Oxysterol-binding protein-related protein 6 regulates neuronal morphology and migration of cerebellar granule cells during cerebellar development in vivo.

Polygenic and spatial insights into the genetic uniqueness of essential tremor using common variants.

GD3 synthase deficiency disrupts Na+/K+-ATPase and plasma membrane Ca2+-ATPase function in mouse brain.

N6-methyladenosine (m6A) is the most abundant transcriptional modification in eukaryotic RNA, regulating RNA fate. While the functions of m6A in the development of the mammalian brain have been extensively studied, its role in synaptic plasticity, and brain function remain underexplored. The involvement of this modification in Parkinson’s disease and other synucleinopathies has only recently been studied, and needs further investigation. Here, we investigated the m6A epitranscriptome using MeRIP-seq in A30P-aSyn transgenic mice (aSyn Tg). We observed hypermethylation of synaptic genes in young aSyn Tg mice compared to age-matched control mice. The methylation was reduced during ageing. Using immunofluorescence imaging and biochemical analysis, we further investigated the levels and distribution of m6A regulatory enzymes—writer, METTL3, reader, YTHDF1, and eraser, FTO, in the cortex, striatum, hippocampus, and cerebellum of aSyn Tg and control mice, and in primary cortical neuronal cultures. While the levels of these proteins were similar, METTL3 was found in the nucleus and in the post-synaptic compartment in neurons, suggesting it may play a role in methylation at the post-synapse. Our findings suggest that alterations in the regulation of m6A RNA methylation may be associated with neurodegeneration and ageing and it may play a significant role at the synapse.

Resveratrol induces molecular changes in cholesterol homeostasis in SAMP8 mice cerebellum.

Smoking is a major public health concern with widespread effects on multiple organ systems, including the immune system. Chronic nicotine exposure can alter immune cell function through nicotinic receptors expressed on peripheral macrophages and microglia in the brain. Recent evidence indicates that the cerebellum is impacted by nicotine, contributing to motor and nonmotor outcomes, during drug use. In this study, we investigated the effect of chronic nicotine on microglia proteomes in the adult mouse cerebellum. Microglia were isolated by fluorescence-activated cell sorting (FACS) based on CD11bhigh CD45low/intermediate expression from male and female mice (n = 9 per group) exposed to 200 μg/mL nicotine (dissolved in 2% saccharin drinking water) for 30 days. Proteomic analysis was performed using liquid chromatography electrospray ionization (LC-ESI) mass spectrometry (MS) comparing the effect of nicotine relative to vehicle control. Our results reveal a sex-dependent effect of nicotine on microglial proteomes. While both males and females exhibited histone-related genomic responsiveness to nicotine, males demonstrated enrichment in cytoskeletal and metabolic proteins, and females showed complement-protein adaptations. The microglial proteome in male mice displayed nicotine-related adaptations in proteins that can contribute to neurodisorders including Huntington's disease and amyotrophic lateral sclerosis (ALS), of which smoking is a known risk factor. Together, these results highlight an effect of nicotine on the proteome of microglia providing insight into immune pathways that can contribute to smoking-related behavior and disease.

The purpose of this study is to non-invasively image subtle blood-brain barrier (BBB) permeability changes in a state of chronic neuroinflammation using radiolabelled human serum albumin (HSA). Although biomarker-based methods for detecting BBB dysfunctions have been established, sensitive spatial and quantitative imaging of low-grade, chronic BBB permeability changes remains limited. This study seeks to establish radiolabelled albumin as a sensitive radiotracer for visualizing and quantifying BBB dysfunction during chronic neuroinflammation. This study evaluated the utility of radiolabeled serum albumin ([89Zr]Zr-DFO-HSA) for assessing BBB permeability using co-registered PET and MR imaging in a transgenic mouse model of neuroinflammation mediated by IKK/NF-κB activation. PET and MRI revealed significantly increased radiotracer accumulation in the cerebellum of neuroinflammatory mice, corroborated by ex vivo analyses. Quantitative PET data revealed elevated intracerebral radiotracer concentrations over 24h, even in the absence of morphological changes in histological evaluations. These findings emphasize the sensitivity of [89Zr]Zr-DFO-HSA in detecting early-stage BBB permeability changes prior to the onset of overt morphological damage or clinical symptoms. As a surrogate marker for macromolecular leakage, this non invasive imaging approach shows promise for advancing preclinical research into neuroinflammatory disorders and barrier dysfunctions.

