Rat Choroid Plexus Research Articles

Progesterone Enhances Transthyretin Expression in the Rat Choroid Plexus In Vitro and In Vivo via Progesterone Receptor

Depletion of ovarian hormones 17β-estradiol (E2) and progesterone (P) after menopause may contribute to the decline in cognitive performance and increases the risk of Alzheimer's disease (AD) in women, striking the importance of understanding the regulation of pivotal proteins involved in AD pathogenesis by ovarian hormones. Transthyretin (TTR) is now recognized as one of such proteins due to its ability to sequester and degrade amyloid β (Aβ) into less harmful peptides and preventing their aggregation. We have previously demonstrated that E2 enhances TTR expression. In this study, we investigate the effects of P on TTR expression in primary cultures of rat choroid plexus epithelial cells and in adult ovariectomized female rats. The results obtained demonstrate that, in vitro and in vivo, TTR is up-regulated by P. In addition, the mechanism underlying the response of TTR to P was investigated, and we provide evidence that this response is achieved through a progesterone receptor-mediated mechanism. Our results reinforce the importance of ovarian hormones on the regulation of TTR, which may reflect on the processing of Aβ peptides and consequently on AD onset and progression.

Lead-induced accumulation of β-amyloid in the choroid plexus: Role of low density lipoprotein receptor protein-1 and protein kinase C

The choroid plexus (CP), constituting the blood–cerebrospinal fluid barrier, has the capacity to remove beta-amyloid (Aβ) from the cerebrospinal fluid. Our previous work indicates that exposure to lead (Pb) results in Aβ accumulation in the CP by decreasing the expression of low density lipoprotein receptor protein-1 (LRP1), a protein involved in the transport and clearance of Aβ. The current study was designed to explore the relationship between Aβ accumulation, protein kinase C (PKC) activity, and LRP1 status in the CP following Pb exposure. Confocal microscopy revealed that LRP1 was primarily localized in the cytosol of the CP in control rats and migrated distinctly towards the apical surface and the microvilli following acute Pb exposure (27 mg Pb/kg, i.p., 24 h). Co-immunostaining revealed a co-localization of both PKC-δ and LRP1 in the cytosol of control rats, with a distinct relocalization of both towards the apical membrane following Pb exposure. Preincubation of the tissues with PKC-δ inhibitor rottlerin (2 μM) prior to Pb exposure in vitro, resulted in abolishing the Pb-induced relocalization of LRP1 to the apical surface. Importantly, a significant elevation in intracellular Aβ levels ( p < 0.01) was observed in the cytosol of the CP following Pb exposure, which was abolished following preincubation with rottlerin. In addition, rottlerin caused a relocalization of Aβ from the cytosol to the nucleus in both Pb-treated and control CP tissues. Finally, co-immunoprecipitation studies revealed a strong protein-protein interaction between LRP1 and PKC-δ in the CP. These studies suggest that Pb exposure disrupts Aβ homeostasis at the CP, owing partly to a Pb-induced relocalization of LRP1 via PKC-δ.

Efflux mechanisms at the developing brain barriers: ABC-transporters in the fetal and postnatal rat

Proteins of the ATP-binding cassette (ABC) family, present at the blood–brain barrier interfaces, have been shown to reduce the entry of compounds from blood into the brain by active efflux. Their substrates are diverse including many drugs and toxins and therefore provide an important mechanism for brain neuroprotection. However, knowledge of their presence and function in the developing brain is very limited. We have used qPCR and immunocytochemistry to determine gene expression and localisation of four main barrier ABC-transporters (pgp/ABCB1, MRP1/ABCC1, MRP4/ABCC4 and BCRP/ABCG2) in the fetal and neonatal rat brain cerebral blood vessels (site of blood–brain barrier) and choroid plexus (site of blood–CSF barrier). The study shows that ABC-transporters localise to the brain barriers even at early fetal stages and although pgp expression was lower in the fetus, the other transporters were expressed at comparable levels in fetal and adult brains suggesting direct neuroprotection of the brain in addition to that provided by the placenta. BCRP was expressed at higher levels in developing choroid plexus and was only detected at fetal stages on the blood-facing side of epithelial cells indicating a particular role of this transporter for early brain efflux mechanisms.

