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Ependymal cells line the wall of cerebral ventricles and ensure the unidirectional cerebrospinal fluid (CSF) flow by beating their motile cilia coordinately. The ependymal denudation or ciliary dysfunction causes hydrocephalus. Here, we report that the deficiency of regulator of G-protein signaling 22 (RGS22) results in severe congenital hydrocephalus in both mice and rats. Interestingly, RGS22 is specifically expressed in ependymal cells within the brain. Using conditional knock-out mice, we further demonstrate that the deletion of Rgs22 exclusively in nervous system is sufficient to induce hydrocephalus. Mechanistically, we show that Rgs22 deficiency leads to the ependymal denudation and impaired ciliogenesis. This phenomenon can be attributed to the excessive activation of lysophosphatidic acid receptor (LPAR) signaling under Rgs22-/- condition, as the LPAR blockade effectively alleviates hydrocephalus in Rgs22-/- rats. Therefore, our findings unveil a previously unrecognized role of RGS22 in the central nervous system, and present RGS22 as a potential diagnostic and therapeutic target for hydrocephalus. [ABSTRACT FROM AUTHOR]

Abstract Hydrocephalus is a common neurological condition, characterized by the excessive accumulation of cerebrospinal fluid in the cerebral ventricles. Primary treatments for hydrocephalus mainly involve neurosurgical cerebrospinal fluid diversion, which hold high morbidity and failure rates, highlighting the necessity for the discovery of novel therapeutic approaches. Although the pathophysiology of hydrocephalus is highly multifactorial, impaired function of the brain ependymal cells plays a fundamental role in hydrocephalus. Here we show that GemC1 and McIdas, key regulators of multiciliated ependymal cell fate determination, induce direct cellular reprogramming towards ependyma. Our study reveals that ectopic expression of GemC1 and McIdas reprograms cortical astrocytes and programs mouse embryonic stem cells into ependyma. McIdas is sufficient to establish functional activity in the reprogrammed astrocytes. Furthermore, we show that McIdas’ expression promotes ependymal cell regeneration in two different postnatal hydrocephalus mouse models: an intracranial hemorrhage and a genetic form of hydrocephalus and ameliorates the cytoarchitecture of the neurogenic niche. Our study provides evidence on the restoration of ependyma in animal models mimicking hydrocephalus that could be exploited towards future therapeutic interventions.

Biondi bodies are filamentous amyloid inclusions of unknown composition in ependymal cells of the choroid plexuses, ependymal cells lining cerebral ventricles and ependymal cells of the central canal of the spinal cord. Their formation is age-dependent and they are commonly associated with a variety of neurodegenerative conditions, including Alzheimer's disease and Lewy body disorders. Here, we show that Biondi bodies are strongly immunoreactive with TMEM239, an antibody specific for inclusions of transmembrane protein 106B (TMEM106B). Biondi bodies were labelled by both this antibody and the amyloid dye pFTAA. Many Biondi bodies were also labelled for TMEM106B and the lysosomal markers Hexosaminidase A and Cathepsin D. By transmission immuno-electron microscopy, Biondi bodies of choroid plexuses were decorated by TMEM239 and were associated with structures that resembled residual bodies or secondary lysosomes. By electron cryo-microscopy, TMEM106B filaments from Biondi bodies of choroid plexuses were similar (Biondi variant), but not identical, to the fold I that was previously identified in filaments from brain parenchyma. [ABSTRACT FROM AUTHOR]

Ependymal cells (ECs) line the ventricular surfaces of the mammalian central nervous system (CNS) and their development is indispensable to structural integrity and functions of the CNS. We previously reported that EC-specific genetic deletion of the myristoylated alanine-rich protein kinase C substrate (Marcks) disrupts barrier functions and elevates oxidative stress and lipid droplet accumulation in ECs causing precocious cellular aging. However, little is known regarding the mechanisms that mediate these changes in ECs. To gain insight into Marcks-mediated mechanisms, we performed mass spectrometric analyses on Marcks-associated proteins in young and aged ECs in the mouse forebrain using an integrated approach. Network analysis on annotated proteins revealed that the identified Marcks-associated complexes are in part involved in protein transport mechanisms in young ECs. In fact, we found perturbed intracellular vesicular trafficking in cultured ECs with selective deletion of Marcks (Marcks-cKO mice), or upon pharmacological alteration to phosphorylation status of Marcks. In comparison, Marcks-associated protein complexes in aged ECs appear to be involved in regulation of lipid metabolism and responses to oxidative stress. Confirming this, we found elevated signatures of inflammation in the cerebral cortices and the hippocampi of young Marcks-cKO mice. Interestingly, behavioral testing using a water maze task indicated that spatial learning and memory is diminished in young Marcks-cKO mice similar to aged wildtype mice. Taken together, our study provides first line of evidence for potential mechanisms that may mediate differential Marcks functions in young and old ECs, and their effect on forebrain homeostasis during aging. [ABSTRACT FROM AUTHOR]

Nestin is expressed extensively in neural stem/progenitor cells during neural development, but its expression is mainly restricted to the ependymal cells in the adult spinal cord. After spinal cord injury (SCI), Nestin expression is reactivated and Nestin-positive (Nestin+) cells aggregate at the injury site. However, the derivation of Nestin+ cells is not clearly defined. Here, we found that Nestin expression was substantially increased in the lesion edge and lesion core after SCI. Using a tamoxifen inducible CreER(T2)-loxP system, we verified that ependymal cells contribute few Nestin+ cells either to the lesion core or the lesion edge after SCI. In the lesion edge, GFAP+ astrocytes were the main cell type that expressed Nestin; they then formed an astrocyte scar. In the lesion core, Nestin+ cells expressed αSMA or Desmin, indicating that they might be derived from pericytes. Our results reveal that Nestin+ cells in the lesion core and edge came from various cell types and rarely from ependymal cells after complete transected SCI, which may provide new insights into SCI repair. [ABSTRACT FROM AUTHOR]

