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The dynamics of cerebrospinal fluid (CSF) are essential for maintaining homeostasis in the central nervous system. Despite insufficiently detailed descriptions of their structural and molecular properties for a century, cranial arachnoid granulations (CAGs) on meninges have been thought to participate in draining CSF from the subarachnoid space into the dural sinuses. However, recent studies have demonstrated the existence of other types of CSF drainage systems, such as lymphatic vessels adjacent to dural sinus and paravascular space in the brain so-called glymphatic system. Therefore, the role of CAGs in CSF drainage has become dubious. To better understand CAG function, we analyzed the ultrastructure and molecular identity of CAG-like structure on meninges adjacent to the superior sagittal sinus of pigs. Transmission electron microscopy analysis revealed that this structure has a reticular conglomerate consisting of endothelial cells that resembles lymphatic linings. Furthermore, immunohistochemistry and immunoelectron microscopy showed that they express molecules specific to lymphatic endothelial cell. We coined a name 'CAG-like dural gap (CAG-LDG)' to this structure and discussed the physiological relevance in terms of CSF drainage., (© The Author(s) 2020. Published by Oxford University Press on behalf of The Japanese Society of Microscopy. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

The leptomeninges, referring to the arachnoid and pia mater and their projections into the perivascular compartments in the central nervous system, actively participate in diverse biological processes including fluid homeostasis, immune cell infiltrations, and neurogenesis, yet their detailed cellular and molecular identities remain elusive. This study aimed to characterize platelet-derived growth factor beta (PDGFR-β)-expressing cells in the leptomeninges in the adult rat brain using light and electron microscopy. PDGFR-β + cells were observed in the inner arachnoid, arachnoid trabeculae, pia mater, and leptomeningeal sheath of the subarachnoid vessels, thereby forming a cellular network throughout the leptomeninges. Leptomeningeal PDGFR-β + cells were commonly characterized by large euchromatic nuclei, thin branching processes forming web-like network, and the expression of the intermediate filaments nestin and vimentin. These cells were typical of active fibroblasts with a well-developed rough endoplasmic reticulum and close spatial correlation with collagen fibrils. Leptomeningeal PDGFR-β + cells ensheathing the vasculature in the subarachnoid space joined with pial PDGFR-β + cells upon entering the cortical parenchyma, yet perivascular PDGFR-β + cells in these penetrating vessels underwent abrupt changes in their morphological and molecular characteristics: they became more flattened with loss of immunoreactivity for nestin and vimentin and deficient collagen deposition, which was indicative of inactive fibroblasts termed fibrocytes. In the cortical parenchyma, PDGFR-β immunoreactivity was almost exclusively localized to larger caliber vessels, and significantly decreased in capillary-like microvessels. Collectively, our data identify PDGFR-β as a novel cellular marker for leptomeningeal fibroblasts comprising the leptomeninges and perivascular adventitial cells of the subarachnoid and penetrating large-sized cortical vasculatures.

Pain in trigeminal areas is driven by nociceptive trigeminal afferents. Transduction molecules, among them the nonspecific cation channels transient receptor potential vanilloid 1 (TRPV1) and ankyrin 1 (TRPA1), which are activated by endogenous and exogenous ligands, are expressed by a significant population of trigeminal nociceptors innervating meningeal tissues. Many of these nociceptors also contain vasoactive neuropeptides such as calcitonin gene-related peptide (CGRP) and substance P. Release of neuropeptides and other functional properties are frequently examined using the cell bodies of trigeminal neurons as models of their sensory endings. Pathophysiological conditions cause phosphorylation, increased expression and trafficking of transient receptor potential (TRP) channels, neuropeptides and other mediators, which accelerate activation of nociceptive pathways. Since nociceptor activation may be a significant pathophysiological mechanism involved in both peripheral and central sensitization of the trigeminal nociceptive pathway, its contribution to the pathophysiology of primary headaches is more than likely. Metabolic disorders and medication-induced painful states are frequently associated with TRP receptor activation and may increase the risk for primary headaches.