In-vivo and in-vitro evidence of a carrier-mediated efflux transport system for oestrone-3-sulphate across the blood-cerebrospinal fluid barrier.

The efflux transport of oestrone-3-sulphate, a steroid hormone sulphate, across the blood-cerebrospinal fluid barrier has been examined following its intracerebroventricular administration. [3H]Oestrone-3-sulphate was eliminated from cerebrospinal fluid (CSF) with an apparent efflux clearance of 205 microL min(-1) per rat. There was 25% of unmetabolized [3H]oestrone-3-sulphate in the plasma 5 min after intracerebroventricular administration, indicating that at least a part of [3H]oestrone-3-sulphate is transported from CSF to the circulating blood across the blood-CSF barrier. This efflux transport was inhibited by co-administration of excess oestrone-3-sulphate (25 mM 10 microL = 0.25 micromol) into rat cerebral ventricle. To characterize the oestrone-3-sulphate transport process, an in-vitro uptake experiment was performed using isolated rat choroid plexus. Oestrone-3-sulphate uptake by isolated rat choroid plexus was found to be a saturable process with a Michaelis-Menten constant (Km) of 18.1 +/- 6.3 microM, and a maximum uptake rate (Vmax) of 48.0 +/- 15.1 pmol min(-1) microL(-1) of tissue. The oestrone-3-sulphate transport process was temperature dependent and was inhibited by metabolic inhibitors such as 2,4-dinitrophenol and rotenone, suggesting an energy dependence. This uptake process was also inhibited by steroid hormone sulphates (1 mM dehydroepiandrosterone sulphate and 1 mM oestrone sulphate), bile acids (1 mM taurocholic acid and 1 mM cholic acid) and organic anions (1 mM sulphobromophthalein and 1 mM phenolsulphonphthalein), whereas 1 mM p-aminohippuric acid, 1 mM p-nitrophenol sulphate, 0.1 mM methotrexate and the cardiac glycoside, 2.5 microM digoxin, had little effect. In conclusion, these results provide evidence that oestrone-3-sulphate is transported from CSF to the circulating blood across the blood-CSF barrier via a carrier-mediated efflux transport system.

Prenatal and Postnatal Development of the Rat Choroid Plexus

Prenatal and Postnatal Development of the Rat Choroid Plexus

Platelet-Derived Growth Factor Receptor Expression and Activation in Choroid Plexus Tumors

Choroid plexus tumors are intraventricular neoplasms predominantly affecting young children. In contrast to choroid plexus papillomas, choroid plexus carcinomas progress frequently, necessitating the development of adjuvant treatment concepts. Platelet derived growth factor (PDGF) signaling has been shown to support growth in a variety of tumors. The finding of PDGF receptor expression in choroid plexus tumors prompted us to elucidate PDGF receptor activation state using a novel method, in situ proximity ligation assay, on formalin-fixed, paraffin-embedded, archival samples of 19 choroid plexus tumors. As assessed by in situ proximity ligation assay, the proportion of phosphorylated PDGF receptor alpha was low in choroid plexus papillomas and choroid plexus carcinomas, whereas phosphorylated PDGF receptor beta was found to be significantly higher in choroid plexus carcinomas. In the immortalized choroid plexus epithelial cell line Z310 expressing PDGF receptor beta, PDGF-BB exhibited a time- and dose-dependent proliferative response, which was significantly attenuated by imatinib (gleevec). In conclusion, our findings suggest that PDGF receptor beta is functionally involved in the biology of choroid plexus tumors and may represent a molecular target for therapy. In addition, this study demonstrates the feasibility and usefulness of in situ proximity ligation assay for monitoring receptor tyrosine kinase activation in formalin-fixed, paraffin-embedded, archival tissues.