Congenital hydrocephalus is a major neurological disorder with high rates of morbidity and mortality; however, the underlying cellular and molecular mechanisms remain largely unknown. Reproducible animal models mirroring both embryonic and postnatal hydrocephalus are also limited. Here, we describe a new mouse model of congenital hydrocephalus through knockout of β-catenin in Nkx2.1-expressing regional neural progenitors. Progressive ventriculomegaly and an enlarged brain were consistently observed in knockout mice from embryonic day 12.5 through to adulthood. Transcriptome profiling revealed severe dysfunctions in progenitor maintenance in the ventricular zone and therefore in cilium biogenesis after β-catenin knockout. Histological analyses also revealed an aberrant neuronal layout in both the ventral and dorsal telencephalon in hydrocephalic mice at both embryonic and postnatal stages. Thus, knockout of β-catenin in regional neural progenitors leads to congenital hydrocephalus and provides a reproducible animal model for studying pathological changes and developing therapeutic interventions for this devastating disease. [ABSTRACT FROM AUTHOR]

Ependymal cells located above the ventricular zone of the lateral, third, and fourth ventricles and the spinal cord are thought to form part of the adult neurogenic niche. Many studies have focused on ependymal cells as potential adult neural stem/progenitor cells. To investigate the functions of ependymal cells, a simple method to isolate subtypes is needed. Accordingly, in this study, we evaluated the expression of cluster of differentiation (CD) 9 in ependymal cells by in situ hybridization and immunohistochemistry. Our results showed that CD9-positive ependymal cells were also immunopositive for SRY-box 2, a stem/progenitor cell marker. We then isolated CD9-positive ependymal cells from the third ventricle using the pluriBead-cascade cell isolation system based on antibody-mediated binding of cells to beads of different sizes and their isolation with sieves of different mesh sizes. As a result, we succeeded in isolating CD9-positive populations with 86% purity of ependymal cells from the third ventricle. We next assayed whether isolated CD9-positive ependymal cells had neurospherogenic potential. Neurospheres were generated from CD9-positive ependymal cells of adult rats and were immunopositve for neuron, astrocyte, and oligodendrocyte markers after cultivation. Thus, based on these findings, we suggest that the isolated CD9-positive ependymal cells from the third ventricle included tanycytes, which are special ependymal cells in the ventricular zone of the third ventricle that form part of the adult neurogenic and gliogenic niche. These current findings improve our understanding of tanycytes in the adult third ventricle in vitro. [ABSTRACT FROM AUTHOR]

Adult neural stem cells (NSCs) reside in specialized niches, which hold a balanced number of NSCs, their progeny, and other cells. How niche capacity is regulated to contain a specific number of NSCs remains unclear. Here, we show that ependyma‐derived matricellular protein CCN1 (cellular communication network factor 1) negatively regulates niche capacity and NSC number in the adult ventricular–subventricular zone (V‐SVZ). Adult ependyma‐specific deletion of Ccn1 transiently enhanced NSC proliferation and reduced neuronal differentiation in mice, increasing the numbers of NSCs and NSC units. Although proliferation of NSCs and neurogenesis seen in Ccn1 knockout mice eventually returned to normal, the expanded NSC pool was maintained in the V‐SVZ until old age. Inhibition of EGFR signaling prevented expansion of the NSC population observed in CCN1 deficient mice. Thus, ependyma‐derived CCN1 restricts NSC expansion in the adult brain to maintain the proper niche capacity of the V‐SVZ. Synopsis: The maintenance and activation of NSCs are regulated by niche components in the adult brain. Matricellular protein CCN1 is secreted from ependymal cells and negatively regulates the number of NSCs through restricting niche capacity in the adult V‐SVZ. Matricellular protein CCN1 is specifically expressed in the ependymal cells in the adult V‐SVZ.CCN1 restricts the capacity of the V‐SVZ NSC niche and the number of NSCs.CCN1 suppresses the expansion of NSC pool through inhibiting EGFR signaling. [ABSTRACT FROM AUTHOR]

Abstract Hydrocephalus is a common congenital anomaly. LCAM1 and MPDZ (MUPP1) are the only known human gene loci associated with non‐syndromic hydrocephalus. To investigate functions of the tight junction‐associated protein Mpdz, we generated mouse models. Global Mpdz gene deletion or conditional inactivation in Nestin‐positive cells led to formation of supratentorial hydrocephalus in the early postnatal period. Blood vessels, epithelial cells of the choroid plexus, and cilia on ependymal cells, which line the ventricular system, remained morphologically intact in Mpdz‐deficient brains. However, flow of cerebrospinal fluid through the cerebral aqueduct was blocked from postnatal day 3 onward. Silencing of Mpdz expression in cultured epithelial cells impaired barrier integrity, and loss of Mpdz in astrocytes increased RhoA activity. In Mpdz‐deficient mice, ependymal cells had morphologically normal tight junctions, but expression of the interacting planar cell polarity protein Pals1 was diminished and barrier integrity got progressively lost. Ependymal denudation was accompanied by reactive astrogliosis leading to aqueductal stenosis. This work provides a relevant hydrocephalus mouse model and demonstrates that Mpdz is essential to maintain integrity of the ependyma.

