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Background: Spontaneous intracerebral hemorrhage (ICH) is associated with alarmingly high rates of disability and mortality, and current therapeutic options are suboptimal. A critical component of ICH pathology is the initiation of a robust inflammatory response, often termed "cytokine storm," which amplifies the secondary brain injury following the initial hemorrhagic insult. The precise sources and consequences of this cytokine-driven inflammation are not fully elucidated, necessitating further investigation., Methods: To address this knowledge gap, our study conducted a comprehensive cytokine profiling using Luminex® assays, assessing 23 key cytokines. We then employed single-cell RNA sequencing and spatial transcriptomics at three critical time points post-ICH: the hyperacute, acute, and subacute phases. Integrating these multimodal analyses allowed us to identify the cellular origins of cytokines and elucidate their mechanisms of action., Results: Luminex® cytokine assays revealed a significant upregulation of IL-6 and IL-1β levels at the 24-h post-ICH time point. Through the integration of scRNA-seq and spatial transcriptomics in the hemorrhagic hemisphere of rats, we observed a pronounced activation of cytokine-related signaling pathways within the choroid plexus. Initially, immune cell presence was sparse, but it surged 24 h post-ICH, particularly in the choroid plexus, indicating a substantial shift in the immune microenvironment. We traced the source of IL-1β and IL-6 to endothelial cells, establishing a link to pyroptosis. Endothelial pyroptosis post-ICH induced the production of IL-1β and IL-6, which activated microglial polarization characterized by elevated expression of Msr1, Lcn2, and Spp1 via the NF-κB pathway in the choroid plexus. Furthermore, we identified neuronal populations undergoing apoptosis, mediated by the Lcn2-SLC22A17 pathway in response to IL-1β and IL-6 signaling. Notably, the inhibition of pyroptosis using VX-765 significantly mitigated neurological impairments., Conclusions: Our study provides evidence that endothelial pyroptosis, characterized by the release of IL-1β and IL-6, triggers microglial polarization through NF-κB pathway activation, ultimately leading to microglia-mediated neuronal apoptosis in the choroid plexus post-ICH. These findings suggest that targeted therapeutic strategies aimed at mitigating endothelial cell pyroptosis and neutralizing inflammatory cytokines may offer neuroprotection for both microglia and neurons, presenting a promising avenue for ICH treatment., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Background: Hydrocephalus is characterized by secretion, circulation, and absorption disorder of cerebrospinal fluid (CSF) with high morbidity and complication rate. The relationship between inflammation and abnormal secretion of CSF by choroid plexus epithelium (CPE) had received more attention. In this study, we aim to detect the role of Toll-like receptor 4/nuclear factor-kappa B/Na+/K+/2Cl-cotransporter 1(TLR4/NF-κB/NKCC1) signal pathway in the development of hydrocephalus., Method: Hydrocephalus was induced in adult rats (8 weeks) by intracisternal kaolin injection, then pyrrolidinedithiocarbamate (PDTC) and bumetanide were administrated to the rats mode. Then the rat model was evaluated, and ventricular volume was calculated at different time points. Then CPE, cortex, preventricular tissue, and CSF were obtained. Protein expressions of TLR-4, NKCC/serine-threonine STE20/SPS1-related, proline-alanine-rich kinase (SPAK), pNKCC1, pSPAK, GFAP, AQP1, and AQP4 were measured by RT-PCR, western blot, and immunofluorescence (IF) stains in CPE, respectively., Result: Our data showed that inflammation factors tumor necrosis factor-(TNF-α), interleukin 18(IL-18), and glial fibrillary acidic protein (GFAP) concentrations were significantly higher in the model group than in controls. The TLR4/NF-κB/NKCC1 signal pathway were actived by NF-κB-p65, NKCC1, pNKCC1- pSPAK complex, and Aquaporin1 (AQP1) high expression. PDTC and bumetanide use can help regular TLR4/NF-κB/NKCC1 expression and reduced AQP1 expression by down-regulate NF-B-p65 and inhibiting NKCC1, respectively. As a result, the treatment groups alleviated CPE abnormal secretion and ventricle enlargement., Conclusion: These results confirmed that the inflammatory reaction contributes TLR4/NF-κB/NKCC1 mediated CPE abnormal secretion and consequent hydrocephalus. Regulation of TLR4/NF-κB/NKCC1 and AQP1 can prevent this process. Our study provides a strong rationale for further exploring alleviating CPE abnormal secretion as a therapeutic perspective of hydrocephalus., (© 2024 The Author(s). CNS Neuroscience & Therapeutics published by John Wiley & Sons Ltd.)

Background: The choroid plexus (CP) is an understudied tissue in the central nervous system and is primarily implicated in cerebrospinal fluid (CSF) production. CP also produces numerous neurotrophic factors (NTF) which circulate to different brain regions. Regulation of NTFs in the CP during natural aging is largely unknown. Here, we investigated the age and gender-specific transcription of NTFs along with the changes in the tight junctional proteins (TJPs) and the water channel protein Aquaporin (AQP1)., Methods: Male and female mice were used for our study. Age-related transcriptional changes were analyzed using quantitative PCR at three different time points: mature adult, middle-aged, and aged. Transcriptional changes during aging were further confirmed with digital droplet PCR. Additionally, we used immunohistochemical analysis (IHC) for the evaluation of in vivo protein expression. We further investigated the cellular phenotype of these NTFS, TJP, and water channel proteins in the mouse CP by co-labeling them with the classical vascular marker, Isolectin B4, and epithelial cell marker, Plectin., Results: Aging significantly altered NTF gene expression in the CP. Brain-derived neurotrophic factor (BDNF), Midkine (MDK), VGF, Insulin-like growth factor (IGF1), IGF2, Klotho (KL), Erythropoietin (EPO), and its receptor (EPOR) were reduced in the aged CP of males and females. Vascular endothelial growth factor (VEGF) transcription was gender-specific; in males, gene expression was unchanged in the aged CP, while females showed an age-dependent reduction. Age-dependent changes in VEGF localization were evident, from vasculature to epithelial cells. IGF2 and klotho localized in the basolateral membrane of the CP and showed an age-dependent reduction in epithelial cells. Water channel protein AQP1 localized in the tip of epithelial cells and showed an age-related reduction in mRNA and protein levels. TJP's JAM, CLAUDIN1, CLAUDIN2 and CLAUDIN5 were reduced in aged mice., Conclusions: Our study highlights transcriptional level changes in the CP during aging. The age-related transcriptional changes exhibit similarities as well as gene-specific differences in the CP of males and females. Altered transcription of the water channel protein AQP1 and TJPs could be involved in reduced CSF production during aging. Importantly, reduction in the neurotrophic factors and longevity factor Klotho can play a role in regulating brain aging., (© 2024. The Author(s).)

Manganese (Mn) is an essential trace mineral for brain function, but excessive accumulation can cause irreversible nervous system damage, highlighting the need for proper Mn balance. ZIP14, ZnT10, and ZIP8 are key transporters involved in maintaining Mn homeostasis, particularly in the absorption and excretion of Mn in the intestine and liver. However, their roles in the brain are less understood. The blood-cerebrospinal fluid barrier and the blood-brain barrier, formed by the choroid plexus and brain blood vessels, respectively, are critical for brain protection and brain metal homeostasis. This study identified ZIP14 on the choroid plexus epithelium, and ZIP8 and ZnT10 in brain microvascular tissue. We show that despite significant Mn accumulation in the CSF of Znt10 knockout mice, ZIP14 expression levels in the blood-cerebrospinal fluid barrier remain unchanged, indicating that ZIP14 does not have a compensatory mechanism for regulating Mn uptake in the brain in vivo. Additionally, Mn still enters the CSF without ZIP14 when systemic levels rise. This indicates that alternative transport mechanisms or compensatory pathways ensure Mn balance in the CSF, shedding light on potential strategies for managing Mn-related disorders.

Background: The choroid plexus (ChP) helps maintain the homeostasis of the brain by forming the blood-CSF barrier via tight junctions (TJ) at the choroid plexus epithelial cells, and subsequently preventing neuroinflammation by restricting immune cells infiltration into the central nervous system. However, whether chronic cerebral hypoperfusion causes ChP structural damage and blood-CSF barrier impairment remains understudied., Methods: The bilateral carotid stenosis (BCAS) model in adult male C57BL/6 J mice was used to induce cerebral hypoperfusion, a model for vascular contributions to cognitive impairment and dementia (VCID). BCAS-mediated changes of the blood-CSF barrier TJ proteins, apical secretory Na + -K + -Cl - cotransporter isoform 1 (NKCC1) protein and regulatory serine-threonine kinases SPAK, and brain infiltration of myeloid-derived immune cells were assessed., Results: BCAS triggered dynamic changes of TJ proteins (claudin 1, claudin 5) accompanied with stimulation of SPAK-NKCC1 complex and NF-κB in the ChP epithelial cells. These changes impacted the integrity of the blood-CSF barrier, as evidenced by ChP infiltration of macrophages/microglia, neutrophils and T cells. Importantly, pharmacological blockade of SPAK with its potent inhibitor ZT1a in BCAS mice attenuated brain immune cell infiltration and improved cognitive neurological function., Conclusions: BCAS causes chronic ChP blood-CSF damage and immune cell infiltration. Our study sheds light on the SPAK-NKCC1 complex as a therapeutic target in neuroinflammation., (© 2024. The Author(s).)