Regulation of cerebrospinal fluid production by caffeine consumption

BackgroundCaffeine is the most commonly consumed psycho-stimulant in the world. The effects of caffeine on the body have been extensively studied; however, its effect on the structure of the brain has not been investigated to date.ResultsIn the present study we found that the long-term consumption of caffeine can induce ventriculomegaly; this was observed in 40% of the study rats. In the caffeine-treated rats with ventriculomegaly, there was increased production of CSF, associated with the increased expression of Na+, K+-ATPase and increased cerebral blood flow (CBF). In contrast to the chronic effects, acute treatment with caffeine decreased the production of CSF, suggesting 'effect inversion' associated with caffeine, which was mediated by increased expression of the A1 adenosine receptor, in the choroid plexus of rats chronically treated with caffeine. The involvement of the A1 adenosine receptor in the effect inversion of caffeine was further supported by the induction of ventriculomegaly and Na+, K+-ATPase, in A1 agonist-treated rats.ConclusionThe results of this study show that long-term consumption of caffeine can induce ventriculomegaly, which is mediated in part by increased production of CSF. Moreover, we also showed that adenosine receptor signaling can regulate the production of CSF by controlling the expression of Na+, K+-ATPase and CBF.

Folate Receptor Alpha Defect Causes Cerebral Folate Transport Deficiency: A Treatable Neurodegenerative Disorder Associated with Disturbed Myelin Metabolism

Sufficient folate supplementation is essential for a multitude of biological processes and diverse organ systems. At least five distinct inherited disorders of folate transport and metabolism are presently known, all of which cause systemic folate deficiency. We identified an inherited brain-specific folate transport defect that is caused by mutations in the folate receptor 1 (FOLR1) gene coding for folate receptor alpha (FRalpha). Three patients carrying FOLR1 mutations developed progressive movement disturbance, psychomotor decline, and epilepsy and showed severely reduced folate concentrations in the cerebrospinal fluid (CSF). Brain magnetic resonance imaging (MRI) demonstrated profound hypomyelination, and MR-based in vivo metabolite analysis indicated a combined depletion of white-matter choline and inositol. Retroviral transfection of patient cells with either FRalpha or FRbeta could rescue folate binding. Furthermore, CSF folate concentrations, as well as glial choline and inositol depletion, were restored by folinic acid therapy and preceded clinical improvements. Our studies not only characterize a previously unknown and treatable disorder of early childhood, but also provide new insights into the folate metabolic pathways involved in postnatal myelination and brain development.

Cellular Transplants, 20 Years Later: The Pharma Initiative

Cellular Transplants, 20 Years Later: The Pharma Initiative

Zinc (Zn) Modulates the Stress Response in Cadmium (Cd) Treated Cultured Choroid Plexus

Our objective is to elucidate how an essential mineral Zn might attenuate the cellular stress response elicited by a contaminant metal, e.g., Cd, in choroid plexus (CP). To begin, primary cultures of neonatal rat CP were supplemented ± 25 µM Zn with 5% NuSerum for 48 h then exposed to 500 nM Cd ± 10 µM Zn in serum‐free medium for 24 h. As per immunoblot analysis, Hsp70 and hemeoxygenase‐1 (HO) were expressed at low levels in control cells (no Zn/no Cd) and cells treated only with Zn. Cd increased Hsp70 and HO by 3‐6‐fold, but co‐exposure with Zn attenuated induction of both stress proteins. Hsp70 expression was also imaged by immunochemistry and fluorescence microscopy; DAPI was used for nuclear staining. Diffuse low fluorescence was detected in controls and in cells treated only Zn. Cd increased Hsp70 throughout the cytosol and altered cell morphology, but in cells co‐exposed to Zn Hsp70 expression pattern was similar to that in controls. Actin cytoskeleton was probed with TRITC‐phalloidin. In control cells, actin filaments were localized at the periphery, whereas in cells treated with Cd plus Zn there was an increase in actin filaments throughout the cell. Interestingly, in cells treated only with Zn actin filaments were localized at the periphery as in control. These preliminary data suggest that Zn attenuates induction of stress proteins and modifies changes in the actin cytoskeleton in Cd‐treated CP.

TWIST-1 is overexpressed in neoplastic choroid plexus epithelial cells and promotes proliferation and invasion.