Spinal cord injury (SCI) is one of the most precarious conditions which have been one of the major reasons for continuous increasing mortality rate of SCI patients. Currently, there is no effective treatment modality for SCI patients posing major threat to the scientific and medical community. The available strategies don't mimic with the natural processes of nervous tissues repair/regeneration and majority of the approaches may induce the additional fibrotic or immunological response at the injury site and are not readily available on demand. To overcome these hurdles, we have developed a ready to use bioengineered human functional neurological construct (BHNC) for regenerative applications in SCI defects. We used cryopreserved meningeal tissues (CMT) for bioengineering these neurological constructs using acellularization and repopulation technology. The technology adopted herein generates intact neurological scaffolds from CMT and retains several crucial structural, biochemical and mechanical cues to enhance the regenerative mechanisms. The neurogenic differentiation on CMT scaffolds was almost similar to the freshly prepared meningeal scaffolds and mimics with the natural nervous tissue developmental mechanisms which offer intact 3D-microarchitecture and hospitable microenvironment enriched with several crucial neurotrophins for long-term cell survival and function. Functional assessment of developed BHNC showed highly increased positive staining for pre-synaptic granules of Synapsis-1 along with MAP-2 antibody with punctuate distribution in axonal regions of the neuronal cells which was well supported by the gene expression analysis of functional transcripts. Given the significant improvement in the field may enable to generate more such ready to use functional BHNC for wider applicability in SCI repair/regeneration., (Copyright © 2019 Elsevier B.V. All rights reserved.)

The identification of the "paucity of transportation vesicles" and "belt-like" tight junctions (TJs) of endothelial cells as the "morphological correlate of a blood-brain barrier" (BBB) by Reese and Karnovsky (J Cell Biol 34:207-217, 1967) has become textbook knowledge, and countless studies have helped to further define the elements, functions, and dynamics of the BBB. Most work, however, has focused on parenchymal capillaries or less clearly defined "microvessels", while a systematic study on similarities and differences between BBB architecture along the vascular tree within the brain and the meninges has been lacking. Since astrocytes induce endothelial cells to display BBB-typical characteristics by sonic hedgehog and Wnt/β-catenin signaling, we hypothesized that BBB-typical features should be most pronounced in parenchymal capillaries, where endothelium and astrocytes are separated by a basement membrane only. In contrast, this intimate contact is absent in leptomeningeal vessels, thereby potentially affecting BBB architecture. However, here, we show that claudin-3, claudin-5, zonula occludens-1, and occludin as typical constitutes of BBB TJs are comparably distributed in all segments of the parenchymal and the meningeal vascular tree of C57Bl6 mice. While electron microscopy revealed equally occluded interendothelial clefts, arterial vessels of the brain parenchyma but not within the meninges exhibited significantly longer TJ overlaps compared to capillaries. The highest density of endothelial vesicles was found in arterial vessels. Thus, endothelial expression of BBB-typical TJ proteins is not reflected by the distance to surrounding astrocytes, but electron microscopy reveals significant differences of endothelial specification along different segments of the CNS vasculature.

One major hallmark of Alzheimer's disease (AD) and cerebral amyloid angiopathy (CAA) is the deposition of extracellular senile plaques and vessel wall deposits composed of amyloid-beta (Aβ). In AD, degeneration of neurons is preceded by the formation of Aβ plaques, which show different morphological forms. Most of them are birefringent owing to the parallel arrangement of amyloid fibrils. Here, we present polarization sensitive optical coherence microscopy (PS-OCM) for imaging mature neuritic Aβ plaques based on their birefringent properties. Formalin-fixed, post-mortem brain samples of advanced stage AD patients were investigated. In several cortical brain regions, neuritic Aβ plaques were successfully visualized in tomographic and three-dimensional (3D) images. Cortical grey matter appeared polarization preserving, whereas neuritic plaques caused increased phase retardation. Consistent with the results from PS-OCM imaging, the 3D structure of senile Aβ plaques was computationally modelled for different illumination settings and plaque sizes. Furthermore, the birefringent properties of cortical and meningeal vessel walls in CAA were investigated in selected samples. Significantly increased birefringence was found in smaller vessels. Overall, these results provide evidence that PS-OCM is able to assess amyloidosis based on intrinsic birefringent properties.