The choroid plexus (ChP) is a vital brain barrier and source of cerebrospinal fluid (CSF). Here, we use longitudinal two-photon imaging in awake mice and single-cell transcriptomics to elucidate the mechanisms of ChP regulation of brain inflammation. We used intracerebroventricular injections of lipopolysaccharides (LPS) to model meningitis in mice and observed that neutrophils and monocytes accumulated in the ChP stroma and surged across the epithelial barrier into the CSF. Bi-directional recruitment of monocytes from the periphery and, unexpectedly, macrophages from the CSF to the ChP helped eliminate neutrophils and repair the barrier. Transcriptomic analyses detailed the molecular steps accompanying this process and revealed that ChP epithelial cells transiently specialize to nurture immune cells, coordinating their recruitment, survival, and differentiation as well as regulation of the tight junctions that control the permeability of the ChP brain barrier. Collectively, we provide a mechanistic understanding and a comprehensive roadmap of neuroinflammation at the ChP brain barrier., Competing Interests: Declaration of interests J.O.-M. reports compensation for consulting services with Cellarity, Tessel Biosciences, and Radera Biotherapeutics. J.C. has been an employee of Dyne Therapeutics since April 2021. Her contributions to this manuscript were made when she was employed by Boston Children’s Hospital., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Background: The blood-cerebrospinal fluid barrier (BCSFB) comprises the choroid plexus epithelia. It is important for brain development, maintenance, function, and especially for maintaining immune homeostasis in the cerebrospinal fluid (CSF). Although previous studies have shown that the peripheral immune function of the body is impaired upon exposure to microgravity, no studies have reported changes in immune cells and cytokines in the CSF that reflect neuroimmune status. The purpose of this study is to investigate the alterations in cerebrospinal fluid (CSF) immune homeostasis induced by microgravity and its mechanisms. This research is expected to provide basic data for brain protection of astronauts during spaceflight., Methods: The proportions of immune cells in the CSF and peripheral blood (PB) of SMG rats were analyzed using flow cytometry. Immune function was evaluated by measuring cytokine concentrations using the Luminex method. The histomorphology and ultrastructure of the choroid plexus epithelia were determined. The concentrations of intercellular junction proteins in choroid plexus epithelial cells, including vascular endothelial-cadherin (VE-cadherin), zonula occludens 1 (ZO-1), Claudin-1 and occludin, were detected using western blotting and immunofluorescence staining to characterize BCSFB injury., Results: We found that SMG caused significant changes in the proportion of CD4 and CD8 T cells in the CSF and a significant increase in the levels of cytokines (GRO/KC, IL-18, MCP-1, and RANTES). In the PB, there was a significant decrease in the proportion of T cells and NKT cells and a significant increase in cytokine levels (GRO/KC, IL-18, MCP-1, and TNF-α). Additionally, we observed that the trends in immune markers in the PB and CSF were synchronized within specific SMG durations, suggesting that longer SMG periods (≥21 days) have a more pronounced impact on immune markers. Furthermore, 21d-SMG resulted in ultrastructural disruption and downregulated expression of intercellular junction proteins in rat choroid plexus epithelial cells., Conclusions: We found that SMG disrupts the BCSFB and affects the CSF immune homeostasis. This study provides new insights into the health protection of astronauts during spaceflight., (© 2024 The Author(s). Brain and Behavior published by Wiley Periodicals LLC.)

Background: Folates are a family of B 9 vitamins essential for normal growth and development in the central nervous system (CNS). Transport of folates is mediated by three major transport proteins: folate receptor alpha (FRα), proton-coupled folate transporter (PCFT), and reduced folate carrier (RFC). Brain folate uptake occurs at the choroid plexus (CP) epithelium through coordinated actions of FRα and PCFT, or directly into brain parenchyma at the vascular blood-brain barrier (BBB), mediated by RFC. Impaired folate transport can occur due to loss of function mutations in FRα or PCFT, resulting in suboptimal CSF folate levels. Our previous reports have demonstrated RFC upregulation by nuclear respiratory factor-1 (NRF-1) once activated by the natural compound pyrroloquinoline quinone (PQQ). More recently, we have identified folate transporter localization at the arachnoid barrier (AB). The purpose of the present study was to further characterize folate transporters localization and function in AB cells, as well as their regulation by NRF-1/PGC-1α signaling and folate deficiency., Methods: In immortalized mouse AB cells, polarized localization of RFC and PCFT was assessed by immunocytochemical analysis, with RFC and PCFT functionality examined with transport assays. The effects of PQQ treatment on changes in RFC functional expression were also investigated. Mouse AB cells grown in folate-deficient conditions were assessed for changes in gene expression of the folate transporters, and other key transporters and tight junction proteins., Results: Immunocytochemical analysis revealed apical localization of RFC at the mouse AB epithelium, with PCFT localized on the basolateral side and within intracellular compartments. PQQ led to significant increases in RFC functional expression, mediated by activation of the NRF-1/PGC-1α signalling cascade. Folate deficiency led to significant increases in expression of RFC, MRP3, P-gp, GLUT1 and the tight junction protein claudin-5., Conclusion: These results uncover the polarized expression of RFC and PCFT at the AB, with induction of RFC functional expression by activation of the NRF-1/PGC-1α signalling pathway and folate deficiency. These results suggest that the AB may contribute to the flow of folates into the CSF, representing an additional pathway when folate transport at the CP is impaired., (© 2024. The Author(s).)

Background: Group B Streptococcus (GBS) is the leading cause of neonatal meningitis responsible for a substantial cause of death and disability worldwide. The vast majority of GBS neonatal meningitis cases are due to the CC17 hypervirulent clone. However, the cellular and molecular pathways involved in brain invasion by GBS CC17 isolates remain largely elusive. Here, we studied the specific interaction of the CC17 clone with the choroid plexus, the main component of the blood-cerebrospinal fluid (CSF) barrier., Methods: The interaction of GBS CC17 or non-CC17 strains with choroid plexus cells was studied using an in vivo mouse model of meningitis and in vitro models of primary and transformed rodent choroid plexus epithelial cells (CPEC and Z310). In vivo interaction of GBS with the choroid plexus was assessed by microscopy. Bacterial invasion and cell barrier penetration were examined in vitro, as well as chemokines and cytokines in response to infection., Results: GBS CC17 was found associated with the choroid plexus of the lateral, 3rd and 4th ventricles. Infection of choroid plexus epithelial cells revealed an efficient internalization of the bacteria into the cells with GBS CC17 displaying a greater ability to invade these cells than a non-CC17 strain. Internalization of the GBS CC17 strain involved the CC17-specific HvgA adhesin and occurred via a clathrin-dependent mechanism leading to transcellular transcytosis across the choroid plexus epithelial monolayer. CPEC infection resulted in the secretion of several chemokines, including CCL2, CCL3, CCL20, CX3CL1, and the matrix metalloproteinase MMP3, as well as immune cell infiltration., Conclusion: Our findings reveal a GBS strain-specific ability to infect the blood-CSF barrier, which appears to be an important site of bacterial entry and an active site of immune cell trafficking in response to infection., (© 2024. The Author(s).)

The brain's choroid plexus (CP), which operates as an anatomical and functional 'checkpoint', regulates the communication between brain and periphery and contributes to the maintenance of healthy brain homeostasis throughout life. Evidence from mouse models and humans reveals a link between loss of CP checkpoint properties and dysregulation of the CP immune milieu as a conserved feature across diverse neurological conditions. In particular, we suggest that an imbalance between different immune signals at the CP, including CD4 + T cell-derived cytokines, type-I interferon, and complement components, can perpetuate brain inflammation and cognitive deterioration in aging and neurodegeneration. Furthermore, we highlight the role of CP metabolism in controlling CP inflammation, and propose that targeting molecules that regulate CP metabolism could be effective in safeguarding brain function., Competing Interests: Declaration of Interests M.S. is an inventor of the intellectual property that forms the basis for development of PD-L1 immunotherapy for AD. M.S. is the cofounder of ImmunoBrain Checkpoint (IBC). The other authors declare no competing interests in relation to this work., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