The stomach-derived hormone ghrelin mainly acts in the brain. Studies in mice have shown that the accessibility of ghrelin into the brain is limited and that it mainly takes place in some circumventricular organs, such as the median eminence. Notably, some known brain targets of ghrelin are distantly located from the circumventricular organs. Thus, we hypothesized that ghrelin could also access the brain via the blood-cerebrospinal fluid (CSF) barrier, which consists of the choroid plexus and the hypothalamic tanycytes. Using systemic injection of ghrelin or fluorescent-ghrelin in mice, we found that cells of the blood-CSF barrier internalize these molecules. In time-response studies, we found that peripherally injected fluorescent-ghrelin quickly reaches hypothalamic regions located in apposition to the median eminence and more slowly reaches the periventricular hypothalamic parenchyma, adjacent to the dorsal part of the third ventricle. Additionally, we found that CSF ghrelin levels increase after the systemic administration of ghrelin, and that central infusions of either an anti-ghrelin antibody, which immuno-neutralizes CSF ghrelin, or a scrambled version of ghrelin, which is also internalized by cells of the blood-CSF barrier, partially impair the orexigenic effect of peripherally injected ghrelin. Thus, current evidence suggests that the blood-CSF barrier can transport circulating ghrelin into the brain, and that the access of ghrelin into the CSF is required for its full orexigenic effect. [ABSTRACT FROM AUTHOR]

IIIG9 is the regulatory subunit 32 of protein phosphatase 1 (PPP1R32), a key phosphatase in the regulation of ciliary movement. IIIG9 localization is restricted to cilia in the trachea, fallopian tube, and testicle, suggesting its involvement in the polarization of ciliary epithelium. In the adult brain, IIIG9 mRNA has only been detected in ciliated ependymal cells that cover the ventricular walls. In this work, we prepared a polyclonal antibody against rat IIIG9 and used this antibody to show for the first time the ciliary localization of this protein in adult ependymal cells. We demonstrated IIIG9 localization at the apical border of the ventricular wall of 17-day-old embryonic (E17) and 1-day-old postnatal (PN1) brains and at the level of ependymal cilia at 10- and 20-day-old postnatal (PN10–20) using temporospatial distribution analysis and comparing the localization with a ciliary marker. Spectral confocal and super-resolution Structured Illumination Microscopy (SIM) analysis allowed us to demonstrate that IIIG9 shows a punctate pattern that is preferentially located at the borders of ependymal cilia in situ and in cultures of ependymocytes obtained from adult rat brains. Finally, by immunogold ultrastructural analysis, we showed that IIIG9 is preferentially located between the axoneme and the ciliary membrane. Taken together, our data allow us to conclude that IIIG9 is localized in the cilia of adult ependymal cells and that its expression is correlated with the process of ependymal differentiation and with the maturation of radial glia. Similarly, its particular localization within ependymal cilia suggests a role of this protein in the regulation of ciliary movement. [ABSTRACT FROM AUTHOR]

Hypothalamic tanycytes are specialized bipolar ependymal cells that line the floor of the third ventricle. Given their strategic location, tanycytes are believed to play several key functions including being a selective barrier and controlling the amount of hypothalamic-derived factors reaching the anterior pituitary. The in vitro culture of these cells has proved to be difficult. Here, we report an improved method for the generation of primary cultures of rat hypothalamic tanycytes. Ependymal cultures were derived from tissue dissected out of the median eminence region of 10-day-old rats and cultured in a chemically defined medium containing DMEM:F12, serum albumin, insulin, transferrin and the antibiotic gentamycin. After 7 days in vitro, ∼30% of the cultured cells exhibited morphological features of tanycytes as observed by phase contrast or scanning electron microscopy. Tanycyte-like cells were strongly immuno-reactive for vimentin and dopamine-cAMP-regulated phospho-protein (DARPP-32) and weakly immune-reactive for glial fibrillary acidic protein. Tanycyte-like cells displayed a stable negative resting plasma membrane potential and failed to show spiking properties in response to current injections. When exposed to fluorescent beads in the culture medium, tanycyte-like cells exhibited a robust endocytosis. Thus, the present method effectively yields cultures containing tanycyte-like cells that resemble in vivo tanycytes in terms of morphologic features and molecular markers as well as electrical and endocytic activity. To our knowledge, this is the first protocol that allows the culturing of tanycyte-like cells that can be individually identified and that conserve the morphology of tanycytes in their natural physiological environment. [ABSTRACT FROM AUTHOR]

During the whole life cycle of mammals, new neurons are constantly regenerated in the subgranular zone of the dentate gyrus and in the subventricular zone of the lateral ventricles. Thanks to emerging methodologies, great progress has been made in the characterization of spinal cord endogenous neural stem cells (ependymal cells) and identification of their role in adult spinal cord development. As recently evidenced, both the intrinsic and extrinsic molecular mechanisms of ependymal cells control the sequential steps of the adult spinal cord neurogenesis. This review introduces the concept of adult endogenous neurogenesis, the reaction of ependymal cells after adult spinal cord injury (SCI), the heterogeneity and markers of ependymal cells, the factors that regulate ependymal cells, and the niches that impact the activation or differentiation of ependymal cells. [ABSTRACT FROM AUTHOR]