We characterized cerebral Oil Red O-positive lipid-laden cells (LLC) of aging mice evaluating their distribution, morphology, density, functional activities and inflammatory phenotype. We identified LLC in meningeal, cortical and neurogenic brain regions. The density of cerebral LLC increased with age. LLC presenting small lipid droplets were visualized adjacent to blood vessels or deeper in the brain cortical and striatal parenchyma of aging mice. LLC with larger droplets were asymmetrically distributed in the cerebral ventricle walls, mainly located in the lateral wall. We also found that LLC in the subventricular region co-expressed beclin-1 or LC3, markers for autophagosome or autophagolysosome formation, and perilipin (PLIN), a lipid droplet-associated protein, suggesting lipophagic activity. Some cerebral LLC exhibited β galactosidase activity indicating a senescence phenotype. Moreover, we detected production of the pro-inflammatory cytokine TNF-α in cortical PLIN(+) LLC. Some cortical NeuN(+) neurons, GFAP(+) glia limitans astrocytes, Iba-1(+) microglia and S100β(+) ependymal cells expressed PLIN in the aging brain. Our findings suggest that cerebral LLC exhibit distinct cellular phenotypes and may participate in the age-associated neuroinflammatory processes.

It is well proved already that neurogenesis does take place in mammals' brain, including human brain. However, neurogenesis by itself is not able to compensate for brain tissue loss in serious neurological diseases, such as stroke, brain trauma or neurodegenerative disorders. Recent evidences show that neural stem cell niches are present not only in classical locations, such as subventricularor subgranular zones, but in other areas as well, including tissues contiguous to the brain (meninges and choroid plexus).In this chapter we revise the relationship of neural stem cells with interstitial cells (mainly telocytes), which we think is significant, and we describe what is known about the juxtacerebral tissue neurogenesis potential.

Seeding of amyloid fibrils into fresh solutions of the same peptide or protein in disaggregated form leads to the formation of replicate fibrils, with close structural similarity or identity to the original fibrillar seeds. Here we describe procedures for isolating fibrils composed mainly of β-amyloid (Aβ) from human brain and from leptomeninges, a source of cerebral blood vessels, for investigating Alzheimer's disease and cerebral amyloid angiopathy. We also describe methods for seeding isotopically labeled, disaggregated Aβ peptide solutions for study using solid-state NMR and other techniques. These methods should be applicable to other types of amyloid fibrils, to Aβ fibrils from mice or other species, tissues other than brain, and to some non-fibrillar aggregates. These procedures allow for the examination of authentic amyloid fibrils and other protein aggregates from biological tissues without the need for labeling the tissue.

The Fras1/Frem family of extracellular matrix proteins consists of Fras1 and its structurally related proteins, Frem1 (Fras1-related extracellular matrix protein 1), Frem2 and Frem3. These are co-localized in embryonic epithelial basement membranes (BMs), where they contribute to epithelial-mesenchymal adhesion. Although Fras1 localization pattern in epithelial BMs has been well defined, it has not yet been comprehensively studied in the central nervous system. Here, we demonstrate the immunohistochemical profile of Fras1 in the developing mouse brain and reveal an exclusively meningeal BM protein deposition. Interestingly, Fras1 displays a segmental localization pattern, which is restricted to certain regions of the meningeal BM. Frem2 protein displays a similar localization pattern, while Frem3 is rather uniformly distributed throughout the meningeal BM. Fras1 and Frem2 proteins are detected in regions of the BM that underlie organizing centers, such as the roof plate (RP) of diencephalon, midbrain and hindbrain, and the RP-derived structures of telencephalon (choroid plexus and hem). Organizing centers exert their activity via the production of bioactive molecules, which are potential Fras1 ligands. The restricted pattern of Fras1 and Frem2 proteins indicates a molecular compartmentalization of the meningeal BM that could reflect, yet unspecified, functional and structural differences.