The human polyomavirus JCPyV is an opportunistic pathogen that infects greater than 60% of the world's population. The virus establishes a persistent and asymptomatic infection in the urogenital system but can cause a fatal demyelinating disease in immunosuppressed or immunomodulated patients following invasion of the CNS. The mechanisms responsible for JCPyV invasion into CNS tissues are not known but direct invasion from the blood to the cerebral spinal fluid via the choroid plexus has been hypothesized. To study the potential of the choroid plexus as a site of neuroinvasion, we used an adult human choroid plexus epithelial cell line to model the blood-cerebrospinal fluid (B-CSF) barrier in a transwell system. We found that these cells formed a highly restrictive barrier to virus penetration either as free virus or as virus associated with extracellular vesicles (EVJC+). The restriction was not absolute and small amounts of virus or EVJC+ penetrated and were able to establish foci of infection in primary astrocytes. Disruption of the barrier with capsaicin did not increase virus or EVJC+ penetration leading us to hypothesize that virus and EVJC+ were highly cell-associated and crossed the barrier by an active process. An inhibitor of macropinocytosis increased virus penetration from the basolateral (blood side) to the apical side (CSF side). In contrast, inhibitors of clathrin and raft dependent transcytosis reduced virus transport from the basolateral to the apical side of the barrier. None of the drugs inhibited apical to basolateral transport suggesting directionality. Pretreatment with cyclosporin A, an inhibitor of P-gp, MRP2 and BCRP multidrug resistance transporters, restored viral penetration in cells treated with raft and clathrin dependent transcytosis inhibitors. Because choroid plexus epithelial cells are known to be susceptible to JCPyV infection both in vitro and in vivo we also examined the release of infectious virus from the barrier. We found that virus was preferentially released from the cells into the apical (CSF) chamber. These data show clearly that there are two mechanisms of penetration, direct transcytosis which is capable of seeding the CSF with small amounts of virus, and infection followed by directional release of infectious virions into the CSF compartment., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 O’Hara et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Background: Structural and functional changes of the choroid plexus (ChP) have been reported in Alzheimer's disease (AD). Nonetheless, the role of the ChP in the pathogenesis of AD remains largely unknown. We aim to unravel the relation between ChP functioning and core AD pathogenesis using a unique proteomic approach in mice and humans., Methods: We used an APP knock-in mouse model, APP NL-G-F , exhibiting amyloid pathology, to study the association between AD brain pathology and protein changes in mouse ChP tissue and CSF using liquid chromatography mass spectrometry. Mouse proteomes were investigated at the age of 7 weeks (n = 5) and 40 weeks (n = 5). Results were compared with previously published human AD CSF proteomic data (n = 496) to identify key proteins and pathways associated with ChP changes in AD., Results: ChP tissue proteome was dysregulated in APP NL-G-F mice relative to wild-type mice at both 7 and 40 weeks. At both ages, ChP tissue proteomic changes were associated with epithelial cells, mitochondria, protein modification, extracellular matrix and lipids. Nonetheless, some ChP tissue proteomic changes were different across the disease trajectory; pathways related to lysosomal function, endocytosis, protein formation, actin and complement were uniquely dysregulated at 7 weeks, while pathways associated with nervous system, immune system, protein degradation and vascular system were uniquely dysregulated at 40 weeks. CSF proteomics in both mice and humans showed similar ChP-related dysregulated pathways., Conclusions: Together, our findings support the hypothesis of ChP dysfunction in AD. These ChP changes were related to amyloid pathology. Therefore, the ChP could become a novel promising therapeutic target for AD., (© 2024. The Author(s).)

Background: The cerebrospinal fluid (CSF), primarily generated by the choroid plexus (ChP), is the major carrier of the glymphatic system. The alternations of CSF production and the ChP can be associated with the Alzheimer's disease (AD). The present work investigated the roles of the ChP in the AD based on a proposed ChP image segmentation pipeline., Methods: A human-in-the-loop ChP image segmentation pipeline was implemented with intermediate and active learning datasets. The performance of the proposed pipeline was evaluated on manual contours by five radiologists, compared to the FreeSurfer and FastSurfer toolboxes. The ChP volume and blood flow were investigated among AD groups. The correlations between the ChP volume and AD CSF biomarkers including phosphorylated tau (p-tau), total tau (t-tau), amyloid-β42 (Aβ42), and amyloid-β40 (Aβ40) was investigated using three models (univariate, multiple variables, and stepwise regression) on two datasets with 806 and 320 subjects., Results: The proposed ChP segmentation pipeline achieved superior performance with a Dice coefficient of 0.620 on the test dataset, compared to the FreeSurfer (0.342) and FastSurfer (0.371). Significantly larger volumes (p < 0.001) and higher perfusion (p = 0.032) at the ChP were found in AD compared to CN groups. Significant correlations were found between the tau and the relative ChP volume (the ChP volume and ChP/parenchyma ratio) in each patient groups and in the univariate regression analysis (p < 0.001), the multiple regression model (p < 0.05 except for the t-tau in the LMCI), and in the step-wise regression model (p < 0.021). In addition, the correlation coefficients changed from - 0.32 to - 0.21 along with the AD progression in the multiple regression model. In contrast, the Aβ42 and Aβ40 shows consistent and significant associations with the lateral ventricle related measures in the step-wise regression model (p < 0.027)., Conclusions: The proposed pipeline provided accurate ChP segmentation which revealed the associations between the ChP and tau level in the AD. The proposed pipeline is available on GitHub ( https://github.com/princeleeee/ChP-Seg )., (© 2024. The Author(s).)

The brain's neuroreparative capacity after injuries such as ischemic stroke is partly contained in the brain's neurogenic niches, primarily the subventricular zone (SVZ), which lies in close contact with the cerebrospinal fluid (CSF) produced by the choroid plexus (ChP). Despite the wide range of their proposed functions, the ChP/CSF remain among the most understudied compartments of the central nervous system (CNS). Here, we report a mouse genetic tool (the ROSA26iDTR mouse line) for noninvasive, specific, and temporally controllable ablation of CSF-producing ChP epithelial cells to assess the roles of the ChP and CSF in brain homeostasis and injury. Using this model, we demonstrate that ChP ablation causes rapid and permanent CSF volume loss in both aged and young adult brains, accompanied by disruption of ependymal cilia bundles. Surprisingly, ChP ablation did not result in overt neurological deficits at 1 mo postablation. However, we observed a pronounced decrease in the pool of SVZ neuroblasts (NBs) following ChP ablation, which occurs due to their enhanced migration into the olfactory bulb. In the middle cerebral artery occlusion model of ischemic stroke, NB migration into the lesion site was also reduced in the CSF-depleted mice. Thus, our study establishes an important role of ChP/CSF in regulating the regenerative capacity of the adult brain under normal conditions and after ischemic stroke., Competing Interests: Competing interests statement:The authors declare no competing interest.

The cells of the choroid plexus (CP) epithelium are specialized ependymal cells (ECs) but have distinct properties. The CP cells and ECs form single-cell sheets contiguous to each other at a transitional zone. The CP is underlined by a basal lamina and has barrier properties, whereas the ECs do not. The basal lamina of the CP is continuous with the glia limitans superficialis and, consequently, the CP stroma is continuous with the meninges along entering blood vessels. The CP has previously been reported to express aquaporin-1 (AQP1) mostly apically, and ECs show mostly basolateral aquaporin-4 (AQP4) expression. Recent evidence in various systems has shown that in changing conditions the expression and distribution of AQP4 can be modified, involving phosphorylation and calmodulin-triggered translocation. Studies on the human CP revealed that AQP4 is also expressed in some CP cells, which is likely to be increased during ageing based on mouse data. Moreover, subependymal astrocytic processes in the ependyma-CP transition, forming a glial plate around blood vessels and facing the CP stroma, were strongly positive for AQP4. We propose that the increased AQP4 expression might be a compensatory mechanism for the observed reduction in CSF production in the ageing human brain. The high AQP4 density in the transition zone might facilitate the transport of water into and out of the CP stroma and serve as a drainage and clearing pathway for metabolites in the CNS., (© 2023 The Authors. The Journal of Physiology published by John Wiley & Sons Ltd on behalf of The Physiological Society.)

The choroid plexus (ChP) produces cerebrospinal fluid (CSF). It also contributes to brain development and serves as the CSF-blood barrier. Prior studies have identified transporters on the epithelial cells that transport water and ions from the blood vasculature to the ventricles and tight junctions involved in the CSF-blood barrier. Yet, how the ChP epithelial cells control brain physiology remains unresolved. We use zebrafish to provide insights into the physiological roles of the ChP. Upon histological and transcriptomic analyses, we identify that the zebrafish ChP is conserved with mammals and expresses transporters involved in CSF secretion. Next, we show that the ChP epithelial cells secrete proteins into CSF. By ablating the ChP epithelial cells, we identify a reduction of the ventricular sizes without alterations of the CSF-blood barrier. Altogether, our findings reveal that the zebrafish ChP is conserved and contributes to the size and homeostasis of the brain ventricles., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Why individuals with Down syndrome (DS) are more susceptible to SARS-CoV-2-induced neuropathology remains elusive. Choroid plexus (ChP) plays critical roles in barrier function and immune response modulation and expresses the ACE2 receptor and the chromosome 21-encoded TMPRSS2 protease, suggesting its substantial role in establishing SARS-CoV-2 infection in the brain. To explore this, we established brain organoids from DS and isogenic euploid iPSC that consist of a core of functional cortical neurons surrounded by a functional ChP-like epithelium (ChPCOs). DS-ChPCOs recapitulated abnormal DS cortical development and revealed defects in ciliogenesis and epithelial cell polarity in ChP-like epithelium. We then demonstrated that the ChP-like epithelium facilitates infection and replication of SARS-CoV-2 in cortical neurons and that this is increased in DS. Inhibiting TMPRSS2 and furin activity reduced viral replication in DS-ChPCOs to euploid levels. This model enables dissection of the role of ChP in neurotropic virus infection and euploid forebrain development and permits screening of therapeutics for SARS-CoV-2-induced neuropathogenesis.

Choroid plexus (ChP), the brain structure primarily responsible for cerebrospinal fluid production, contains a robust circadian clock, whose role remains to be elucidated. The aim of our study was to [1] identify rhythmically controlled cellular processes in the mouse ChP and [2] assess the role and nature of signals derived from the master clock in the suprachiasmatic nuclei (SCN) that control ChP rhythms. To accomplish this goal, we used various mouse models (WT, mPer2 Luc , ChP-specific Bmal1 knockout) and combined multiple experimental approaches, including surgical lesion of the SCN (SCNx), time-resolved transcriptomics, and single cell luminescence microscopy. In ChP of control (Ctrl) mice collected every 4 h over 2 circadian cycles in darkness, we found that the ChP clock regulates many processes, including the cerebrospinal fluid circadian secretome, precisely times endoplasmic reticulum stress response, and controls genes involved in neurodegenerative diseases (Alzheimer's disease, Huntington's disease, and frontotemporal dementia). In ChP of SCNx mice, the rhythmicity detected in vivo and ex vivo was severely dampened to a comparable extent as in mice with ChP-specific Bmal1 knockout, and the dampened cellular rhythms were restored by daily injections of dexamethasone in mice. Our data demonstrate that the ChP clock controls tissue-specific gene expression and is strongly dependent on the presence of a functional connection with the SCN. The results may contribute to the search for a novel link between ChP clock disruption and impaired brain health., (© 2024. The Author(s).)