Benchtop sucrose-gradient pelleting enables rapid nuclei purification from myelin-rich adult mouse brainScales across tissue inputs (e.g., hippocampus ∼15-20 mg; cerebellum ∼50-70 mg) without ultracentrifugation or 15 mL gradientsMagnetic enrichment as the recommended final cleanup step further reduces myelin/debris carryover and is compatible with 10x Flex and PARSE WT workflows.

Organ development and function are orchestrated by intricate transcriptional circuits. Here, we present a comprehensive atlas profiling 1,904 transcription regulators in the brain, cerebellum, heart, kidney, liver, ovary, and testis of fetal, neonatal, and adult mice. Using this dataset, we uncover Thymocyte Selection-Associated High Mobility Group Box Family Member 3 (TOX3) as a potential coactivator of Atoh1 in cerebellar granule neuron progenitors (GNPs). Tox3-deficient mice display severe ataxia and cerebellar hypoplasia, driven by depletion of GNPs, diminished Atoh1 expression, and impaired primary cilia. Single-nucleus RNA-sequencing analyses reveals compromised maintenance of the progenitor pool. TOX3 is also highly expressed in subsets of medulloblastoma, and its deletion reduces cerebellar neoplasia and prolongs survival in a mouse model. Mechanistically, how lineage-defining factors such as Atoh1 drive robust gene expression despite weak intrinsic transactivation activity remains unclear. We show that Tox3 physically associates with Atoh1 and co-occupies shared regulatory elements, converting an otherwise weak single-copy Atoh1-responsive E-box into a highly active enhancer that drives transcriptional activation by up to 120-fold, including at an ultraconserved E-box downstream of Atoh1 itself. Cross-species single-cell comparisons further show an association between Tox3 expression and cerebellum expansion during vertebrate evolution. Together, this work supports Tox3 as a critical Atoh1 coactivator in cerebellar development, tumorigenesis, and evolution, while providing an atlas and screening strategy as a valuable resource for exploring novel transcriptional regulators in organogenesis and tissue physiology.

Multiple sclerosis (MS) is an autoimmune demyelinating disease of the central nervous system (CNS) characterized by multifocal inflammation and axonal degeneration, driven by innate and adaptive immune cells. The Janus Kinase (JAK)/Signal Transducers and Activators of Transcription (STAT)/Suppressors Of Cytokine Signaling (SOCS) pathway regulates immune cell activity, with SOCS proteins functioning as negative regulators. Using the Experimental Autoimmune Encephalomyelitis (EAE) model of MS, our prior work demonstrated that mice lacking Socs3 in myeloid cells (Socs3ΔLysM) developed severe, brain-targeted EAE (btEAE), with increased cerebellar infiltration of activated neutrophils. To define neutrophil-specific roles, we generated mice with Socs3 deletion restricted to neutrophils (Socs3ΔLy6G). Following MOG-induced EAE, these mice exhibited clinical features identical to Socs3ΔLysM mice, including severe cerebellar demyelination, increased cerebellar infiltration of activated neutrophils and CD4+ T-cells, and clinical symptoms of both btEAE and classical EAE (cEAE), the latter involving the spinal cord (SC). Cerebellar neutrophils from Socs3ΔLy6G mice exhibited a primed, inflammatory phenotype with elevated reactive oxygen species, neutrophil extracellular traps (NETs) and heightened production of pro-inflammatory cytokines/chemokines. Neutrophil depletion alleviated btEAE, confirming their pathogenic role. Single-cell RNA Sequencing (scRNA-Seq) of cerebellum (CB) and SC neutrophils revealed five clusters in naïve and EAE mice, with expansion of two clusters (Neu2 and Neu4) in Socs3ΔLy6G mice with EAE. Neu2, Neu3 and Neu4 clusters showed high expression of Saa3, Il1b and Cxcl2, with Neu4 enriched in cytokine signaling pathways and inflammatory responses. Strikingly, Saa3 mRNA and protein expression were markedly increased in the CB and SC of Socs3ΔLy6G mice with EAE compared to controls. Translationally, the human orthologue SAA1 was significantly elevated in plasma from MS patients relative to healthy controls. Collectively, these findings demonstrate that Socs3 deficiency unleashes pathogenic neutrophil activity in Socs3ΔLy6G mice with EAE. They further demonstrate neutrophil heterogeneity within the inflamed CNS and define inflammatory transcriptional states, with Saa3/SAA1 as a potential biomarker and/or target in autoimmune neuroinflammation.