Ependymal cells form an epithelium lining the ventricular cavities of the vertebrate brain. Numerous methods to obtain primary culture ependymal cells have been developed. Most of them use foetal or neonatal rat brain and the few that utilize adult brain hardly achieve purity. Here, we describe a simple and novel method to obtain a pure non-adherent ependymal cell culture from explants of the striatal and septal walls of the lateral ventricles. The combination of a low incubation temperature followed by a gentle enzymatic digestion allows the detachment of most of the ependymal cells from the ventricular wall in a period of 6 h. Along with ependymal cells, a low percentage (less than 6 %) of non-ependymal cells also detaches. However, they do not survive under two restrictive culture conditions: (1) a simple medium (alpha-MEM with glucose) without any supplement; and (2) a low density of 1 cell/µl. This purification method strategy does not require cell labelling with antibodies and cell sorting, which makes it a simpler and cheaper procedure than other methods previously described. After a period of 48 h, only ependymal cells survive such conditions, revealing the remarkable survival capacity of ependymal cells. Ependymal cells can be maintained in culture for up to 7-10 days, with the best survival rates obtained in Neurobasal supplemented with B27 among the tested media. After 7 days in culture, ependymal cells lose most of the cilia and therefore the mobility, while acquiring radial glial cell markers (GFAP, BLBP, GLAST). This interesting fact might indicate a reprogramming of the cell identity. [ABSTRACT FROM AUTHOR]

Early cerebellum development in humans is poorly understood. The present study histologically examined sections from 20 human embryos and fetuses at 6 weeks (12-16 mm crown-rump length (CRL); 4 specimens), 7-9 weeks (21-39 mm CRL; 8 specimens), 11-12 weeks (70-90 mm CRL; 4 specimens) and 15-16 weeks (110-130 mm CRL; 4 specimens). During 7-9 weeks (approximate CRL 28 mm), the rhombic lip (a pair of thickenings of the alar plate) protruded dorsally, bent laterally, extended ventrolaterally and fused with the medially located midbrain. During that process, the primitive choroid plexus appeared to become involved in the cerebellar hemisphere to form a centrally located eosinophilic matrix. At that stage, the inferior olive had already developed in the thick medulla. Thus, the term 'bulbo-pontine extension' may represent an erroneous labeling of a caudal part of the rhombic lip. The cerebellar vermis developed much later than the hemisphere possibly from a midline dark cell cluster near the aqueduct. In the midline area after 12 weeks (80 mm CRL), the growing bilateral hemispheres seem to provide mechanical stress such as rotation and shear that cause the development of several fissures much deeper than those on the hemisphere. The rapidly growing surface germinal layer may be a minor contributor to this vermian fissure formation. The vermian fissures seem to enable inside involvement of the surface germinal cells, and to induce cytodifferentiation of the vermis. Consequently, in the early stages, it appears that the cerebellar hemisphere and vermis develop independently of each other. [ABSTRACT FROM AUTHOR]

During the last stage of infection in the experimental scrapie-infected hamster model, light microscopy reveals typical immunostaining of PrPsc in the subependymal region and at the apical ependymal cell borders. Whereas the subependymal immuno-staining is known to originate from extracellular amyloid filaments and residual membranes of astrocytes as constituents of plaque-like structures, the ultrastructural correlate of the supraependymal PrPsc staining remains uncertain. To decipher this apical PrPsc immunopositivity and subsequently the ependymocyte–scrapie agent interaction, we employed highly sensitive immuno-electron microscopy for detecting PrPsc in 263K scrapie-infected hamster brains. The results revealed the supraependymal PrPsc signal to be correlated not only with extracellular accumulation of amyloid filaments, but also with three distinct ependymal cell structures: (1) morphologically intact or altered microvilli associated with filaments, (2) the ependymal cell cytoplasm in proximity of apical cell membrane, and (3) intracytoplasmic organelles such as endosomes and lysosomal-like structures. These findings suggest a strong ependymotrope feature of the scrapie agent and recapitulate several aspects of the cell–prion interaction leading to the formation and production of PrPsc amyloid filaments. Our data demonstrate that in addition to neurons and astrocytes, ependymocytes constitute a new cellular target for the scrapie agent. In contrast, the absence of PrPsc labeling in choroid plexus and brain vascular endothelial cells indicates that these cells are not susceptible to the infection and may inhibit passage of the infectious agent across the blood–brain barrier. [ABSTRACT FROM AUTHOR]

The present work was undertaken to reveal the mechanism of cerebral aqueduct agenesis found to result in hydrocephalus following intrauterine exposure to model teratogen, cyclophosphamide, in murine fetuses. A single dose of 10-mg/kg body weight cyclophosphamide was injected intaperitoneally to pregnant mice on day 10, 11 or 12 of gestation. Fetuses were collected through abdominal incision on day 18 and studied for various malformations of brain and cranium including hydrocephalus. Incomplete development and failure of canalization of the cerebral aqueduct were detected when serial sections of brain in coronal and transverse planes were studied under the microscope. Biotechnological investigations such as % DNA fragmentation, % viable cell count and cell proliferation assay were carried out on brain cells for further studies. Agenesis and non-canalization of the cerebral aqueduct resulted in increased pressure of CSF, which led to rupture of the aqueduct complicated by leakage and accumulation of CSF in brain substance forming a cavity containing CSF parallel and lateral to the unopened part of the cerebral aqueduct. Incomplete development along with non-canalization of the cerebral aqueduct resulted in blockage of CSF flow through the ventricles that manifest as internal hydrocephalus. External hydrocephalus on the other hand was detected where the CSF accumulated in the cavity formed inside the brain substance and established communication with the CSF in the subarachnoid space. Cyclophosphamide induced inhibition of mitosis and cell differentiation of ependymal cells reflecting a decreased % viable cell count and cell proliferation assay along with augmentation of apoptosis of brain cells quantified as increased % DNA fragmentation count, which were identified as the contributing factors underlying the agenesis and incomplete development of the cerebral aqueduct. The study also suggests that cell survival, proliferation, migration or differentiation of ependymal cells might have been affected, and we speculate that CSF may have an inducing role in the development and canalization of the cerebral aqueduct. [ABSTRACT FROM AUTHOR]