GH acts in numerous organs expressing the GH receptor (GHR), including the brain. However, the mechanisms behind the brain's permeability to GH and how this hormone accesses different brain regions remain unclear. It is well-known that an acute GH administration induces phosphorylation of the signal transducer and activator of transcription 5 (pSTAT5) in the mouse brain. Thus, the pattern of pSTAT5 immunoreactive cells was analyzed at different time points after IP or intracerebroventricular GH injections. After a systemic GH injection, the first cells expressing pSTAT5 were those near circumventricular organs, such as arcuate nucleus neurons adjacent to the median eminence. Both systemic and central GH injections induced a medial-to-lateral pattern of pSTAT5 immunoreactivity over time because GH-responsive cells were initially observed in periventricular areas and were progressively detected in lateral brain structures. Very few choroid plexus cells exhibited GH-induced pSTAT5. Additionally, Ghr mRNA was poorly expressed in the mouse choroid plexus. In contrast, some tanycytes lining the floor of the third ventricle expressed Ghr mRNA and exhibited GH-induced pSTAT5. The transport of radiolabeled GH into the hypothalamus did not differ between wild-type and dwarf Ghr knockout mice, indicating that GH transport into the mouse brain is GHR independent. Also, single-photon emission computed tomography confirmed that radiolabeled GH rapidly reaches the ventral part of the tuberal hypothalamus. In conclusion, our study provides novel and valuable information about the pattern and mechanisms behind GH transport into the mouse brain., (© The Author(s) 2024. Published by Oxford University Press on behalf of the Endocrine Society. All rights reserved. For commercial re-use, please contact reprints@oup.com for reprints and translation rights for reprints. All other permissions can be obtained through our RightsLink service via the Permissions link on the article page on our site—for further information please contact journals.permissions@oup.com. See the journal About page for additional terms.)

Hibernation is an extreme state of seasonal energy conservation, reducing metabolic rate to as little as 1% of the active state. During the hibernation season, many species of hibernating mammals cycle repeatedly between the active (aroused) and hibernating (torpid) states (T-A cycling), using brown adipose tissue (BAT) to drive cyclical rewarming. The regulatory mechanisms controlling this process remain undefined but are presumed to involve thermoregulatory centres in the hypothalamus. Here, we used the golden hamster (Mesocricetus auratus), and high-resolution monitoring of BAT, core body temperature and ventilation rate, to sample at precisely defined phases of the T-A cycle. Using c-fos as a marker of cellular activity, we show that although the dorsomedial hypothalamus is active during torpor entry, neither it nor the pre-optic area shows any significant changes during the earliest stages of spontaneous arousal. Contrastingly, in three non-neuronal sites previously linked to control of metabolic physiology over seasonal and daily time scales - the choroid plexus, pars tuberalis and third ventricle tanycytes - peak c-fos expression is seen at arousal initiation. We suggest that through their sensitivity to factors in the blood or cerebrospinal fluid, these sites may mediate metabolic feedback-based initiation of the spontaneous arousal process., Competing Interests: Competing interests The authors declare no competing or financial interests., (© 2024. Published by The Company of Biologists Ltd.)

Donepezil (DNPZ) is a cholinesterase inhibitor used for the management of Alzheimer's disease (AD) and is dependent on membrane transporters such as ABCG2 to actively cross brain barriers and reach its target site of action in the brain. Located in the brain ventricles, the choroid plexus (CP) forms an interface between the cerebrospinal fluid (CSF) and the bloodstream, known as the blood-CSF barrier (BCSFB). Historically, the BCSFB has received little attention as a potential pathway for drug delivery to the central nervous system (CNS). Nonetheless, this barrier is presently viewed as a dynamic transport interface that limits the traffic of molecules into and out of the CNS through the presence of membrane transporters, with parallel activity with the BBB. The localization and expression of drug transporters in brain barriers represent a huge obstacle for drug delivery to the brain and a major challenge for the development of therapeutic approaches to CNS disorders. The widespread interest in understanding how circadian clocks modulate many processes that define drug delivery in order to predict the variability in drug safety and efficacy is the next bridge to improve effective treatment. In this context, this study aims at characterizing the circadian expression of ABCG2 and DNPZ circadian transport profile using an in vitro model of the BCSFB. We found that ABCG2 displays a circadian pattern and DNPZ is transported in a circadian way across this barrier. This study will strongly impact on the capacity to modulate the BCSFB in order to control the penetration of DNPZ into the brain and improve therapeutic strategies for the treatment of AD according to the time of the day.

The brain's ventricles are filled with a colorless fluid known as cerebrospinal fluid (CSF). When there is an excessive accumulation of CSF in the ventricles, it can result in high intracranial pressure, ventricular enlargement, and compression of the surrounding brain tissue, leading to potential damage. This condition is referred to as hydrocephalus. Hydrocephalus is classified into two categories: congenital and acquired. Congenital hydrocephalus (CH) poses significant challenges for affected children and their families, particularly in resource-poor countries. Recognizing the psychological and economic impacts is crucial for developing interventions and support systems that can help alleviate the distress and burden faced by these families. As our understanding of CSF production and circulation improves, we are gaining clearer insights into the causes of CH. In this article, we will summarize the current knowledge regarding CSF circulation pathways and the underlying causes of CH. The main causes of CH include abnormalities in the FoxJ1 pathway of ventricular cilia, dysfunctions in the choroid plexus transporter Na + -K + -2Cl - contransporter isoform 1, developmental abnormalities in the cerebral cortex, and structural abnormalities within the brain. Understanding the causes of CH is indeed crucial for advancing research and developing effective treatment strategies. In this review, we will summarize the findings from existing studies on the causes of CH and propose potential research directions to further our understanding of this condition., (© 2024. The Author(s).)

Background: In the choroid plexus and pituitary gland, vasculature is known to have a permeable, fenestrated phenotype which allows for the free passage of molecules in contrast to the blood brain barrier observed in the rest of the CNS. The endothelium of these compartments, along with secretory, neural-lineage cells (choroid epithelium and pituitary endocrine cells) have been studied in detail, but less attention has been given to the perivascular mesenchymal cells of these compartments., Methods: The Hic1 CreERT2 Rosa26 LSL-TdTomato mouse model was used in conjunction with a Pdgfra H2B-EGFP mouse model to examine mesenchymal cells, which can be subdivided into Pdgfra + fibroblasts and Pdgfra - pericytes within the choroid plexus (CP) and pituitary gland (PG), by histological, immunofluorescence staining and single-cell RNA-sequencing analyses., Results: We found that both CP and PG possess substantial populations of distinct Hic1 + mesenchymal cells, including an abundance of Pdgfra + fibroblasts. Within the pituitary, we identified distinct subpopulations of Hic1 + fibroblasts in the glandular anterior pituitary and the neurosecretory posterior pituitary. We also identified multiple distinct markers of CP, PG, and the meningeal mesenchymal compartment, including alkaline phosphatase, indole-n-methyltransferase and CD34., Conclusions: Novel, distinct subpopulations of mesenchymal cells can be found in permeable vascular interfaces, including the CP, PG, and meninges, and make distinct contributions to both organs through the production of structural proteins, enzymes, transporters, and trophic molecules., (© 2024. The Author(s).)

The cerebrospinal fluid (CSF) fills the brain ventricles and the subarachnoid space surrounding the brain and spinal cord. The fluid compartment of the brain ventricles communicates with the interstitial fluid of the brain across the ependyma. In comparison to blood, the CSF contains very little protein to buffer acid-base challenges. Nevertheless, the CSF responds efficiently to changes in systemic pH by mechanisms that are dependent on the CO 2 /HCO 3 - buffer system. This is evident from early studies showing that the CSF secretion is sensitive to inhibitors of acid/base transporters and carbonic anhydrase. The CSF is primarily generated by the choroid plexus, which is a well-vascularized structure arising from the pial lining of the brain ventricles. The epithelial cells of the choroid plexus host a range of acid/base transporters, many of which participate in CSF secretion and most likely contribute to the transport of acid/base equivalents into the ventricles. This review describes the current understanding of the molecular mechanisms in choroid plexus acid/base regulation and the possible role in CSF pH regulation., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Epithelial cells comprising the choroid plexus (CP) form a crucial barrier between the blood and the cerebrospinal fluid, thereby assuming a central position in brain homeostasis and signaling. Mounting evidence suggests that the impairment of CP function may be a significant contributor to Alzheimer's disease (AD) pathogenesis. CP function relies on the expression of specific receptors, and the potential involvement of olfactory receptors (ORs) and taste receptors (TASRs) in chemical surveillance within the CP is being investigated. Previous studies have implicated ORs and TASRs in neurodegenerative disorders like AD, although the direct evidence of their expression in the human CP remains to be established. In this study, we conducted a transcriptomic analysis encompassing eleven ORs and TASRs in the CP, comparing samples from healthy age-matched controls to those from patients with AD spanning Braak stages I to VI. Among these receptors, a striking finding emerged- OR2K2 exhibited robust expression, with a statistically significant upregulation noted at Braak stage I. Surprisingly, at the protein level, OR2K2 showed a significant decrease in both Braak stage I and VI. Additionally, we identified CP epithelial cells as the source of OR2K2 expression, where it colocalized with autophagy markers LC3 and p62. We postulate that OR2K2 could be subjected to degradation by autophagy in the early stages of AD, triggering a compensatory mechanism that leads to increased OR2K2 mRNA transcription. This study uncovers a potential role for OR2K2 in AD pathogenesis, offering a novel perspective on the intricate dynamics at play in this neurodegenerative disorder.

