Glia最新文献

In the mammalian central nervous system, axonal myelination, executed by mature oligodendrocytes (MOLs), enables rapid neural transmission. Conversely, myelin deficiencies are hallmark features of multiple sclerosis, optic neuromyelitis, and some leukodystrophies. Recent studies have highlighted that MOLs are heterogeneous; however, how MOL subpopulations are specified and balanced in physiological settings is poorly understood. Previous works have demonstrated an essential role of the small GTPases R-Ras1 and R-Ras2 in the survival and myelination of oligodendrocytes. In this study, we aimed to determine how R-Ras1 and R-Ras2 contribute to the heterogeneity of MOL subpopulations. Our results evidence that R-Ras1 and R-Ras2 affect specification into the distinct subpopulations MOL1, MOL2, and MOL5/6, which in turn vary in their dependence of these GTPases. In R-Ras1 and/or R-Ras2 mutant mice, we observed an increase in the MOL1 subpopulation and a decrease in the MOL2 and MOL5/6 subpopulations. We identified R-Ras1 and R-Ras2 as key elements in balancing the heterogeneity of MOLs. Our results contribute to the understanding of the molecular mechanisms underlying the heterogeneity of MOLs and the myelination processes, which is crucial for innovating regenerative therapies for nervous system disorders.

Single-cell transcriptomics, epigenomics, and other 'omics applied at single-cell resolution can significantly advance hypotheses and understanding of glial biology. Omics technologies are revealing a large and growing number of new glial cell subtypes, defined by their gene expression profile. These subtypes have significant implications for understanding glial cell function, cell-cell communications, and glia-specific changes between homeostasis and conditions such as neurological disease. For many, the training in how to analyze, interpret, and understand these large datasets has been through reading and understanding literature from other fields like biostatistics. Here, we provide a primer for glial biologists on experimental design and analysis of single-cell RNA-seq datasets. Our goal is to further the understanding of why decisions are made about datasets and to enhance biologists' ability to interpret and critique their work and the work of others. We review the steps involved in single-cell analysis with a focus on decision points and particular notes for glia. The goal of this primer is to ensure that single-cell 'omics experiments continue to advance glial biology in a rigorous and replicable way.

Mature hippocampal astrocytes exhibit a linear current-to-voltage (I-V) K⁺ membrane conductance called passive conductance. It is estimated to enable astrocytes to keep potassium homeostasis in the brain. We previously reported that the TWIK-1/TREK-1 heterodimeric channels are crucial for astrocytic passive conductance. However, the regulatory mechanism of these channels by other binding proteins remains elusive. Here, we identified Na+/H+ exchange regulator-1 (NHERF-1), a protein highly expressed in astrocytes, as a novel interaction partner for these channels. NHERF-1 endogenously bound to TWIK-1/TREK-1 in hippocampal cultured astrocytes. When NHERF-1 is overexpressed or silenced, surface expression and activity of TWIK-1/TREK-1 heterodimeric channels are inhibited or enhanced, respectively. Furthermore, we confirmed that reduced astrocytic passive conductance by NHERF-1 overexpressing in the hippocampus increases kainic acid (KA)-induced seizure sensitivity. Taken together, these results suggest that NHERF-1 is a key regulator of TWIK-1/TREK-1 heterodimeric channels in astrocytes and suppression of TREK-1 surface expression by NHERF-1 increases KA-induced seizure susceptibility via reduction of astrocytic passive conductance.