GFAP expression patterns were compared between the brains of a spiny dogfish (Squalusacanthias) and a little skate (Raiaerinacea). After anesthesia, the animals were perfused with paraformaldehyde. Serial vibratome sections were immunostained against GFAP using the avidin-biotin method. Spiny dogfish brain contained mainly uniformly-distributed, radially arranged ependymoglia. From GFAP distribution, the layered organization in both the telencephalon and the tectum were visible. In the cerebellum, the molecular and granular layers displayed conspicuously different glial structures; in the former a Bergmann glia-like population was found. No true astrocytes (i.e., stellate-shaped cells) were found. Radial glial endfeet lined all meningeal surfaces. Radial fibers also seemed to form endfeet and en passant contacts on the vessels. Plexuses of fine perivascular glial fibers also contributed to the perivascular glia. Compared with spiny dogfish brain, GFAP expression in the little skate brain was confined. Radial glia were limited to a few areas, e.g., segments of the ventricular surface of the telencephalon, and the midline of the diencephalon and mesencephalon. Scarce astrocytes occurred in every brain part, but only the optic chiasm, and the junction of the tegmentum and optic tectum contained large numbers of astrocytes. Astrocytes formed the meningeal glia limitans and the perivascular glia. No GFAP-immunopositive Bergmann glia-like structure was found. Astrocytes seen in the little skate were clearly different from the mammalian and avian ones; they had a different process system – extra large forms were frequently seen, and the meningeal and perivascular cells were spread along the surface instead of forming endfeet by processes. The differences between Squalus and Raia astroglia were much like those found between reptiles versus mammals and birds. It suggests independent and parallel glial evolutionary processes in amniotes and chondrichthyans, seemingly correlated with the thickening of the brain wall, and the growing complexity of the brain. There is no strict correlation, however, between the replacement of radial ependymoglia with astrocytes, and the local thickness of the brain wall. [ABSTRACT FROM AUTHOR]

Testican is a putative extracellular heparan/chondroitin sulfate proteoglycan of unknown function that is expressed in a variety of human tissues at widely different levels but is most abundant in the brain. In mice, testican mRNA has been detected only in brain and it is therefore likely to have an important function in the central nervous system. RNA blot analysis reveals the relative intensity of testican in various regions of the human brain. Levels of testican message are most pronounced in the thalamus, hippocampus, occipital lobe, nucleus accumbens, temporal lobe, and caudate nucleus, with somewhat lower levels in the cerebral cortex, medulla oblongata, frontal lobe, amygdala, putamen, spinal cord, substantia nigra, and cerebellum. In situ hybridization reveals the cellular distribution of the mRNA within these areas to be highest in neurons and in choroid plexus epithelium, and moderately lower in ependymal cells lining the ventricles and in vascular endothelial cells. Testican mRNA is not detected in oligodendrocytes or in most astrocytes. However, astrocytes in regions of reactive gliosis do express testican mRNA. These findings, along with a cysteine-rich pattern similarity to neurocan, brevican, versican, and other proteoglycans found in brain, suggest that testican may be a part of the specialized extracellular matrix of the brain. [ABSTRACT FROM AUTHOR]

The signaling action of neuropeptides in the brain is terminated by breakdown through extracellular peptidases and subsequent removal of the peptide fragments from the extracellular fluid via specific transporter proteins. Here we describe the anatomical distribution in the rat nervous system of the recently isolated high affinity peptide transporter PEPT2. Using nonisotopic in situ hybridization we demonstrate that PEPT2 mRNA is expressed in brain by astrocytes, subependymal cells, ependymal cells and epithelial cells of choroid plexus. Furthermore, PEPT2 is expressed in retina by Müller cells and in dorsal root ganglia by satellite cells. The mRNA levels of PEPT2 in astrocytes are moderate and relatively homogenous throughout the brain except for an area in ventral forebrain where PEPT2 levels are below average. PEPT2 mRNA expression is weakly upregulated in reactive astrocytes that were stimulated through an injection of the glutamatergic neurotoxin quinolinic acid. These data suggest that removal of neuropeptide fragments from brain extracellular fluid occurs via PEPT2 expressed in astrocytes, ependymal cells and choroid plexus epithelial cells. [ABSTRACT FROM AUTHOR]

LRH injected into the third ventricle induced increases of serum LH and FSH in the rat. Following injection of LRH-I into the third ventricle, autoradiographic grains were concentrated in the ependymal cell bodies and ependymal processes in the median eminence. Aggregations of grains were also found in the capillaries of the portal vessels, and in both sinusoids and parenchymal cells of the adenohypophysis. These findings show that the ependymal cells represent a link between the cerebrospinal fluid and the portal blood. [ABSTRACT FROM AUTHOR]

Images of ependymal cell proliferation after experimental spinal cord injury in the rabbit are presented. This finding suggests that segmental central canal obliteration after injury could be considered in the pathogenesis of posttraumatic syringomyelia. [ABSTRACT FROM AUTHOR]

Ependymal cells in the ventricular wall and in several circumventricular organs of the rat were compared by means of freeze-fracturing. In principle, tight junctions and orthogonal arrays of particles (OAP) do not coexist in the cells bordering the ventricular wall: (1) Ordinary ependymal cells of the rat possess OAP and are devoid of tight junctions. (2) Epithelial cells of the rat choroid plexus are connected by tight junctions; OAP are lacking here. In some cases, however, tight junctions and OAP coexist in the same cell. In the boundary zone between choroid plexus and ependyma of the rat, the density of OAP is very low, whereas the tight junctions are well developed. In the subfornical and the subcommissural organ (SCO) of the rat both structures are poorly developed; in the SCO they occur segregated in different membranous areas. An overview of the literature confirms that tight junctions and OAP mostly exclude each other. The possibility that in astrocytes and ependymal cells tight junctions may have been replaced by OAP during phylogeny is briefly discussed. [ABSTRACT FROM AUTHOR]