Background: The dysregulation of the gut-brain axis in chronic inflammatory bowel diseases can cause neuro-psychological disturbances, but the underlying mechanisms are still not fully understood. The choroid plexus (CP) maintains brain homeostasis and nourishment through the secretion and clearance of cerebrospinal fluid. Recent research has demonstrated the existence of a CP vascular barrier in mice which is modulated during intestinal inflammation. This study investigates possible correlations between CP modifications and inflammatory activity in patients with Crohn's disease (CD)., Methods: In this prospective study, 17 patients with CD underwent concomitant abdominal and brain 3 T MRI. The volume and permeability of CP were compared with levels of C-reactive protein (CRP), fecal calprotectin (FC), sMARIA and SES-CD scores., Results: The CP volume was negatively correlated with CRP levels (R = -0.643, p-value = 0.024) and FC (R = -0.571, p-value = 0.050). DCE metrics normalized by CP volume were positively correlated with CRP (K-trans: R = 0.587, p-value = 0.045; Vp: R = 0.706, p-value = 0.010; T1: R = 0.699, p-value = 0.011), and FC (Vp: R = 0.606, p-value = 0.037)., Conclusions: Inflammatory activity in patients with CD is associated with changes in CP volume and permeability, thus supporting the hypothesis that intestinal inflammation could affect the brain through the modulation of CP vascular barrier also in humans., Competing Interests: Declaration of competing interest The following authors disclose conflicts of interest: Federica Furfaro received consulting fees from Amgen, AbbVie and lecture fees from Janssen and Pfizer; Mariangela Allocca received consulting fees from Nikkiso Europe, Mundipharma, Janssen, AbbVie and Pfizer; Gionata Fiorino received consultancy fees from Ferring, MSD, AbbVie, Takeda, Janssen, Amgen, Sandoz, Samsung Bioepis, Celltrion; Silvio Danese served as a speaker, consultant, and advisory board member for Schering-Plow, Abbott (AbbVie) Laboratories, Merck, UCB Pharma, Ferring, Cellerix, Millenium Takeda, Nycomed, Pharmacosmos, Actelion, Alfa Wasserman, Genentech, Grunenthal, Pfizer, AstraZeneca, Novo Nordisk, Cosmo Pharmaceuticals, Vifor, and Johnson and Johnson; Alessandro Armuzzi has received consulting and/or advisory board fees from AbbVie, Allergan, Amgen, Arena, Biogen, Boehringer Ingelheim, Bristol-Myers Squibb, Celltrion, Eli-Lilly, Ferring, Galapagos, Gilead, Janssen, MSD, Mylan, Pfizer, Protagonist Therapeutics, Roche, Samsung Bioepis, Sandoz, and Takeda; lecture and/or speaker bureau fees from AbbVie, Amgen, Arena, Biogen, Bristol-Myers Squibb, Celltrion, Eli-Lilly, Ferring, Galapagos, Gilead, Janssen, MSD, Mitsubishi Tanabe, Novartis, Pfizer, Roche, Samsung Bioepis, Sandoz, Takeda, and Tigenix; and research grants from MSD, Pfizer, Takeda and Biogen; Maria Rescigno reports payment or honoraria for speaking engagements from Fondazione Internazionale Menarini; Riccardo Levi has received funds from Esaote. The authors have no other relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript apart from those disclosed. All remaining authors declare no financial or non-financial competing interests. All authors have submitted the ICMJE Form for Disclosure of Potential Conflicts of Interest., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

The choroid plexus (ChP) in the brain ventricles has a major influence on brain homeostasis. In this study, we aimed to determine whether the circadian clock located in ChP is affected by chronodisruption caused by misalignment with the external light/dark cycle and/or inflammation. Adult mPer2 Luc mice were maintained in the LD12:12 cycle or exposed to one of two models of chronic chronodisruption - constant light for 22-25 weeks (cLL) or 6-hour phase advances of the LD12:12 cycle repeated weekly for 12 weeks (cLD-shifts). Locomotor activity was monitored before the 4 th ventricle ChP and suprachiasmatic nuclei (SCN) explants were recorded in real time for PER2-driven population and single-cell bioluminescence rhythms. In addition, plasma immune marker concentrations and gene expression in ChP, prefrontal cortex, hippocampus and cerebellum were analyzed. cLL dampened the SCN clock but did not shorten the inactivity interval (sleep). cLD-shifts had no effect on the SCN clock, but transiently affected sleep duration and fragmentation. Both chronodisruption protocols dampened the ChP clock. Although immune markers were elevated in plasma and hippocampus, levels in ChP were unaffected, and unlike the liver clock, the ChP clock was resistant to lipopolysaccharide treatment. Importantly, both chronodisruption protocols reduced glucocorticoid signaling in ChP. The data demonstrate the high resistance of the ChP clock to inflammation, highlighting its role in protecting the brain from neuroinflammation, and on the other hand its high sensitivity to chronodisruption. Our results provide a novel link between human lifestyle-induced chronodisruption and the impairment of ChP-dependent brain homeostasis., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

The choroid plexus is composed of epithelial cells situated on the basal layer. The tight junctions between adjacent choroid plexus epithelial cells establish the blood-cerebrospinal fluid barrier. This barrier, in conjunction with the blood-brain barrier, is crucial for the homeostasis of the brain microenvironment. The choroid plexus epithelium secretes cerebrospinal fluid, growth factors, neuropeptides, and lipids into the ventricles and also serves as a gateway for immune cells to enter the brain. The pathophysiology of aging and neurodegenerative diseases remains largely enigmatic, with an increasing body of research linking the choroid plexus to the etiology of these age-related disorders. In this review, we summarize the known relationship between the choroid plexus epithelium and age-related diseases, aiming to provide new therapeutic clues for these disorders.

Depressed patients exhibit altered levels of immune-inflammatory markers both in the peripheral blood and in the cerebrospinal fluid (CSF) and inflammatory processes have been widely implicated in the pathophysiology of mood disorders. The Choroid Plexus (ChP), located at the base of each of the four brain ventricles, regulates the exchange of substances between the blood and CSF and several evidence supported a key role for ChP as a neuro-immunological interface between the brain and circulating immune cells. Given the role of ChP as a regulatory gate between periphery, CSF spaces and the brain, we compared ChP volumes in patients with bipolar disorder (BP) or major depressive disorder (MDD) and healthy controls, exploring their association with history of illness and levels of circulating cytokines. Plasma levels of inflammatory markers and MRI scans were acquired for 73 MDD, 79 BD and 72 age- and sex-matched healthy controls (HC). Patients with either BD or MDD had higher ChP volumes than HC. With increasing age, the bilateral ChP volume was larger in patients, an effect driven by the duration of illness; while only minor effects were observed in HC. Right ChP volumes were proportional to higher levels of circulating cytokines in the clinical groups, including IFN-γ, IL-13 and IL-17. Specific effects in the two diagnostic groups were observed when considering the left ChP, with positive association with IL-1ra, IL-13, IL-17, and CCL3 in BD, and negative associations with IL-2, IL-4, IL-1ra, and IFN-γ in MDD. These results suggest that ChP could represent a reliable and easy-to-assess biomarker to evaluate the brain effects of inflammatory status in mood disorders, contributing to personalized diagnosis and tailored treatment strategies., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Membrane transporters expressed in the choroid plexus (CP) are involved in the transport of substances between the blood and cerebrospinal fluid (CSF). Carnitine/organic cation transporter 1 (OCTN1, also known as SLC22A4) is expressed in rodent CP; however, its specific roles in blood-CSF transport remain unclear. Therefore, in this study, we aimed to evaluate the potential role of OCTN1 in the elimination of substances from CSF. Tritium-labeled ergothioneine ([ 3 H]ERGO), a typical in vivo substrate of OCTN1, was injected into the lateral ventricles of wild-type and octn1 gene knockout (octn1 -/- ) mice. Clearance of [ 3 H]ERGO from CSF was higher than that of the bulk flow marker, [ 14 C]mannitol, in wild-type mice. However, [ 3 H]ERGO clearance was significantly lower in octn1 -/- mice than in wild-type mice. Furthermore, OCTN1 expression in CP was determined via immunohistochemical analysis. CP/CSF ratio of [ 3 H]ERGO was significantly lower in octn1 -/- mice than in wild-type mice. These results suggest that OCTN1 is functionally expressed in CP and involved in the elimination of ERGO from CSF in mice.