The pathogenesis of choroid plexus papillomas, intraventricular papillary neoplasms most often occurring sporadically in children and young adults, remains poorly understood. To identify pathways operative in the development of choroid plexus papillomas, gene expression profiles obtained from laser-microdissected human choroid plexus papilloma cells (n = 7) were compared with that of normal choroid plexus epithelial cells laser microdissected from autopsy tissue (n = 8). On DNA microarray data analysis, 53 probe sets were differentially expressed in choroid plexus papilloma tumor cells (>7-fold). Up-regulation of TWIST1, WIF1, TRPM3, BCLAF1, and AJAP1, as well as down-regulation of IL6ST was confirmed using quantitative reverse transcription-PCR. Knockdown of Twist1 gene expression in the rat choroid plexus epithelial cell line Z310 significantly reduced proliferation as assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and cell invasion in a Matrigel assay, whereas cell migration was not affected. Screening for expressional changes of cancer-related genes upon Twist1 knockdown revealed up-regulation of Cdkn1a, Cflar, and Serpinb2 and down-regulation of Figf. To conclude, using gene expression profiling, several genes differentially expressed in human choroid plexus papillomas could be identified. Among those, TWIST1 is highly expressed in choroid plexus papillomas and promotes proliferation and invasion.

Altered Iron Metabolism Is Part of the Choroid Plexus Response to Peripheral Inflammation

Iron is essential for normal cellular homeostasis but in excess promotes free radical formation and is detrimental. Therefore, iron metabolism is tightly regulated. Here, we show that mechanisms regulating systemic iron metabolism may also control iron release into the brain at the blood-choroid plexus-cerebrospinal fluid (CSF) barrier. Intraperitoneal administration of lipopolysaccharide (LPS) in mice triggers a transient transcription of the gene encoding for hepcidin, a key regulator of iron homeostasis, in the choroid plexus, which correlated with increased detection of pro-hepcidin in the CSF. Similarly, the expression of several other iron-related genes is influenced in the choroid plexus by the inflammatory stimulus. Using primary cultures of rat choroid plexus epithelial cells, we show that this response is triggered not only directly by LPS but also by molecules whose expression increases in the blood in response to inflammation, such as IL-6. Intracellular conveyors of these signaling molecules include signal transducer and activator of transcription 3, which becomes phosphorylated, and SMAD family member 4, whose mRNA levels increase soon after LPS administration. This novel role for the choroid plexus-CSF barrier in regulating iron metabolism may be particularly relevant to restrict iron availability for microorganism growth, and in neurodegenerative diseases in which an inflammatory underlying component has been reported.

17beta-estradiol induces transthyretin expression in murine choroid plexus via an oestrogen receptor dependent pathway.

Oestrogen protects against AD by multiple mechanisms, including the enhancement of Abeta clearance. Transthyretin (TTR) is a homotetrameric protein mainly synthesized by the liver and choroid plexus (CP) of the brain that sequesters the amyloid beta (Abeta) peptide. In this study we examined the effects of 17beta-estradiol (E2) on TTR protein and mRNA levels, in primary cultures of rat CP epithelial cells (CPEC) by Western blot and Real Time PCR, respectively. Moreover, the localization of oestrogen receptors alpha (ERalpha) and beta (ERbeta) in response to E2 treatment was analysed by confocal microscopy in these cells. The expression of TTR, ERalpha and ERbeta was also compared in the CP of castrated female mice treated with E2 to vehicle-treated animals by Real Time PCR. TTR concentration in the CSF of all these animals was measured by radioimmunoassay. E2 treatment induced TTR transcription and increased TTR protein content in CPEC. Pre-treatment with ICI 182,780 (ICI) abrogated E2-induced TTR expression suggesting that, TTR is up-regulated via an ER-dependent pathway. Confocal microscopy demonstrated extranuclear ERalpha and ERbeta localization in untreated CPEC. Upon E2 treatment, translocation of ERalpha to the nucleus occurred, while ERbeta remained in the cytosol. These data was concurrent with the up-regulation of TTR expression detected in the CP of castrated female mice subjected to E2 treatment. Our results highlight the importance of E2 on the regulation of TTR, which may participate in the oestrogen-induced decrease in Abeta levels and deposition described in the literature.