Glycoproteins rich in mannosyl or glucosyl residues were analyzed in the subcommissural organ (SCO) and the pineal organ of the sheep ( Ovis aries). By use of concanavalin A labelled with fluorescein isothiocyanate, fluorescent material was found both in ependymal and hypendymal cells of the SCO. In the pineal organ, either isolated or grouped parenchymal cells showed a marked fluorescence. These cells may correspond to ependymal elements also called 'interstitial cells' or 'supporting cells'. In addition, scarce slender, fluorescent processes were observed in the pineal parenchyma. The techniques of electrophoresis and electrotransfer on nitrocellulose paper have been applied to analyze the glycopeptide content of the SCO and the pineal organ in comparison to cerebellar and cerebral fractions solubilized by use of Triton X 100. Approximately 30 different concanavalin A-reactive glycopeptides were revealed in each fraction. In the SCO extract four glycopeptides (30, 54, 72, 100 kd) might correspond to subunits of the glycoprotein(s) characteristically stored in the ependymal cells of the SCO. In addition, two glycopeptides (32/33, 115 kd) are specific to the pineal organ extract. The possible similarity of the concanavalin A-reactive material in both organs is discussed and a putative secretory activity of the pineal ependymal cells is postulated. [ABSTRACT FROM AUTHOR]

The architecture of astrocytic membranes in the optic nerve and the spinal cord of the lizard, Anolis carolinensis, was investigated by use of the freeze-fracturing technique. Whereas astrocytes in mammals reveal so-called orthogonal arrays of particles (OAPs) in their membranes, astrocytes in lower vertebrates lack these structures. This study demonstrates for the first time OAPs in astrocytes from a submammalian species. They were found commonly in the optic nerve and less frequently in the spinal cord. However, the OAPs in astrocytes of spinal cord were confined to midtrunk levels; the astrocytes in the caudal spinal cord failed to reveal OAPs. Additionally, the ependymal cells around the central canal did not show any OAPs, either in the thoracic or in the caudal spinal cord. They were interconnected by gap and tight junctions, which were not intercalated with each other. The findings support our current working hypothesis according to which the presence and absence of OAPs in astrocytes may be correlated with regenerative incapability or capability of CNS-structures; i.e., whereas the thoracic spinal cord in Anolis carolinensis is known to be incapable of regeneration after injury, the caudal spinal cord is regenerative. [ABSTRACT FROM AUTHOR]

The synchronizing effect of ethinylestradiol (4 μg/g b.w.) on neurons of the arcuate nucleus 700-950 μm caudal to the posterior edge of the optic chiasma was studied by karyometry in 6-week-old albino mice during proestrus. The caudal portion of the arcuate nucleus was identified as the most estrogen-sensitive subdivision; all neurons showed an increase in their nuclear area (mean transect, profile area of the nucleus) 1 h following administration of ethinylestradiol. This hypothalamic region was selected for the subsequent electron-microscopic cytometric study to analyze functional interrelationships among neurons, ependymal cells and glial cells. Six and 12 days after ovariectomy no significant change in the nuclear area of neurons and ependymal cells was found 850-950 μm behind the posterior slope of the optic chiasma, but the neurons exhibited a decrease in the number of polyribosomes, the volume fraction (V) and the surface density of the inner membrane of mitochondria (S). A similar decrease in V and S was measured in the apical part of ependymal cells and in the pericapillary profiles of ependymal and glial cells, which was accompanied by a reduction in the surface density of ependymal processes extending into the ventricular lumen. In addition, no change of V and S was seen in the basal subnuclear part of ependymal cells. This bipolar functional reaction of ependymal cells after ovariectomy is discussed as an indicator of ependymal control of neuronal activity by sequestering biologically active agents, e.g., transmitters of neurohormones, in their apical and basal extensions facing the ventricular surface or the pericapillary space. [ABSTRACT FROM AUTHOR]

In ependymal cells of the mouse the neck region of all cilia examined by means of transmission electron microscopy exhibited rows of electron-dense spines. These structures correspond to the ciliary necklace reported from freeze-etch studies, a structure presumed to serve as an energy-regulating system in motile cilia. [ABSTRACT FROM AUTHOR]

The distribution of the tracer substance horseradish peroxidase (HRP, Mw 40,000) in the neuro-intermediate lobe of the lizard, Anolis carolinensis, was studied at various time intervals (13 min to 24 h) after vascular injection. HRP rapidly entered the extracellular lumen of the neural lobe, but did not penetrate into the third ventricle. The tracer was found in micropinocytotic vesicles (MPVs) of ependymal cells within 13 min after injection. The number of cellular inclusions containing HRP increased during the period of observation (24 h). The tracer was sparsely taken up by aminergic and peptidergic nerve terminals of the external layer. After transection of the hypophysial stalk, numerous dense, labelled droplets were found in the peptidergic terminals, and the number of labelled inclusions in ependymal cells increased. MPVs were frequently found in extensions of stellate cells of the intermediate lobe, and endocytotic vacuoles (EVs) developed especially in the perikaryon. HRP was also found in large cisternae of the secretory cells, appearing predominantly towards the perivascular septum (PVS). These cisternae were found to communicate with the extracellular lumen, probably representing a system of the extracellular space extending into the secretory cell. After transection of the hypophysial stalk, there was an increase in the number of small EVs in secretory cells of the intermediate lobe. The results are discussed in terms of MSH-release regulation and possible participation of the extracellular lumen, glial and stellate cells in the transport of regulating factors and secretory material. [ABSTRACT FROM AUTHOR]