Purpose: Choroid plexus carcinomas (CPCs) are extremely rare brain tumors and carry a dismal prognosis. Treatment options are limited and there is an urgent need to develop models to further research. In the present study, we established two CPC cell lines and performed multi-omics analyses. These cell lines serve as valuable models to propose new treatments in these rare but deadly brain tumors., Methods: Multi-omic profiling including, (i) methylation array (EPIC 850 K), (ii) whole genome sequencing (WGS), (iii) CANCERPLEX cancer genome panel testing, (iv) RNA sequencing (RNA-seq), and (v) proteomics analyses were performed in CCHE-45 and NGT131 cell lines., Results: Both cell lines were classified as methylation class B. Both harbored pathogenic TP53 point mutations; CCHE-45 additionally displayed TP53 loss. Furthermore, alterations of the NOTCH and WNT pathways were also detected in both cell lines. Two protein-coding gene fusions, BZW2-URGCP, and CTTNBP2-ERBB4, mutations of two oncodrivers, GBP-4 and KRTAP-12-2, and several copy number alterations were observed in CCHE-45, but not NGT131. Transcriptome and proteome analysis identified shared and unique signatures, suggesting that variability in choroid plexus carcinoma tumors may exist. The discovered difference's importance and implications highlight the possible diversity of choroid plexus carcinoma and call for additional research to fully understand disease pathogenesis., Conclusion: Multi-omics analyses revealed that the two choroid plexus carcinoma cell lines shared TP53 mutations and other common pathway alterations and activation of NOTCH and WNT pathways. Noticeable differences were also observed. These cell lines can serve as valuable models to propose new treatments in these rare but deadly brain tumors., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Purpose: Our aims were to investigate the presence of choroid plexus (CP) inflammation in chronic-phase intracerebral hemorrhage (ICH) patients and to characterize any inflammatory cells in the CP., Patients and Methods: An in vivo 18 F-DPA714 PET study was undertaken in 22 chronic-phase ICH patients who were admitted to the First Affiliated Hospital of Fujian Medical University or Tianjin Medical University General Hospital from April 2017 to June 2020. Ten control participants with nonhemorrhagic central nervous system diseases were included. Choroid plexus 18 F-DPA714 uptake was calculated as the average SUVR. To aid the interpretation of the 18 F-DPA714 uptake results at the CP level, Cy5-DPA714 in vivo imaging and immunofluorescence staining were used to show the presence of CP inflammation in an ICH mouse model during the chronic phase (14 weeks after ICH). Then immunofluorescence staining against translocator protein and other specific biomarkers was used to characterize the cells present in the inflamed CP of ICH mice in the chronic phase., Results: PET imaging showed that CP DPA714 SUVRs in chronic-phase ICH patients were higher than in controls (mean CP SUVR ± SD; ICH group: 1.05 ± 0.35; control group: 0.81 ± 0.21; P = 0.006). Immunofluorescence staining of the CP in ICH model mice identified a population of CD45 + immune cells, peripheral monocyte-derived CD14 + cells, CD68 + phagocytes, and CD11b + resident microglia/macrophages expressing translocator protein, possibly contributing to the increased 18 F-DPA714 uptake., Conclusions: Our study shows that CP DPA714 uptake in chronic-phase ICH patients was higher than that of participants with nonhemorrhagic central nervous system diseases, which means that CP inflammation is still active in chronic-phase ICH patients., Competing Interests: Conflicts of interest and sources of funding: The authors declared no conflicts of interest. The study was supported by the National Natural Science Foundation of China (grant no. 82171327, 82171982, and 82371466), the Stroke Prevention and Treatment Project of the National Health Commission–Research and Popularization of Appropriate Intervention Technology for the Stroke High Risk Group in China (GN2018R002), the Fujian Science and Technology Innovation Joint Fund Project (2019Y9118), the Technology Platform Construction Project of Fujian Province (2020Y2003 and 2021Y2001), the Major Scientific Research Specialty Project of Fujian Province (2022ZD01003), and the Natural Science Foundation of Fujian Province (2023J01590 and 2023J06031). The funders had no role in the design and conduct of the study; collection, management, analysis, and interpretation of the data; preparation, review, or approval of the article; and decision to submit the article for publication., (Copyright © 2023 The Author(s). Published by Wolters Kluwer Health, Inc.)

Tumor-associated hydrocephalus (TAH) is a common and lethal complication of brain metastases. Although other factors beyond mechanical obstructions have been suggested, the exact mechanisms are unknown. Using single-nucleus RNA sequencing and spatial transcriptomics, we find that a distinct population of mast cells locate in the choroid plexus and dramatically increase during TAH. Genetic fate tracing and intracranial mast-cell-specific tryptase knockout showed that choroid plexus mast cells (CPMCs) disrupt cilia of choroid plexus epithelia via the tryptase-PAR2-FoxJ1 pathway and consequently increase cerebrospinal fluid production. Mast cells are also found in the human choroid plexus. Levels of tryptase in cerebrospinal fluid are closely associated with clinical severity of TAH. BMS-262084, an inhibitor of tryptase, can cross the blood-brain barrier, inhibit TAH in vivo, and alleviate mast-cell-induced damage of epithelial cilia in a human pluripotent stem-cell-derived choroid plexus organoid model. Collectively, we uncover the function of CPMCs and provide an attractive therapy for TAH., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2023 Elsevier Inc. All rights reserved.)

The choroid plexus is a small monolayered epithelium located in the brain ventricles and serves to secrete the cerebrospinal fluid (CSF) that envelops the brain and fills the central ventricles. The CSF secretion is sustained with a concerted effort of a range of membrane transporters located in a polarized fashion in this tissue. Prominent amongst these are the Na + /K + -ATPase, the Na + ,K + ,2Cl - cotransporter (NKCC1), and several HCO 3 - transporters, which together support the net transepithelial transport of the major electrolytes, Na + and Cl - , and thus drive the CSF secretion. The choroid plexus, in addition, serves an important role in keeping the CSF K + concentration at a level compatible with normal brain function. The choroid plexus Na + /K + -ATPase represents a key factor in the barrier-mediated control of the CSF K + homeostasis, as it increases its K + uptake activity when faced with elevated extracellular K + ([K + ] o ). In certain developmental or pathological conditions, the NKCC1 may revert its net transport direction to contribute to CSF K + homeostasis. The choroid plexus ion transport machinery thus serves dual, yet interconnected, functions with its contribution to electrolyte and fluid secretion in combination with its control of brain K + levels., Competing Interests: Declaration of Competing Interest None., (Copyright © 2023 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Background: The choroid plexus (CP) is the principal source of cerebrospinal fluid (CSF). It can produce and release a wide range of materials, including growth and neurotrophic factors which have a crucial role in the maintenance and proper functioning of the brain. Tramadol is a synthetic analog of codeine, mainly prescribed to alleviate mild to moderate pains. Nevertheless, it causes several side effects, such as emotional instability and anxiety., Methods: In this study, we focused on alterations in the expression of inflammatory and apoptotic genes in the CP under chronic tramadol exposure. Herein, rats were treated daily with tramadol at 50 mg/kg doses for three weeks. CSF samples were collected, with superoxide dismutase (SOD) and glutathione (GSH) measured in the CSF., Results: We found that tramadol reduced the SOD and GSH levels in the CSF. Furthermore, the stereological analysis revealed a significant increase in the CP volume, epithelial cells, and capillary number upon tramadol administration. Tramadol elevated the number of blob mitochondria in CP. Also, we observed the upregulation of inflammatory and apoptosis genes following tramadol administration in the CP., Conclusions: Our findings indicate that tramadol induces neurotoxicity in the CP via apoptosis, inflammation, and oxidative stress., (© 2023. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Background: The choroid plexus (CP) is involved in the clearance of harmful metabolites from the brain, as a part of the glymphatic system. This study aimed to investigate the association between CP volume (CPV), nigrostriatal dopaminergic degeneration and motor outcomes in Parkinson's disease (PD)., Methods: We retrospectively searched drug-naïve patients with early-stage PD who underwent dopamine transporter (DAT) scanning and MRI. Automatic CP segmentation was performed, and the CPV was calculated. The relationship between CPV, DAT availability and Unified PD Rating Scale Part III (UPDRS-III) scores was assessed using multivariate linear regression. We performed longitudinal analyses to assess motor outcomes according to CPV., Results: CPV was negatively associated with DAT availability in each striatal subregion (anterior caudate, β=-0.134, p=0.012; posterior caudate, β=-0.162, p=0.002; anterior putamen, β=-0.133, p=0.024; posterior putamen, β=-0.125, p=0.039; ventral putamen, β=-0.125, p=0.035), except for the ventral striatum. CPV was positively associated with the UPDRS-III score even after adjusting for DAT availability in the posterior putamen (β=0.121; p=0.035). A larger CPV was associated with the future development of freezing of gait in the Cox regression model (HR 1.539, p=0.027) and a more rapid increase in dopaminergic medication in the linear mixed model (CPV×time, p=0.037), but was not associated with the risk of developing levodopa-induced dyskinesia or wearing off., Conclusion: These findings suggest that CPV has the potential to serve as a biomarker for baseline and longitudinal motor disabilities in PD., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2023. No commercial re-use. See rights and permissions. Published by BMJ.)

The choroid plexus (CP) plays a key role in remotely controlling brain function in health, aging, and disease. Here, we report that CP epithelial cells express the brain-specific cholesterol 24-hydroxylase (CYP46A1) and that its levels are decreased under different mouse and human brain conditions, including amyloidosis, aging, and SARS-CoV-2 infection. Using primary mouse CP cell cultures, we demonstrate that the enzymatic product of CYP46A1, 24(S)-hydroxycholesterol, downregulates inflammatory transcriptomic signatures within the CP, found here to be elevated across multiple neurological conditions. In vitro, the pro-inflammatory cytokine tumor necrosis factor α (TNF-α) downregulates CYP46A1 expression, while overexpression of CYP46A1 or its pharmacological activation in mouse CP organ cultures increases resilience to TNF-α. In vivo, overexpression of CYP46A1 in the CP in transgenic mice with amyloidosis is associated with better cognitive performance and decreased brain inflammation. Our findings suggest that CYP46A1 expression in the CP impacts the role of this niche as a guardian of brain immune homeostasis., Competing Interests: Declaration of interests M.S. is an inventor of the intellectual property that forms the basis for the development of PD-L1 immunotherapy for AD., (Copyright © 2023 The Author(s). Published by Elsevier Inc. All rights reserved.)