Background: Clinical studies of β-amyloid (Aβ) immunotherapy in Alzheimer's disease (AD) patients have demonstrated reduction of central Aβ plaque by positron emission tomography (PET) imaging and the appearance of amyloid-related imaging abnormalities (ARIA). To better understand the relationship between ARIA and the pathophysiology of AD, we undertook a series of studies in PDAPP mice evaluating vascular alterations in the context of central Aβ pathology and after anti-Aβ immunotherapy., Methods: We analyzed PDAPP mice treated with either 3 mg/kg/week of 3D6, the murine form of bapineuzumab, or isotype control antibodies for periods ranging from 1 to 36 weeks and evaluated the vascular alterations in the context of Aβ pathology and after anti-Aβ immunotherapy. The number of mice in each treatment group ranged from 26 to 39 and a total of 345 animals were analyzed., Results: The central vasculature displayed morphological abnormalities associated with vascular Aβ deposits. Treatment with 3D6 antibody induced clearance of vascular Aβ that was spatially and temporally associated with a transient increase in microhemorrhage and in capillary Aβ deposition. Microhemorrhage resolved over a time period that was associated with a recovery of vascular morphology and a decrease in capillary Aβ accumulation., Conclusions: These data suggest that vascular leakage events, such as microhemorrhage, may be related to the removal of vascular Aβ. With continued treatment, this initial susceptibility period is followed by restoration of vascular morphology and reduced vulnerability to further vascular leakage events. The data collectively suggested a vascular amyloid clearance model of ARIA, which accounts for the currently known risk factors for the incidence of ARIA in clinical studies., (Copyright © 2013. Published by Elsevier Inc.)

Headaches can be evoked by activation of meningeal nociceptors, but an involvement of pericranial tissues is debated. We aimed to examine a possible extracranial innervation by meningeal afferents in the rat. For in vivo neuronal tracing, dextran amines were applied to the periosteum underlying the temporal muscle. Labeling was observed 2 days later in the parietal dura mater, trigeminal ganglion, and spinal trigeminal nucleus with confocal and electron microscopy. In the hemisected rat head, extracellular recordings were made from meningeal nerve fibers. Release of calcitonin gene-related peptide (CGRP) from the cranial dura mater during noxious stimulation of pericranial muscles was quantified. In vivo capsaicin was injected into the temporal muscle while meningeal blood flow was recorded. In the parietal dura mater, labeled C- and Aδ fibers ramified extensively, accompanied the middle meningeal artery, and passed through the spinosus nerve into the maxillary and mandibular, but not the ophthalmic division of the trigeminal ganglion. Some fibers could be traced into the ipsilateral spinal trigeminal nucleus. Electrophysiological recordings revealed afferent fibers with mechanosensitive receptive fields both in the dura mater and in the parietal periosteum. Noxious stimulation of the temporal muscle caused CGRP release from the dura mater and elevated meningeal blood flow. Collaterals of meningeal nerve fibers project through the skull, forming functional connections between extra- and intracranial tissues. This finding offers a new explanation of how noxious stimulation of pericranial tissues can directly influence meningeal nociception associated with headache generation and why manual therapies of pericranial muscles may be useful in headaches., (Copyright © 2013 International Association for the Study of Pain. Published by Elsevier B.V. All rights reserved.)