The ultrastructure of the ependymal cells in the area postrema of the domestic fowl was studied by scanning and transmission electron microscopy. The ependymal surface of the area postrema is covered with many furrows and ridges. These ridges consist of ependymal cells aggregated in a fan-like shape. The ependymal cell lacks clustered cilia, microvilli are few, and a long basal process extends through the parenchymal layer of the area postrema. Within the cytoplasm as well as in the basal process, a spherical body with a diameter ranging from 1.5 to 2 gmm is occasionally observed. [ABSTRACT FROM AUTHOR]

Intranuclear microfilament bundles were observed in ependymal cells of the third ventricle of the rat. They appeared as single, cylindricallyshaped fibrillar bands up to 4.9 μm in length. In cross sections of bundles, the microfilaments exhibited an apparently regular arrangement. They were separated by a distance of 4 to 5 nm, measuring approximately 11 nm from centre to centre. One bundle consisted of 55 to 88 microfilaments. The intranuclear bundles terminated at the inner membrane of the nucleus. They exhibited close spatial relationship to perinuclear chromatin, chromatin centres, intrachromatin granules and fibrillar structures. The average ratio of nuclear sections with and without bundles was 1∶15. [ABSTRACT FROM AUTHOR]

Freeze-etched preparations of the ventricular surfaces of ependymal cells clearly reveal the presence of pinocytotic vesicles opening into the third ventricle and large vacuoles formed by broad cell projections. The density of the vesicular openings is approximately 20 per μm. The ependymal cells in the median eminence of the frog are adjoined by tight junctions comprised of five to eight interconnected junctional strands, whereas near the median eminence in the mouse only one to two such strands form the tight junction of the ependymal cells. Gap junctions between the adjacent ependymal cells are detected near the median eminence in the mouse but not in the frog. [ABSTRACT FROM AUTHOR]

Actin and myosin were located in astrocytes, marginal glial and ependymal cells in rat diencephalon by using antibodies against highly purified chicken gizzard actin and myosin. On the basis of these findings it is suggested that glial cell motility in vivo and in vitro is due to the presence of an intracellular actin/myosin system. [ABSTRACT FROM AUTHOR]

The ultrastructure of the normal median eminence of the male rhesus monkey ( Macaca mulatta) is described using high-voltage electron microscopy. Surface specializations of ependymal cells lining the infundibular recess included cilia, apical extrusions, and microvilli. Supraependymal cells were predominantly macrophage-like, but examples of lymphocytic types were also seen. Tanycytes had long, branching, basal processes filled with numerous microtubules, some lipid droplets, and granules. The zona interna was composed of large unmyelinated neurosecretory fibers. A few myelinated fibers were also seen, but their character as neurosecretory fibers could not be established. The zona externa was composed of densely-packed profiles of neurosecretory fibers of small diameter, was well-vascularized and contained the terminations of tanycytes. Perivascular glial cells, vesiculated elements, pituicytes, and cellular elements common to connective tissue were observed. The intricate relationships between both the cellular and fibrous elements of the median eminence can be appreciated with the capability of high-voltage electron microscopy to discern ultrastructure in sections 10 times thicker than those used for low-voltage electron microscopy. The median eminence of this primate species has an ultrastructural organization similar to that described for most other species. [ABSTRACT FROM AUTHOR]

Immunohistochemical localization of luteinizing hormone-releasing hormone (LH-RH), during different phases of the estrus cycle, in the preoptic, suprachiasmatic and arcuate nuclei, and in the OVLT of rats, with special emphasis on the ependymal cells, was studied by light, fluorescent and electron microscopy, by using rabbit anti serum to synthetic LH-RH. The LH-RH neurons in the above mentioned areas, were very active during late diestrus and early proestrus phases. Specialized ependymal cells bordering the 3rd ventricle also showed varied LH-RH positive reaction during different phases of the estrus cycle. Immunofluorescent studies showed cyclic variations in the LH-RH material in the CSF of the preoptic and infundibular recesses, as well as in the 3rd ventricle near OVLT, in that, it was maximum during late diestrus and early proestrus phases. Immediately after this, the LH-RH material started reducing in these areas, and had almost vanished by the time late proestrus was reached. We have also observed that during the proestrus phase, as the LH-RH material started declining in the CSF, it had started building up in the specialized ependyma. Estrus, metaestrus and early diestrus phases showed very weak immunofluorescent LH-RH material in the lumen of the infundibular recess and in the specialized ependyma. Our immuno-electron microscopic observations showed pleomorphic LH-RH granules in the specialized ependyma during late diestrus and proestrus phases. All these observations lead us to believe that LH-RH is not synthesized in the ependymal cells, but is phagocytosed from the CSF of the 3rd ventricle by the specialized ependyma, which transports it to the ME portal system. In males, the fluorescent LH-RH material did not show any noticeable changes. With the present and previous work, it is concluded that the neurons in different nuclei synthesize LH-RH and transport it to the ME portal system, primarily through the nerve fibers and secondarily by the ventricular route. It is also sugested that the ependymal transport of LH-RH to the ME portal system is cyclic and thus controls the gonadotropin secretion. [ABSTRACT FROM AUTHOR]

