and Seed Grants from the David Geffen School of Medicine at UCLA that have made this work possible.**