Background: Fibromyalgia is a chronic disease that predominantly affects women and lasts over several months, causing problems both for individuals and society. While several studies have demonstrated the potential of electroacupuncture (EA) to alleviate fibromyalgia pain in mice, further research is needed to investigate its underlying mechanisms. Programmed cell death ligand 1 (PD-L1)/PD-1 were first identified to be involved in cancer immunotherapy, and their application to pain management has not been yet investigated. Methods: In this study, we aimed to explore the mechanism underlying the action of PD-L1 on the PD-1 pathway in a mouse model of fibromyalgia. Results: We established such a mouse model using intermittent cold stress (ICS) and confirmed mechanical (D4: 2.02 ± 0.13 g, n = 9) and thermal (D4: 4.28 ± 0.21 s, n = 9) hyperalgesia. We found that EA, intracerebral ventricle (ICV) PD-L1 injection, and transient receptor potential vanilloid 1 (Trpv1) knockout effectively counteracted hyperalgesia. We observed low PD-1 expression in the cerebellum of fibromyalgia mice but increased expression of TRPV1 and pain-related kinases. These phenomena could be further reversed by EA, ICV PD-L1 injection, and Trpv1 knockout. To confirm that these effects were caused by PD-L1 release, we added PD-L1-neutralizing antibodies to the EA and PD-L1 treatment. The analgesic effects and EA and PD-L1 mechanisms were inhibited. Conclusions: Our results elucidate the role of the PD-L1/PD-1 pathway in EA treatment of fibromyalgia and reveal its potential value for fibromyalgia management.

Rett syndrome (RTT) is a neurodevelopmental disorder characterized by motor deficits, partly attributed to cerebellar dysfunction. RTT is primarily caused by mutations in the gene encoding the methyl-CpG-binding protein 2 (MECP2), which has been implicated in cholesterol homeostasis by mechanisms that remain poorly understood. Given that brain cholesterol is primarily synthesized de novo and that disrupted cholesterol homeostasis is linked to various neurological disorders, we aimed to investigate cholesterol regulation in the cerebellum of Mecp2-null mice, a well-established RTT model. We measured total cholesterol levels in cerebellar tissue and cerebellar synaptosomes and assessed the expression of genes involved in cholesterol biosynthesis and intracellular transport. Our results show significantly elevated total cholesterol in both cerebellar tissue and synaptosomes. Furthermore, we identified a marked reduction in CYP46A1 expression, which is essential for the elimination of encephalon sterols. In contrast, key cholesterol biosynthetic regulators (Srebp2, Hmgcs1, Sqle) showed no significant changes in expression, suggesting an impaired cerebellar cholesterol turnover-driven by defective clearance-rather than enhanced synthesis may underlie the metabolic imbalance observed in the cerebellum of the RTT mouse model. Altogether, these findings provide a mechanistic insight into how MeCP2 deficiency disrupts cerebellar cholesterol homeostasis and highlight cholesterol clearance pathways as potential contributors to RTT pathology and a factor to consider for further RTT therapeutic approaches.

Primary coenzyme Q10 (CoQ10) deficiency is a rare mitochondrial disorder caused by mutations in genes involved in CoQ biosynthesis (e.g., COQ4) that result in impaired mitochondrial respiration, oxidative stress, and dysfunction across multiple organ systems because of decreased mitochondrial levels of CoQ10. Although oral CoQ10 supplementation has been examined for standard of care, poor absorption and inadequate tissue and intracellular distribution have resulted in a lack of clinically significant efficacy. BPM31510 is a lipid nanoparticle containing oxidized CoQ10 designed to improve bioavailability and targeted uptake into the mitochondria. In the current study, we assessed the efficacy of BPM31510 to increase CoQ levels in Coq4F147C mice, a novel genetic knock-in model of primary CoQ deficiency. Coenzyme Q9, the main form of CoQ in mice, and CoQ10 were significantly decreased in the brain, kidney, heart, and muscle of Coq4F147C mice compared with Coq4+/+ mice. BPM31510 treatment significantly increased oxidized CoQ10 levels across all tissues, mediated by the nanoliposome biodistribution of oxidized CoQ10 in BPM31510. MALDI-MS imaging demonstrated regional and spatial restoration of CoQ10 within the brain, including the cerebellum, myocardium, and renal cortex of Coq4F147C mice. These results demonstrate that BPM31510 successfully concentrates pharmacologically active CoQ10 in target tissues that are not reachable with oral therapy in a genetic model of primary CoQ deficiency. We enabled the visualization of suborgan CoQ10 localization to specifically demonstrate CoQ10 restoration. This study establishes proof of concept for spatial quinomics, a new methodology that combines spatial metabolomics with quinomics to evaluate next-generation CoQ10-based therapeutics for mitochondrial disorders.