Efficient clearance of hematomas is crucial for improving clinical outcomes in patients with intracerebral hemorrhage (ICH). The glymphatic system, facilitated by aquaporin-4 (AQP4), plays a crucial role in cerebrospinal fluid (CSF) entry and metabolic waste clearance. This study examined the role of the glymphatic system in ICH pathology, with a focus on AQP4. Collagenase-induced ICH models were established, with AQP4 expression regulated through mifepristone as an agonist, TGN-020 as an inhibitor, and Aqp4 gene knockout. Fluorescence tracing and multimodal magnetic resonance imaging (MRI) were employed to observe glymphatic system functionality, hematoma, and edema volumes. Neurological deficit scoring was performed using the modified Garcia Scale. AQP4 expression was quantified using RT-qPCR and Western blotting, and cellular localization was explored using immunofluorescence. The brain tissue sections were examined for neuronal morphology, degenerative changes, and iron deposition. Three days post-ICH, the AQP4 agonist group showed increased AQP4 protein expression and perivascular polarization, decreased hemoglobin levels, and reduced iron deposition. Conversely, the inhibition group exhibited contrasting trends. AQP4 activation improved glymphatic system function, leading to a wider distribution, improved neurological function, and reduced hematoma. Pharmacological inhibition and genetic knockout of AQP4 have opposing effects. The glymphatic system, facilitated by AQP4, plays a crucial role in hematoma clearance following cerebral hemorrhage. Upregulation of AQP4 improves glymphatic system function, facilitates hematoma clearance, and promotes brain tissue recovery.

Ubiquitination is a major post-translational regulatory mechanism that tunes numerous aspects of ubiquitinated target proteins, including localization, stability, and function. During differentiation and myelination, Oligodendrocytes (OLs) in the central nervous system and Schwann cells (SCs) in the peripheral nervous system undergo major cellular changes, including the tightly controlled production of large and accurate amounts of proteins and lipids. Such processes have been implied to depend on regulatory aspects of ubiquitination, with E3 ubiquitin ligases being generally involved in the selection of specific ubiquitination substrates by ligating ubiquitin to targets and granting target selectivity. In this study, we have used multiple transgenic mouse lines to investigate the functional impact of the E3 ubiquitin ligase Nedd4 in the OL- and SC-lineages. Our findings in the developing spinal cord indicate that Nedd4 is required for the correct accumulation of differentiated OLs and ensures proper myelination, supporting and further expanding previously suggested conceptual models. In sciatic nerves, we found that Nedd4 is required for timely radial sorting of axons by SCs as a pre-requirement for correct onset of myelination. Moreover, Nedd4 ensures correct myelin thickness in both SCs and spinal cord OLs.

Oligodendrocytes (OLs) of the central nervous system require iron for proteolipid biosynthesis during the myelination process. Although most heme is found complexed to hemoglobin in red blood cells, surprisingly, we found that Slc48a1, encoding the heme transporter Hrg1, is expressed at higher levels in OLs than any other cell type in rodent and humans. We confirmed in situ that Hrg1 is expressed in OLs but not their precursors (OPCs) and found that Hrg1 proteins in CNS white matter co-localized within myelin sheaths. In older Hrg1 null mutant mice we observed reduced expression of myelin associated glycoprotein (Mag) and ultrastructural myelin defects reminiscent of Mag-null animals, suggesting myelin adhesion deficiency. Further, we confirmed reduced myelin iron levels in Hrg1 null animals in vivo, and show that OLs in vitro can directly import both the fluorescent heme analogue ZnMP and heme itself, which rescued iron deficiency induced inhibition of OL differentiation in a heme-oxidase-dependent manner. Together these findings indicate OL Hrg1 encodes a functional heme transporter required for myelin integrity.

The Enteric Nervous System is composed of a vastly interconnected network of neurons and glial cells that coordinate to regulate homeostatic gut function including intestinal motility, nutrient sensing, and mucosal barrier immunity. Enteric Glial Cells (EGCs) are a heterogeneous cell population located throughout the gastrointestinal tract and have well described roles in regulating intestinal immune responses. Enteric Glial Cells have been suggested to act as nonconventional antigen presenting cells via the Major Histocompatibility Complex II (MHC II), though this has not been confirmed functionally. Here, we investigate the capability of EGCs to present antigen on MHC I and MHC II using in vitro antigen presentation assays performed with primary murine EGC cultures. We found that EGCs are capable of functional antigen presentation on MHC I, including antigen cross-presentation, but are not capable of functional antigen presentation on MHC II. We also determined EGC cell surface MHC I and MHC II expression levels by flow cytometry during intestinal inflammation during Dextran Sodium Sulfate-induced colitis or acute Toxoplasma gondii infection. We found that EGCs upregulate MHC I during acute T. gondii infection and induce low-level MHC II expression. These findings suggest that EGCs may be important in the regulation of CD8⁺ T cell responses via MHC I mediated antigen (cross) presentation but may not be relevant for MHC II-mediated antigen presentation.