Meninges, the connective tissue of the vertebrate central nervous system (CNS), have not been recognized in invertebrates. We describe the ultrastructure of the adult brain, antennules, and cord in five marine copepods: Calanus finmarchicus, Gaussia princeps, Bestiolina similis, Labidocera madurae, and Euchaeta rimana. In all of these locations we identified cell types with characteristics of the typical cells of vertebrate meninges and of their peripheral nervous system (PNS) connective tissue counterpart: fibroblasts, having flattened twisting processes with labyrinthine cavities communicating with the extracellular space, and macrophages, containing prominent lysosomes, well-developed endoplasmic reticulum, Golgi apparatus, and indented heterochromatin. The vertebrate distinction between electron-dense cells in the most external connective tissues (dura mater and epineurium) versus electron-lucent cells in the more internal connective tissues (pia-arachnoid and endoneurium-perineurium) was also found in the copepod CNS and PNS. Similar to the vertebrate organization, electron-dense cell networks penetrated from the outer layer (subcuticle) to surround inner substructures of the copepod nervous systems, and electron-lucent networks penetrated deeply from the brain and nerve surfaces to form intertwined associations with neural cells. Moreover, the association of these cells with basement membranes, glycocalyx, and fibrils of collagen in copepods conforms to a meningeal organization. The primary deviation from the vertebrate ultrastructural organization was the often tight investment of axons by the meningeal-like cells, with an intercalated basement membrane. Together, these data suggest that the tissues investing the copepod nervous system possess an organization that is analogous in many respects to that of vertebrate meninges., (Copyright © 2012 Wiley Periodicals, Inc.)

Background: Newts have the remarkable ability to regenerate their spinal cords as adults. Their spinal cords regenerate with the regenerating tail after tail amputation, as well as after a gap-inducing spinal cord injury (SCI), such as a complete transection. While most studies on newt spinal cord regeneration have focused on events occurring after tail amputation, less attention has been given to events occurring after an SCI, a context that is more relevant to human SCI. Our goal was to use modern labeling and imaging techniques to observe axons regenerating across a complete transection injury and determine how cells and the extracellular matrix in the injury site might contribute to the regenerative process., Results: We identify stages of axon regeneration following a spinal cord transection and find that axon regrowth across the lesion appears to be enabled, in part, because meningeal cells and glia form a permissive environment for axon regeneration. Meningeal and endothelial cells regenerate into the lesion first and are associated with a loose extracellular matrix that allows axon growth cone migration. This matrix, paradoxically, consists of both permissive and inhibitory proteins. Axons grow into the injury site next and are closely associated with meningeal cells and glial processes extending from cell bodies surrounding the central canal. Later, ependymal tubes lined with glia extend into the lesion as well. Finally, the meningeal cells, axons, and glia move as a unit to close the gap in the spinal cord. After crossing the injury site, axons travel through white matter to reach synaptic targets, and though ascending axons regenerate, sensory axons do not appear to be among them. This entire regenerative process occurs even in the presence of an inflammatory response., Conclusions: These data reveal, in detail, the cellular and extracellular events that occur during newt spinal cord regeneration after a transection injury and uncover an important role for meningeal and glial cells in facilitating axon regeneration. Given that these cell types interact to form inhibitory barriers in mammals, identifying the mechanisms underlying their permissive behaviors in the newt will provide new insights for improving spinal cord regeneration in mammals.

Primary malignant tumours arising from the meninges are distinctly uncommon, and when they occur, they are usually sarcomas. In contrast, metastatic meningeal involvement is increasingly seen as advances in cancer therapy have changed the natural history of malignant disease and prolonged the life span of cancer patients. The meninges can either be infiltrated by contiguous extension of primary tumours of the central nervous system, paranasal sinuses and skull base origin or can be diffusely infiltrated from haematogenous dissemination from distant primary malignancies. Imaging in these patients provides crucial information in planning management. This article reviews the pertinent anatomy that underlies imaging findings, discusses the mechanism of meningeal metastasis and highlights different imaging patterns of meningeal carcinomatosis and the pitfalls.