Maternal exposure to polypropylene nanoplastics disrupts sex- and region-specific lipid metabolism in the brains of C57BL/6N mouse offspring.

Interleukin-17 (IL-17) is a pleiotropic cytokine produced mainly by peripheral T helper 17 cells. Yet, the brain functions of IL-17 derived from central nervous cells remain poorly understood. Here, we find an aberrant IL-17A signaling in the cerebellum of Fmr1-KO mice, a well-established genetic model for autism spectrum disorder (ASD). Cerebellar IL-17A, derived exclusively from microglia, is essential for the regulation of social behaviors by maintaining neuronal excitability and selectively suppressing inhibitory neurotransmission of Purkinje cells (PCs) in the cerebellar Crus I, a brain region critically involved in social cognition. Specific downregulation of IL-17 receptor-mediated signaling in cerebellar PCs recapitulates ASD-like social deficits and repetitive behaviors. Notably, both direct administration of IL-17A and induction of IL-17A release from cerebellar microglia by poly(I:C) effectively restore PC excitability and ameliorate ASD-like symptoms. The findings uncover an indispensable role of microglia-derived IL-17A for cerebellar social processing and suggest potential therapeutic strategies targeting IL-17A signaling for ASD.

The poor clinical outcomes of pediatric high-grade glioma (pHGG) highlight the urgent need for new therapies. Oligodendrocyte lineage transcription factor 2 (OLIG2) is a pro-mitotic transcription factor highly expressed in glioma stem cells and may represent a novel therapeutic target. To evaluate the therapeutic efficacy of an OLIG2 inhibitor CT-179 in pHGG, we determined the OLIG2 mRNA expression in 10 patient-derived orthotopic xenograft (PDOX) models. In vitro activities of CT-179 were analyzed in monolayer and neurosphere cells (0-10 µM) with and without radiation (XRT) (0-8 Gy), brain penetration was evaluated in tumor-bearing PDOX mice, and in vivo efficacy was determined at 15-240 mg/kg (oral) alone or combined with XRT (2 Gy/day × 5 days). Changes in animal survival times were analyzed using the Kaplan-Meier method, followed by pair-wise comparisons. Increased OLIG2 mRNA expression was detected in seven out of ten PDOX models. CT-179 inhibited cell viability in a time- and dose-dependent manner in all eight pGBM xenograft tumors (IC50 0.03-10 µM) and was potentiated by XRT (0.03-1 µM). Oral gavage (24 mg/kg) of CT-179 for 5 days led to effective penetration in mouse cerebrum (3232.7 ± 569.2 ng/g), cerebellum (1563.3 ± 269.6 ng/g), brain stem (1685.3 ± 309 ng/g), and PDOX tumors (1814 ± 110.3 ng/g) vs. 361.3 ± 1.5 ng/mL in serum. CT-179 alone was not active at 200 mg/kg in four models, although it was moderately effective at 240 mg/kg in one model. When combined with XRT, a significant extension of animal survival times was observed in two out of four models. Doses needed to eliminate OLIG2 expression in vitro varied from 0.3 to >1 µM in pGBM cells. In summary, our data showed that orally administered CT-179 penetrated the blood-brain barrier (BBB) and exhibited potential for inhibiting pGBM growth when combined with XRT.