The mainstays of primary treatment of ependymoma are aggressive surgery followed by radiotherapy. Although spreading occasionally occurs in the cerebrospinal pathways, chemotherapy is still not established and no ultimate drug has so far been found. Estramustine-phosphate (EMP), with a demonstrated effect on astrocytoma in vitro, has been shown to penetrate the blood-tumor barrier and to accumulate in human brain tumor tissue including ependymoma. It has been proposed that the cytotoxic effect of EMP depends on the presence of a binding protein, estramustine-binding protein (EMBP). In the present paper we have, for the first time, immunohistochemically demonstrated an EMBP-like protein in a series of ependymomas. Immunoreactivity was found within the cytoplasm of the tumor cells with a tendency to increase with increasing malignancy of the tumor. In addition, the occurence of EMBP-like protein was demonstrated in human ependymal cells. In the rat brain, a weak immunoreactivity was detected in early fetal neuroepithelial cells while the staining intensity was increased in mature ependymal cells in late fetal, neonatal, and adult rat. Thus, immunoreactivity for an EMBP-like protein was demonstrated in ependymoma tissue, normal human ependyma and in the developing rat ependymal cells. [ABSTRACT FROM AUTHOR]

The ultrastructural characteristics of a concentration of mitotically active cells near the lateral margin of the lateral ventricle was studied in the neonatal rabbit brain. This area, termed the caudato-pallial area, consists of subependymal cells, neuroblasts, microglial cells and astroblasts bordering a layer of ependymal epithelial cells. The character of the ependymal cells in this area indicates a persistence of embryonic conditions favoring the development of neuroblasts and astroblasts. It is not known what type of cells are produced in the subependymal layer since the primitive subependymal cells have no features which would clearly relate them to nerve or glial cells. Of the 80 subependymal cells which were reconstructed from serial sections none have processes extending beyond the limits of the subependymal layer. They range in form from simple cells devoid of processes to complex cells with several short cellular extensions. The presence of intermediate forms with transitional cytologic features indicates simple and complex cells may represent different stages in the mitotic cycle of a single cell type. No direct evidence was obtained to support the concept that the small ramified microglial cells of the caudato-pallial area or round and amoeboid microglial cells near the corpus callosum arise through mitosis of nearby ependymal or subependymal cells. [ABSTRACT FROM AUTHOR]

The extremely hypertrophied hypothalamo-hypophysial neurosecretory system of the hereditary nephrogenic diabetes insipidus (DI Os/+) mice, shows distinctly hypertrophied median eminence (ME) indicated by the hypertrophied ependymal cells, increased number of Herring bodies and a great number of neurosecretory fibers loaded with secretion converged towards blood capillaries and towards pars tuberalis. The other distinct feature of the DI Os/+ mice is the marked hypertrophy and hyperplasia of the pars intermedia (PI) infiltrated with many beaded neurosecretory fibers. The majority of the PI glandular cells are of the large light type rather than the small type, the latter being comparatively numerous in normal mice. Very often these large light glandular cells and the marginal cuboidal epithelial cells of the hypophysial cleft contain or are surrounded by AF granules and/or colloid. When the SON is destroyed bilaterally by means of electrolytic lesions, loss of NSM from the ME, infundibulum and the pars nervosa, is accompanied by the disappearance of AF granules in the PI, and the shrinkage of all the above mentioned parts. The changes also occur in the three glandular cell types. The large light glandular cells, decrease in number as well as in size, while the granularity of the dark cells increases. The marginal cuboidal cells of the hypophysial cleft and the large light glandular cells loose most of the AF material from the cells as well as from the surroundings. Wherever the AF material does not disappear in the PI, the PI cells remain much big, as in the controls. The colloidal secretion on the margin of the PI and from the cleft may be the secretion of PI glandular cells, as a result of the stimulation of the neurosecretory fibers in the PI of DI Os/+ mice. This colloid disappears when the neurosecretory fibers are destroyed from the PI as a result of lesioning. Therefore, it suggests that this secretory material may belong to SON, or possibly to some other secretory cells residing in the anterior hypothalamic region. Or more probable explanation could be that with the disappearance of supraoptic neurons and/or anterior hypothalamic neurons due to electryolytic lesions, the secretory activity of the PI cells and its AF material, might have been hampered due to lack of stimulation of these fibers in PI. Therefore, from the previous work and from the present one, it is suggested that the NSM may contain some substance which stimulates the glandular cells of the PI and this substance may be either vasopressin and/or a type of CRF. However, it is concluded that the activated neurosecretory system plays an important role in the hypertrophy of the ME and PI and certain changes in PI cell types. The impairment of catecholamine fibers and their lack of influence on synthesis and/or release in PI parenchyma cells is discussed. [ABSTRACT FROM AUTHOR]

In the developing cerebellum of 8-day old rats surgical lesions were made. During regeneration of the cerebellum the pia mater was found to penetrate inside the neural tissue. Partially differentiated Purkinje cells and granule cells, that were in close contact with the pial cells, were found atrophied. When the profilerative cells of external granular layer came into contact with the pial cells, they were reduced to a primitive type of epitheloid cells. In this instance epithelio-mesenchymal interaction was found deleterious to the precursors of neurons. However, when the epithelioid cells were freed from the contact with the pial cells by intervening basement membrane, they differentiated into ependymal cells. Such ependymal cells gave rise to small as well as large new ventriculer structures, and structures resembling chorioid plexus. [ABSTRACT FROM AUTHOR]

Studies of the effects of neurotransmitter amino acids on the motor activity of the ciliary apparatus of ependymal cells of the midbrain cerebral aqueduct in neonatal white mice showed that addition of glutamate, GABA-aminobutyric acid, glycine, and taurine to the culture medium induced slowing and then complete cessation of the activity of the ciliary apparatus. Inhibition and cessation of the activity of cilia in response to neurotransmitters, especially glutamate at high concentrations, demonstrated the existence of the corresponding receptors on the membranes of ependymal cells in the cerebral aqueduct, and this was supported by experiments with prior addition of ion channel blockers to the growth medium, i.e., ketamine, strychnine, and bicuculline, which weakened the destructive influences of neurotransmitters and increased the duration of functioning of the ciliary apparatus. [ABSTRACT FROM AUTHOR]