Stroke remains a major health problem worldwide, and is the leading cause of serious long-term disability. Recent findings now suggest that strategies to enhance angiogenesis after focal cerebral ischemia may provide unique opportunities to improve clinical outcomes during stroke recovery. In this mini-review, we survey emerging mechanisms and potential targets for angiogenic therapies in brain after stroke. Multiple elements may be involved, including growth factors, adhesion molecules and progenitor cells. Furthermore, cross talk between angiogenesis and neurogenesis may also provide additional substrates for plasticity and remodeling in the recovering brain. A better understanding of the molecular interplay between all these complex pathways may lead to novel therapeutic avenues for tackling this difficult disease.

Objective: To evaluate leptomeningeal and subpial inflammatory responses of experimental Staphylococcus aureus bacteriemia following intraperitoneal and intravenous applications and to compare the inflammatory reactions in different regions of central nervous system., Material and Methods: Forty anesthetized rats were divided into four groups equal in number. The rats in group-I were given 1 ml suspension of Staphylococcus aureus intraperitoneally. Group-II was the control group of group I; it was administrated 1 ml 0.9% NaCl in water intraperitoneally. The rats in group-III were given the same amount of bacteria intravenously. Group IV was the control group of the group-III; it was administrated 1 ml 0.9% NaCl solution intravenously. The rats were sacrificed on the 21st day. Inflammatory changes of different regions of the central nervous system were examined under transmission electron microscopy. Statistical analysis was done by using variance analysis, Bonferroni, Tamhane post hoc, Student's t and univariate tests., Results: Thoracic and occipital regions were the most vulnerable zones. Increasing of collagen tissue was the most detected inflammatory change., Conclusion: This experimental model can be used for inducing subpial and leptomeningeal inflammations and it may be developed for investigations of pathogenesis of leptomeningitis during systemic infections.

Immunohistochemical and electron microscopic studies were carried out to elucidate the structure of the serotonergic neuron system in the spinal cord of the spotted gar, Lepisosteus oculatus, a nonteleost actinopterygian. Serotonin-immunoreactive (5HT-IR) cell bodies and fibers were widely distributed in the spinal cord, constituting an intrinsic neuron system. This system comprised three anatomical cell groups in different portions of the spinal cord, i.e., the rostromedial cell group, the paired ventrolateral cell groups, and the ventral superficial cell group. The rostromedial cell group included cerebrospinal fluid-contacting neurons with intraventricular processes. The immunostained fibers projecting from all three of these cell groups ran in various directions, mainly ventrally and ventrolaterally, and partly gave rise to a dense plexus at the ventrolateral surface of the spinal cord. Immunoelectron microscopy of the relevant portion demonstrated many varicose fibers containing 5HT-immunopositive vesicles. Conventional electron microscopy of the plexus showed that the constituent varicose fibers were unmyelinated and frequently made a direct contact with the basement membrane contiguous to the leptomeniges (meninx primitiva). There, exocytotic figures of cytoplasmic vesicles were demonstrated, suggesting that 5HT may be secreted, in a paracrine way, into the extraspinal space. This specialized area in the gar spinal cord may be referred to as the juxtameningeal serotonergic plexus.

We present a case of squamous spindle cell carcinoma of the external auditory meatus in a 38-year-old man. The tumour was extended to the inner ear, the temporal bone, the middle cranial fossa and the meningo-cerebral tissue. The surgical intervention of temporo-occipital craniotomy removed most of the neoplasia. At pathologic examination, the tumour showed an undifferentiated spindle cell pattern. Immunohistochemistry with a large antibody panel found a weak positivity only to EMA. The diagnosis was made when the electron microscopy showed rare junctional structures and tonofilaments.