Expression of HNF4alpha in the human and rat choroid plexus – Implications for drug transport across the blood-cerebrospinal-fluid (CSF) barrier

BackgroundThe choroid plexus consists of highly differentiated epithelium and functions as a barrier at the interface of the blood-cerebrospinal-fluid (CSF). This tissue may therefore determine the bioavailability and transport of drugs to the brain. Little is known about the expression of drug and xenobiotic metabolizing enzymes (DME) and of drug transporters in the human choroid plexus. Notably, the transcription factor and zinc finger protein HNF4alpha is a master regulator of DMEs and of drug transporters. As of today its activity in the blood-CSF barrier is unknown. Here we report our efforts in determining HNF4alpha activity in the regulation of ABC transporters in the human and rat choroid plexus.ResultsWe report expression of HNF4alpha by qRT-PCR and by immunohistochemistry and evidence transcript expression of the ATP-binding cassette transporters ABCB1, ABCB4, ABCC1-6 in choroid plexus. Additionally, HNF4alpha DNA binding activity at regulatory sequences of ABCB4 and ABCC1 was determined by EMSA bandshift assays with a specific antibody. We then performed siRNA mediated functional knock down of HNF4alpha in Caco-2 cells and found ABCC1 gene expression to be repressed in cell culture experiments.ConclusionOur study evidences activity of HNF4alpha in human and rat choroid plexus. This transcription factor targets DMEs and drug transporters and may well determine availability of drugs at the blood-CSF barrier.

Progesterone up-regulates transthyretin levels in primary cultures of choroid plexus epithelial cells

Event Abstract Back to Event Progesterone up-regulates transthyretin levels in primary cultures of choroid plexus epithelial cells Telma Alexandra-Quintela-Paixao1* 1 University of Beira Interior, Centre of Investigation in Health Sciences, Portugal Transthyretin (TTR) is a protein mainly synthesized by the liver and choroid plexus (CP) of the brain. Besides its role as a carrier for thyroid hormones, TTR also sequesters the amyloid beta (Aβ) peptide impairing its deposition in nervous tissues, and its concentrations in the cerebrospinal fluid (CSF) appear to be inversely correlated with Alzheimer´s disease (AD) onset and progression. Abundant evidence suggests that the depletion of progesterone (PROG) at menopause is a significant risk factor for the development of AD in women, but the mechanisms involved are poorly understood. So, we examined the effects of PROG on TTR protein and mRNA levels, in primary cultures of rat CP epithelial cells (CPEC) by Western blot, and Real Time PCR, respectively. We show that, PROG (100 nM) induced TTR transcription in these cells. The combination of mifepristone (RU486) with PROG in the treatment of CPEC abrogated the induction of TTR expression by PROG. Pre-treatment with ICI 182,780 and flutamide, specific oestrogen and androgen receptor antagonists, respectively, had no effect on TTR levels. These data suggest that the effects observed are due to PROG itself and not from downstream products from its metabolization, such as testosterone and estradiol, and indicates that TTR is up-regulated via a progesterone receptor (PR)-dependent pathway. Our results highlight the importance of PROG on the regulation of TTR, which may be involved in the neuroprotective role of PROG in AD described in the literature. Conference: 11th Meeting of the Portuguese Society for Neuroscience, Braga, Portugal, 4 Jun - 6 Jun, 2009. Presentation Type: Poster Presentation Topic: Abstracts Citation: Alexandra-Quintela-Paixao T (2009). Progesterone up-regulates transthyretin levels in primary cultures of choroid plexus epithelial cells. Front. Neurosci. Conference Abstract: 11th Meeting of the Portuguese Society for Neuroscience. doi: 10.3389/conf.neuro.01.2009.11.042 Copyright: The abstracts in this collection have not been subject to any Frontiers peer review or checks, and are not endorsed by Frontiers. They are made available through the Frontiers publishing platform as a service to conference organizers and presenters. The copyright in the individual abstracts is owned by the author of each abstract or his/her employer unless otherwise stated. Each abstract, as well as the collection of abstracts, are published under a Creative Commons CC-BY 4.0 (attribution) licence (https://creativecommons.org/licenses/by/4.0/) and may thus be reproduced, translated, adapted and be the subject of derivative works provided the authors and Frontiers are attributed. For Frontiers’ terms and conditions please see https://www.frontiersin.org/legal/terms-and-conditions. Received: 06 Aug 2009; Published Online: 06 Aug 2009. * Correspondence: Telma Alexandra-Quintela-Paixao, University of Beira Interior, Centre of Investigation in Health Sciences, Covilhã, Portugal, nemoABS01@frontiersin.org Login Required This action requires you to be registered with Frontiers and logged in. To register or login click here. Abstract Info Abstract The Authors in Frontiers Telma Alexandra-Quintela-Paixao Google Telma Alexandra-Quintela-Paixao Google Scholar Telma Alexandra-Quintela-Paixao PubMed Telma Alexandra-Quintela-Paixao Related Article in Frontiers Google Scholar PubMed Abstract Close Back to top Javascript is disabled. Please enable Javascript in your browser settings in order to see all the content on this page.