Background: Female sex is a known risk factor of brain disorders with raised intracranial pressure (ICP) and sex hormones have been suggested to alter cerebrospinal fluid (CSF) dynamics, thus impairing ICP regulation in CSF disorders such as idiopathic intracranial hypertension (IIH). The choroid plexus (CP) is the tissue producing CSF and it has been hypothesized that altered hormonal composition could affect the activity of transporters involved in CSF secretion, thus affecting ICP. Therefore, we aimed to investigate if expression of various transporters involved in CSF secretion at CP were different between males and females and between females in different estrous cycle states. Steroid levels in serum was also investigated., Methods: Female and male rats were used to determine sex-differences in the genes encoding for the transporters Aqp1 and 4, NKCC1, NBCe2, NCBE; carbonic anhydrase enzymes II and III (CA), subunits of the Na + /K + -ATPase including Atp1a1, Atp1b1 and Fxyd1 at CP. The estrous cycle stage metestrus (MET) and estrous (ES) were determined before euthanasia. Serum and CP were collected and subjected to RT-qPCR analysis and western blots. Serum was used to measure steroid levels using liquid chromatography tandem mass spectrometry (LC-MS/MS)., Results: Significant differences in gene expression and steroid levels between males and ES females were found, while no differences were found between male and MET females. During ES, expression of Aqp1 was lower (p < 0.01) and NKCC1 was higher in females compared to males. CAII was lower while CAIII was higher in ES females (p < 0.0001). Gene expression of Atp1a1 was lower in ES compared to male (p = 0.0008). Several of these choroidal genes were also significantly different in MET compared to females in ES. Differences in gene expression during the estrus cycle were correlated to serum level of steroid hormones. Protein expression of AQP1 (p = 0.008) and CAII (p = 0.035) was reduced in ES females compared to males., Conclusions: This study demonstrates for the first time that expression at CP is sex-dependent and markedly affected by the estrous cycle in female rats. Further, expression was related to hormone levels in serum. This opens a completely new avenue for steroid regulation of the expression of CSF transporters and the close link to the understanding of CSF disorders such as IIH., (© 2023. The Author(s).)

Background: As a consequence of SARS-CoV-2 infection various neurocognitive and neuropsychiatric symptoms can appear, which may persist for several months post infection. However, cell type-specific routes of brain infection and underlying mechanisms resulting in neuroglial dysfunction are not well understood., Methods: Here, we investigated the susceptibility of cells constituting the blood-brain barrier (BBB) and the blood-cerebrospinal fluid barrier (BCSFB) of the choroid plexus (ChP) to SARS-CoV-2 infection using human induced pluripotent stem cell (hiPSC)-derived cellular models and a ChP papilloma-derived epithelial cell line as well as ChP tissue from COVID-19 patients, respectively., Results: We noted a differential infectibility of hiPSC-derived brain microvascular endothelial cells (BMECs) depending on the differentiation method. Extended endothelial culture method (EECM)-BMECs characterized by a complete set of endothelial markers, good barrier properties and a mature immune phenotype were refractory to SARS-CoV-2 infection and did not exhibit an activated phenotype after prolonged SARS-CoV-2 inoculation. In contrast, defined medium method (DMM)-BMECs, characterized by a mixed endothelial and epithelial phenotype and excellent barrier properties were productively infected by SARS-CoV-2 in an ACE2-dependent manner. hiPSC-derived brain pericyte-like cells (BPLCs) lacking ACE2 expression were not susceptible to SARS-CoV-2 infection. Furthermore, the human choroid plexus papilloma-derived epithelial cell line HIBCPP, modeling the BCSFB was productively infected by SARS-CoV-2 preferentially from the basolateral side, facing the blood compartment. Assessment of ChP tissue from COVID-19 patients by RNA in situ hybridization revealed SARS-CoV-2 transcripts in ChP epithelial and ChP stromal cells., Conclusions: Our study shows that the BCSFB of the ChP rather than the BBB is susceptible to direct SARS-CoV-2 infection. Thus, neuropsychiatric symptoms because of COVID-19 may rather be associated with dysfunction of the BCSFB than the BBB. Future studies should consider a role of the ChP in underlying neuropsychiatric symptoms following SARS-CoV-2 infection., (© 2023. BioMed Central Ltd., part of Springer Nature.)

Intraventricular hemorrhage is a potentially life-threatening condition. Approximately 20% of patients develop posthemorrhagic hydrocephalus with increased ventricular volume and intracranial pressure. Hydrocephalus develops partially due to increased secretion of cerebrospinal fluid by the choroid plexus. During hemorrhage a multitude of factors are released into the cerebrospinal fluid. Many of these have been implicated in the hypersecretion. In this study, we have investigated the isolated effect of inflammatory components, on the abundance of two membrane transporters involved in cerebrospinal fluid secretion by the choroid plexus: the Na + -dependent Cl - /HCO 3 - exchanger, Ncbe, and the Na + , K + , 2Cl - cotransporter, NKCC1. We have established a primary choroid plexus epithelial cell culture from 1 to 7 days old mouse pups. Seven days after seeding, the cells formed a monolayer. The cells were treated with either tumor necrosis factor alpha (TNFα), interleukin 1 beta (IL-1β), or interleukin 6 (IL-6) to mimic inflammation. The data show that treatment with TNFα, and IL-1β only transiently increased NKCC1 abundance whereas the effect on Ncbe abundance was a transient decrease. IL-6 however significantly increased NKCC1 (242%), the phosphorylated NKCC1 (147%), as well as pSPAK (406%) abundance, but had no effect on Ncbe. This study suggests that the inflammatory pathway involved in hypersecretion primarily is mediated by activation of basolateral receptors in the choroid plexus, mainly facilitated by IL-6. This study highlights the complexity of the pathophysiological circumstances occurring during intraventricular hemorrhage., (© 2023. BioMed Central Ltd., part of Springer Nature.)

Myotonic dystrophy type 1 is a dominantly inherited multisystemic disease caused by CTG tandem repeat expansions in the DMPK 3' untranslated region. These expanded repeats are transcribed and produce toxic CUG RNAs that sequester and inhibit activities of the MBNL family of developmental RNA processing factors. Although myotonic dystrophy is classified as a muscular dystrophy, the brain is also severely affected by an unusual cohort of symptoms, including hypersomnia, executive dysfunction, as well as early onsets of tau/MAPT pathology and cerebral atrophy. To address the molecular and cellular events that lead to these pathological outcomes, we recently generated a mouse Dmpk CTG expansion knock-in model and identified choroid plexus epithelial cells as particularly affected by the expression of toxic CUG expansion RNAs. To determine if toxic CUG RNAs perturb choroid plexus functions, alternative splicing analysis was performed on lateral and hindbrain choroid plexi from Dmpk CTG knock-in mice. Choroid plexus transcriptome-wide changes were evaluated in Mbnl2 knockout mice, a developmental-onset model of myotonic dystrophy brain dysfunction. To determine if transcriptome changes also occurred in the human disease, we obtained post-mortem choroid plexus for RNA-seq from neurologically unaffected (two females, three males; ages 50-70 years) and myotonic dystrophy type 1 (one female, three males; ages 50-70 years) donors. To test that choroid plexus transcriptome alterations resulted in altered CSF composition, we obtained CSF via lumbar puncture from patients with myotonic dystrophy type 1 (five females, five males; ages 35-55 years) and non-myotonic dystrophy patients (three females, four males; ages 26-51 years), and western blot and osmolarity analyses were used to test CSF alterations predicted by choroid plexus transcriptome analysis. We determined that CUG RNA induced toxicity was more robust in the lateral choroid plexus of Dmpk CTG knock-in mice due to comparatively higher Dmpk and lower Mbnl RNA levels. Impaired transitions to adult splicing patterns during choroid plexus development were identified in Mbnl2 knockout mice, including mis-splicing previously found in Dmpk CTG knock-in mice. Whole transcriptome analysis of myotonic dystrophy type 1 choroid plexus revealed disease-associated RNA expression and mis-splicing events. Based on these RNA changes, predicted alterations in ion homeostasis, secretory output and CSF composition were confirmed by analysis of myotonic dystrophy type 1 CSF. Our results implicate choroid plexus spliceopathy and concomitant alterations in CSF homeostasis as an unappreciated contributor to myotonic dystrophy type 1 CNS pathogenesis., (© The Author(s) 2023. Published by Oxford University Press on behalf of the Guarantors of Brain. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Posthemorrhagic hydrocephalus occurs in up to 30% of infants with high-grade intraventricular hemorrhage and is associated with the worst neurocognitive outcomes in preterm infants. The mechanisms of posthemorrhagic hydrocephalus after intraventricular hemorrhage are unknown; however, CSF levels of iron metabolic pathway proteins including hemoglobin have been implicated in its pathogenesis. Here, we develop an animal model of intraventricular hemorrhage using intraventricular injection of hemoglobin at post-natal day 4 that results in acute and chronic hydrocephalus, pathologic choroid plexus iron accumulation, and subsequent choroid plexus injury at post-natal days 5, 7, and 15. This model also results in increased expression of aquaporin-1, Na + /K + /Cl - cotransporter 1, and Na + /K + /ATPase on the apical surface of the choroid plexus 24 h post-intraventricular hemorrhage. We use this model to evaluate a clinically relevant treatment strategy for the prevention of neurological sequelae after intraventricular hemorrhage using intraventricular administration of the iron chelator deferoxamine at the time of hemorrhage. Deferoxamine treatment prevented posthemorrhagic hydrocephalus for up to 11 days after intraventricular hemorrhage and prevented the development of sensorimotor gating deficits. In addition, deferoxamine treatment facilitated acute iron clearance through the choroid plexus and subsequently reduced choroid plexus iron levels at 24 h with reversal of hemoglobin-induced aquaporin-1 upregulation on the apical surface of the choroid plexus. Intraventricular administration of deferoxamine at the time of intraventricular hemorrhage may be a clinically relevant treatment strategy for preventing posthemorrhagic hydrocephalus and likely acts through promoting iron clearance through the choroid plexus to prevent hemoglobin-induced injury., (© 2022. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