• Tobacco smoke exposure consistently increased cAMP response element-binding protein (CREB) phosphorylation in mice cerebellum under different exposure regimens. • AB-free kava effectively suppressed tobacco smoke-induced p-CREB elevation in mice cerebellum. • The impact of AB-free kava suppression of tobacco smoke-induced p-CREB elevation remains to be determined. Our recent studies revealed the potential of AB-free kava as a novel therapeutic candidate against tobacco use disorder while the underlying mechanism remains to be elucidated. The cerebellum region in the brain has been reported to be involved in tobacco withdrawal. The cAMP response element-binding protein (CREB) activation via phosphorylation has also been reported to contribute to substance abuse, including tobacco use disorder. Our early pre-clinical work revealed the potential of AB-free kava to suppress tobacco smoke-induced CREB activation in lung tissues. In this study, we investigated the impact of tobacco smoke exposure on the levels of phosphorylated CREB (p-CREB) in the mouse cerebellum and evaluated the effects of AB-free kava. Four different regimens of tobacco smoke exposure consistently increased the levels of p-CREB in the cerebellum. AB-free kava effectively suppressed tobacco smoke-induced increase in p-CREB to levels comparable to mice without tobacco smoke exposure. Our data provide preliminary evidence that CREB activation in the cerebellum is a potential mechanism involved in tobacco use disorder and the protective potential of AB-free kava.

Functional neural circuits are sculpted by strengthening frequently used synapses and removing unnecessary connections. At birth, cerebellar Purkinje cells receive inputs with similar synaptic strengths from multiple climbing fibers (CFs). During postnatal development, a single "winner" CF is selectively strengthened and expands its dendritic innervation territory, while somatic "loser" synapses are eliminated. Here, we report that deleting metabotropic glutamate receptor 1 (mGluR1) or protein kinase Cγ (PKCγ) in mice disrupts this selective strengthening and territory expansion of "winner" CFs during early development. This impairment leads to weaker synaptic transmission and diminished dendritic innervation territory of "winner" CFs at later stages. Notably, "winner" CF synapses in these mutants exhibit impaired long-term potentiation, reduced AMPA receptor expression, and simpler postsynaptic organizations. These findings reveal a previously unappreciated role for mGluR1-PKCγ signaling, besides its established role in eliminating "loser" CFs, in promoting the functional and structural maturation of "winner" CF synapses.

Immunohistochemically-detected differential localization of CXCL14 in mouse cerebellum suggesting the presence of neuronal and glial forms of CXCL14.

This study aimed to investigate the mechanisms by which Acanthopanax senticosus extract(ASH) exerts effects on α-synuclein(α-syn) overexpressing transgenic mouse model of Parkinson's disease(PD), with a focus on its regulation of brain lipid metabolism. Twenty PD model mice were randomly assigned to a model group or an ASH treatment group(45.5 mg·kg~(-1) by gavage for 4 weeks), and 10 C57BL/6 mice served as a normal control group. Behavioral assessments revealed that, compared with controls, PD model mice showed prolonged pole test time, reduced spontaneous locomotor activity, shorter latency to fall in the rotarod test, and decreased total distance traveled in the open field test. Serum levels of tumor necrosis factor-α(TNF-α), interleukin-6(IL-6), caspase-9 were significantly elevated, B-cell lymphoma-2(Bcl-2), and proliferating cell nuclear antigen(PCNA) expression was reduced, and marked neuronal damage was observed in brain tissue. ASH intervention significantly improved these behavioral and biochemical parameters and attenuated neuronal injury. Untargeted lipidomics analysis revealed significant alterations in sphingomyelin(SM), ceramide(Cer), phosphatidylcholine(PC), and phosphatidylserine(PS) across multiple brain regions(cortex, substantia nigra, cerebellum, and striatum) in PD mice, which were notably restored by ASH treatment. Pathway analysis indicated that these metabolites were predominantly involved in sphingolipid metabolism. Western blot further demonstrated that ASH downregulated the expression of key sphingolipid metabolic enzymes serine palmitoyltransferase long-chain base subunits 1 and 2(SPTLC1 and SPTLC2) and upregulated UDP-glucose ceramide glucosyltransferase(UGCG), β-galactosylceramidase(GALC), and sphingosine kinase 2(SPHK2), thereby suppressing abnormal SM and Cer accumulation in the substantia nigra and elevating PS and PC levels in the striatum. Spearman's correlation analysis supported the modulatory effect of ASH on brain lipid metabolic profiles. In conclusion, ASH improves behavioral deficits, exerts anti-inflammatory effects, and regulates sphingolipid metabolism to correct disordered lipid profiles, thereby providing neuroprotective effects in PD mice.