Drug metabolizing enzyme expression in rat choroid plexus: effects of in vivo xenobiotics treatment

The presence of drug metabolizing enzymes in extrahepatic tissues such as the choroid plexus (CP) suggests that the CP, like the blood-brain barrier, affords a metabolic protection to the brain against xenobiotics. The CP, which is the principal site of formation of the cerebrospinal fluid (CSF), controls the exchange of many endogenous compounds and exogenous molecules between brain tissue and CSF. We present the changes in mRNA expression and enzymatic activities of UDP-glucuronosyltransferase, UGT1A6 isoform and NADPH-cytochrome P450 reductase, after in vitro treatment with xenobiotic molecules known to act in the liver as inducers or inhibitors of these drug metabolizing enzymes. Five study groups of male Sprague-Dawley rats were treated separately with 3-methylcholantrene (3-MC), phenobarbital (PB), dexamethasone (DEX), cyclosporine (CsA) or paraquat (PQ). Choroidal 1-naphthol glucuronidation activities were significantly induced by 3-MC and PQ administration (354 +/- 85 and 257 +/- 49 vs. 115 +/- 24 nmol/h per mg protein, in control group), whereas the other molecules were without effect. Accordingly, UGT1A6 mRNA expression, measured by RT-PCR, was 2.3-fold higher after 3-MC treatment and 2.1-fold higher after PQ administration. By contrast, reductase activities and mRNA expression remained unchanged in the isolated choroids plexus in these experimental conditions. We present for the first time evidences that the choroids plexus express transcripts for both UGT1A6 and NADPH-cytochrome P450 reductase, and their mRNA expression can be differently regulated by exogenous factors. These results emphasize that xenobiotics could modulate the biotransformation of exogenous and/or endogenous compounds in the choroids plexus, and underline the role of UGTs in the maintenance of brain homeostasis.

Androgen Receptor is Expressed in Murine Choroid Plexus and Downregulated by 5α-Dihydrotestosterone in Male and Female Mice

The choroid plexuses (CPs) of the brain form a unique interface between the peripheral blood and the cerebrospinal fluid (CSF). CPs produce several neuroprotective peptides, which are secreted into the CSF. Despite their importance in neuroprotection, the mechanisms underlying the regulation of most of these peptides in CPs remain unknown. Androgens regulate the expression of neuroprotective peptides in several tissues where the androgen receptor (AR) is coexpressed, including the brain. The presence of AR in CPs has never been investigated, but recent studies in our laboratory show that the CP is an androgen-responsive tissue. In order to fulfill this gap, we investigated and characterized AR distribution and expression in male and female rat CPs and in primary cultures from rat CP epithelial cells. In addition, the response of AR to 5alpha-dihydrotestosterone (DHT) in castrated male and female mice subjected to DHT replacement was analyzed. We show that rat CP epithelial cells contain AR mRNA and protein. Moreover, we demonstrate that AR is downregulated by DHT in mice CPs.