While some vivid memories are unyielding and unforgettable, others fade with time. Astrocytes are recognized for their role in modulating the brain's environment and have recently been considered integral to the brain's information processing and memory formation. This suggests their potential roles in emotional perception and memory formation. In this study, we delve into the impact of amygdala astrocytes on fear behaviors and memory, employing astrocyte-specific optogenetic manipulations in mice. Our findings reveal that astrocytic photoactivation with channelrhodopsin-2 (ChR2) provokes aversive behavioral responses, while archaerhodopsin-T (ArchT) photoactivation diminishes fear perception. ChR2 photoactivation amplifies fear perception and fear memory encoding but obstructs its consolidation. On the other hand, ArchT photoactivation inhibits memory formation during intense aversive stimuli, possibly due to weakened fear perception. However, it prevents the decay of remote fear memory over three weeks. Crucially, these memory effects were observed when optogenetic manipulations coincided with the aversive experience, indicating a deterministic role of astrocytic states at the exact moment of fear experiences in shaping long-term memory. This research underscores the significant and multifaceted role of astrocytes in emotional perception, fear memory formation, and modulation, suggesting a sophisticated astrocyte-neuron communication mechanism underlying basic emotional state transitions of information processing in the brain.

Timely differentiation and myelin formation by oligodendrocytes are essential for the physiological functioning of the central nervous system (CNS). While the Rho GTPase RhoA has been hinted as a negative regulator of myelin sheath formation, the precise in vivo mechanisms have remained elusive. Here we show that RhoA controls the timing and progression of myelination by oligodendrocytes through a fine-tuned balance between cortical tension, membrane tension and cell shape. Using a conditional mouse model, we observe that Rhoa ablation results in the acceleration of myelination driven by hastened differentiation and facilitated through membrane expansion induced by changes in MLCII activity and in F-actin redistribution and turnover within the cell. These findings reveal RhoA as a central molecular integrator of alterations in actin cytoskeleton, actomyosin contractility and membrane tension underlying precise morphogenesis of oligodendrocytes and normal myelination of the CNS.

Phagocytosis is an indispensable function of microglia, the brain professional phagocytes. Microglia is particularly efficient phagocytosing cells that undergo programmed cell death (apoptosis) in physiological conditions. However, mounting evidence suggests microglial phagocytosis dysfunction in multiple brain disorders. These observations prompted us to search for phagocytosis modulators (enhancers or inhibitors) with therapeutic potential. We used a bottom-up strategy that consisted on the identification of phagocytosis modulators using phenotypic high throughput screenings (HTSs) in cell culture and validation in organotypic cultures and in vivo. We performed two complementary HTS campagnes: at Achucarro, we used primary cultures of mouse microglia and compounds of the Prestwick Chemical Library; at Roche, we used human iPSC derived macrophage-like cells and a proprietary chemo-genomic library with 2200 compounds with known mechanism-of-action. Next, we validated the more robust compounds using hippocampal organotypic cultures and identified two phagocytosis inhibitors: trifluoperazine, a dopaminergic and adrenergic antagonist used as an antipsychotic and antineoplastic; and deoxytubercidin, a ribose derivative. Finally, we tested whether these compounds were able to modulate phagocytosis of apoptotic newborn cells in the adult hippocampal neurogenic niche in vivo by administering them into the mouse hippocampus using osmotic minipumps. We confirmed that both trifluoperazine and deoxytubercidin have anti-phagocytic activity in vivo, and validated our bottom-up strategy to identify novel phagocytosis modulators. These results show that chemical libraries with annotated mechanism of action are an starting point for the pharmacological modulation of microglia in drug discovery projects aiming at the therapeutic manipulation of phagocytosis in brain diseases.