The choroid plexus (CP) is a source of trophic factors for the developing and mature brain. Insulin is produced in epithelial cells of the CP (EChPs), and its secretion is stimulated by Htr2c-mediated signaling. We modulated insulin expression in EChPs with intracerebroventricular injections of AAV5. Insulin overexpression in CP decelerates food intake, whereas its knockdown has the opposite effect. Insulin overexpression also results in reduced anxious behavior. Transcriptomic changes in the hypothalamus, especially in synapse-related processes, are also seen in mice overexpressing insulin in CP. Last, activation of Gq signaling in CP leads to acute Akt phosphorylation in neurons of the arcuate nucleus, indicating a direct action of CP-derived insulin on the hypothalamus. Taken together, our findings signify that CP is a relevant source of insulin in the central nervous system and that CP-derived insulin should be taken into consideration in future work pertaining to insulin actions in the brain., Competing Interests: Declaration of interests The authors declare no competing interests., (Published by Elsevier Inc.)

Insulin-like growth factor-1 (IGF-1) is essential for normal brain development and regulates essential processes of vascular maturation and stabilization. Importantly, preterm birth is associated with reduced serum levels of IGF-1 as compared to in utero levels. Using a preterm rabbit pup model, we investigated the uptake of systemic recombinant human (rh) IGF-1 in complex with its main binding protein IGF-binding protein 3 (BP-3) to the brain parenchyma via the choroid plexus. Five hours after subcutaneous administration, labeled rhIGF-1/rhIGFBP-3 displayed a widespread presence in the choroid plexus of the lateral and third ventricle, however, to a less degree in the fourth, as well as in the perivascular and subarachnoid space. We found a time-dependent uptake of IGF-1 in cerebrospinal fluid, decreasing with postnatal age, and a translocation of IGF-1 through the choroid plexus. The impact of systemic rhIGF-1/rhIGFBP-3 on IGF-1 receptor activation in the choroid plexus decreased with postnatal age, correlating with IGF-1 uptake in cerebrospinal fluid. In addition, choroid plexus gene expression was observed to increase with postnatal age. Moreover, using choroid plexus in vitro cell cultures, gene expression and protein synthesis were further investigated upon rhIGF-1/rhIGFBP-3 stimulation as compared to rhIGF-1 alone, and found not to be differently altered. Here, we characterize the uptake of systemic rhIGF-1/rhIGFBP-3 to the preterm brain, and show that the interaction between systemic rhIGF-1/rhIGFBP-3 and choroid plexus varies over time., (© 2023. BioMed Central Ltd., part of Springer Nature.)

The choroid plexus (ChP) produces and is bathed in the cerebrospinal fluid (CSF), which in aging and Alzheimer's disease (AD) shows extensive proteomic alterations including evidence of inflammation. Considering inflammation hampers functions of the involved tissues, the CSF abnormalities reported in these conditions are suggestive of ChP injury. Indeed, several studies document ChP damage in aging and AD, which nevertheless remains to be systematically characterized. We here report that the changes elicited in the CSF by AD are consistent with a perturbed aging process and accompanied by aberrant accumulation of inflammatory signals and metabolically active proteins in the ChP. Magnetic resonance imaging (MRI) imaging shows that these molecular aberrancies correspond to significant remodeling of ChP in AD, which correlates with aging and cognitive decline. Collectively, our preliminary post-mortem and in vivo findings reveal a repertoire of ChP pathologies indicative of its dysfunction and involvement in the pathogenesis of AD. HIGHLIGHTS: Cerebrospinal fluid changes associated with aging are perturbed in Alzheimer's disease Paradoxically, in Alzheimer's disease, the choroid plexus exhibits increased cytokine levels without evidence of inflammatory activation or infiltrates In Alzheimer's disease, increased choroid plexus volumes correlate with age and cognitive performance., (© 2023 The Authors. Alzheimer's & Dementia published by Wiley Periodicals LLC on behalf of Alzheimer's Association.)

Background: The choroid plexus (CP) produces and secretes most of the cerebrospinal fluid (CSF) of the central nervous system. The CP is suggested to be regulated by descending neurons and by circulating factors and is involved in the interaction between central and peripheral inflammation. Quantitative imaging has demonstrated volumetric CP changes in psychosis, schizophrenia and depression. This study histologically examines CP epithelial cell morphology in these illnesses to identify the biological source of such volumetric changes., Methods: Formalin-fixed paraffin-embedded (FFPE) blocks were obtained bilaterally from the lateral ventricles of 13 cases of sex- and age-matched brains from each of schizophrenia (SZ) with psychosis, major depressive disorder (MDD) and matched controls (NPD). FFPE blocks were sectioned at 7 μm and routinely stained for H&E. Morphological analysis of 180 CP epithelia/case was conducted blindly on digital images collected at x600 magnification. Calcification was assessed in all CP regions manually., Results: Analysis with a General Linear Model demonstrated a significant effect of diagnosis on somal width (p = 0.006, R 2  = 0.33 R 2 (adj) = 0.25) demonstrating increased somal width in SZ without psychotic medication versus controls (p = 0.032), but not in medicated SZ cases. No effects were observed in calcification., Discussion: The epithelial cells that were examined were attached to the CP fibrous surface, so width expansion describes the primary methods for these cells to expand with adherence to this surface in SZ. The interaction of antipsychotic medication and diagnosis demonstrates that this is an illness-specific change mediated through the DA-system with likely neuronal origin. CP alterations were not found in MDD where they are instead generally associated with heightened allostatic load that was unknown in this cohort., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Author(s). Published by Elsevier Inc. All rights reserved.)

Transmission and secretion of signals via the choroid plexus (ChP) brain barrier can modulate brain states via regulation of cerebrospinal fluid (CSF) composition. Here, we developed a platform to analyze diurnal variations in male mouse ChP and CSF. Ribosome profiling of ChP epithelial cells revealed diurnal translatome differences in metabolic machinery, secreted proteins, and barrier components. Using ChP and CSF metabolomics and blood-CSF barrier analyses, we observed diurnal changes in metabolites and cellular junctions. We then focused on transthyretin (TTR), a diurnally regulated thyroid hormone chaperone secreted by the ChP. Diurnal variation in ChP TTR depended on Bmal1 clock gene expression. We achieved real-time tracking of CSF-TTR in awake Ttr mNeonGreen mice via multi-day intracerebroventricular fiber photometry. Diurnal changes in ChP and CSF TTR levels correlated with CSF thyroid hormone levels. These datasets highlight an integrated platform for investigating diurnal control of brain states by the ChP and CSF., (© 2023. The Author(s).)

The choroid plexus (ChP) serves as a critical gateway for immune cell infiltration into the central nervous system (CNS) under both physiological and pathological conditions. Recent research has shown that regulating ChP activity may offer protection against CNS disorders. However, studying the biological function of the ChP without affecting other brain regions is challenging due to its delicate structure. This study presents a novel method for gene knockdown in ChP tissue using adeno-associated viruses (AAVs) or cyclization recombination enzyme (Cre) recombinase protein consisting of TAT sequence (CRE-TAT). The results demonstrate that after injecting AAV or CRE-TAT into the lateral ventricle, the fluorescence was exclusively concentrated in the ChP. Using this approach, the study successfully knocked down the adenosine A2A receptor (A2AR) in the ChP using RNA interference (RNAi) or Cre/locus of X-overP1 (Cre/LoxP) systems, and showed that this knockdown could alleviate the pathology of experimental autoimmune encephalomyelitis (EAE). This technique may have important implications for future research on the ChP's role in CNS disorders.

Regulation of the volume and electrolyte composition of the cerebrospinal fluid (CSF) is vital for brain development and function. The Na-K-Cl co-transporter NKCC1 in the choroid plexus (ChP) plays key roles in regulating CSF volume by co-transporting ions and mediating same-direction water movements. Our previous study showed ChP NKCC1 is highly phosphorylated in neonatal mice as the CSF K + level drastically decreases and that overexpression of NKCC1 in the ChP accelerates CSF K + clearance and reduces ventricle size [1]. These data suggest that NKCC1 mediates CSF K + clearance following birth in mice. In this current study, we used CRISPR technology to create a conditional NKCC1 knockout mouse line and evaluated CSF K + by Inductively Coupled Plasma Optical Emission spectroscopy (ICP-OES). We demonstrated ChP-specific reduction of total and phosphorylated NKCC1 in neonatal mice following embryonic intraventricular delivery of Cre recombinase using AAV2/5. ChP-NKCC1 knockdown was accompanied by a delayed perinatal clearance of CSF K + . No gross morphological disruptions were observed in the cerebral cortex. We extended our previous results by showing embryonic and perinatal rats shared key characteristics with mice, including decreased ChP NKCC1 expression level, increased ChP NKCC1 phosphorylation state, and increased CSF K + levels compared to adult. Collectively, these follow up data support ChP NKCC1's role in age-appropriate CSF K + clearance during neonatal development., (© 2023. The Author(s).)