Copper transport to the brain by the blood-brain barrier and blood-CSF barrier

The mechanism of copper (Cu) transport into the brain is unclear. This study evaluated the main species and route of Cu transport into the brain using in situ brain perfusion technique, and assessed the levels of mRNA encoding Cu transporters using real time RT-PCR. Free 64Cu uptake in rat choroid plexus (CP), where the blood-cerebrospinal fluid barrier (BCB) is primarily located, is about 50 and 1000 times higher than 64Cu-albumin and 64Cu-ceruloplasmin uptake, respectively. The unidirectional transport rate constants ( K in) for Cu in the CP and brain capillaries of the blood-brain barrier (BBB) were 1034 and 319 μl/s/g, respectively, while K in in CSF and capillary-depleted parenchyma were much reduced, 0.8 and 112 μl/s/g, respectively. The K in in cerebellum was significantly lower than that in hippocampus. The mRNAs encoding Cu transporter-1 (Ctr1) and ATP7A were higher in the CP than those in brain capillaries and parenchyma, whereas ATP7B mRNA was higher in brain capillaries than those in the CP and brain parenchyma. Taken together, these data suggest that the expression of Cu transporters is higher in brain barriers than in brain parenchyma; the Cu transport into the brain is mainly achieved through the BBB as a free Cu ion and the BCB may serve as a main regulatory site of Cu in the CSF.

Expression and possible role of creatine transporter in the brain and at the blood‐cerebrospinal fluid barrier as a transporting protein of guanidinoacetate, an endogenous convulsant

Little is known about the cerebral distribution and clearance of guanidinoacetate (GAA), the accumulation of which induces convulsions. The purpose of the present study was to identify creatine transporter (CRT)-mediated GAA transport and to clarify its cerebral expression and role in GAA efflux transport at the blood-cerebrospinal fluid barrier (BCSFB). CRT mediated GAA transport with a K(m) value of 269 microM/412 microM which was approximately 10-fold greater than that of CRT for creatine. There was wide and distinct cerebral expression of CRT and localization of CRT on the brush-border membrane of choroid plexus epithelial cells. The in vivo elimination clearance of GAA from the CSF was 13-fold greater than that of d-mannitol reflecting bulk flow of the CSF. This process was partially inhibited by creatine. The characteristics of GAA uptake by isolated choroid plexus and an immortalized rat choroid plexus epithelial cell line (TR-CSFB cells) used as an in vitro model of BCSFB are partially consistent with those of CRT. These results suggest that CRT plays a role in the cerebral distribution of GAA and GAA uptake by the choroid plexus. However, in the presence of endogenous creatine in the CSF, CRT may make only a limited contribution to the GAA efflux transport at the BCSFB.

Texas Red transport across rat and dogfish shark (Squalus acanthias) choroid plexus

Confocal microscopy and image analysis were used to compare driving forces, specificity, and regulation of transport of the fluorescent organic anion, Texas Red (sulforhodamine 101 free acid; TR), in lateral choroid plexus (CP) isolated from rat and an evolutionarily ancient vertebrate, dogfish shark (Squalus acanthias). CP from both species exhibited concentrative, specific, and metabolism-dependent TR transport from bath to subepithelial/vascular space; at steady state, TR accumulation in vascular/subepithelial space was substantially higher than in epithelial cells. In rat CP, steady-state TR accumulation in subepithelial/vascular spaces was reduced by Na(+)-replacement, but was not affected by a 10-fold increase in buffer K(+). In shark CP, Na(+)-replacement did not alter TR accumulation in either tissue compartment; subepithelial/vascular space levels of TR were reduced in high-K(+) medium. In both species, steady-state TR accumulation was not affected by p-aminohippurate or leukotriene C4, suggesting that neither organic anion transporters (SLC22A family) nor multidrug resistance-associated proteins (ABCC family) contributed. In rat CP, digoxin was without effect, indicating that organic anion transporting polypeptide isoform 2 was not involved. Several organic anions reduced cellular and subepithelial/vascular space TR accumulation in both tissues, including estrone sulfate, taurocholate, and the Mrp1 inhibitor MK571. In rat CP, TR accumulation in subepithelial/vascular spaces increased with PKA activation (forskolin), but was not affected by PKC activation (phorbol ester). In shark, neither PKA nor PKC activation specifically affected TR transport. Thus, rat and dogfish shark CP transport TR but do so using different basic mechanisms that respond to different regulatory signals.