Background: The safety of ketamine when administered by the spinal route must be confirmed in various animal species before it is approved for use in humans. This study evaluates the ultrastructure of canine meninges after repeated doses of epidural S(+)-ketamine., Methods: Five dogs received S(+)-ketamine 5%, 1 mg/kg, twice a day for 10 days through an epidural catheter with its tip located at the L5 level. One dog received the same volume of normal saline at the same times. The spinal cord and meninges were processed for histopathological and ultrastructural studies. Clinical effects were assessed after each injection., Results: Motor and sensory block appeared after each injection of S(+)-ketamine, but not in the dog receiving saline. No signs of clinical or neurologic alterations were observed. Using light microscopy, no meningeal layer showed alterations except focal infiltration at the catheter tip level by macrophages, lymphocytes, and a few mast cells. The cells of different layers were studied by electron microscopy and interpreted according to data from human and other animal species because no ultrastructural description of the canine meninges is currently available. There were no cellular signs of inflammation, phagocytosis, or degeneration in meningeal layers and no signs of atrophy, compression, or demyelinization in the areas of dorsal root ganglia and spinal cord around the arachnoid. These findings were common for dogs receiving S(+)-ketamine and the dog receiving saline., Conclusion: Repeated doses of epidural S(+)-ketamine 5%, 1 mg/kg, twice a day for 10 days was not associated to cellular alterations in canine meninges.

Communicating hydrocephalus may occur spontaneously in elderly patients or occur as a complication of meningitis or intracranial hemorrhage, typically as a result of fibrosis along the route of cerebrospinal fluid (CSF) flow. Hepatocyte growth factor (HGF) has anti-fibrotic properties and is a promising candidate for the treatment of various fibrotic diseases. Thus, the goal of this study was to examine the effect of exogenous HGF (30 microg of human recombinant (hr) HGF intraventricularly for 7 or 14 days) in a model of hr transforming growth factor beta1-induced communicating hydrocephalus in C57BL/6 mice. HGF treatment resulted in a reduction of ventriculomegaly, as demonstrated by magnetic resonance imaging, and improved spatial memory. Further, ink passage test demonstrated improvement of normalized CSF in flow in mice receiving HGF treatment as opposed to delayed CSF flow in the hydrocephalic mice at baseline. Finally, histological examination in hydrocephalic mice undergoing HGF treatment revealed reduction of collagen fibers in the meninges and normalization of their structures. These results indicate that exogenous HGF may be of utility in the treatment of hydrocephalus in humans.

The meninges comprise the dura mater and the leptomeninges (arachnoid and pia mater). Dura forms an outer endosteal layer related to the bones of the skull and spine and an inner layer closely applied to the arachnoid mater. Leptomeninges have multiple functions and anatomical relationships. The outer parietal layer of arachnoid is impermeable to CSF due to tight intercellular junctions; elsewhere leptomeningeal cells form demosomes and gap junctions. Trabeculae of leptomeninges compartmentalize the subarachnoid space and join the pia to arachnoid mater. In bacterial meningitis leptomeningeal cells secrete cytokines. Pia mater is reflected from the surface of the brain and spinal cord onto arteries and veins, thus separating the subarachnoid space from the brain and cord. A sheath of leptomeninges accompanies arteries into the brain and is related to the pathways for the drainage of interstitial fluid that play a role in inflammatory responses in the brain and appear to be blocked by amyloid-beta in Alzheimer's disease. Specialised leptomeningeal cells in the stroma of the choroid plexus form collagen whorls that become calcified with age. Leptomeningeal cells also form channels in the core and apical cap of arachnoid granulations for the drainage of CSF into venous sinuses. In the spine, leptomeninges form highly perforated intermediate sheets of arachnoid and delicate ligaments that compartmentalize the subarachnoid space; dentate ligaments anchor subpial collagen to the dura mater and stabilize the spinal cord. Despite the multiple anatomical arrangements and physiological functions, leptomeningeal cells retain many histological features that are similar from site to site.

Unlabelled: We examined ultrastructural details such as the cellular component and membrane thickness of human spinal pia mater with the aim of determining whether fenestrations are present. We hypothesized that pia mater is not a continuous membrane but, instead, that there are fenestrations across the pial cellular membrane. The lumbar dural sac from 7 fresh human cadavers was removed, and samples from lumbar spinal pia mater were studied by special staining techniques, immunohistochemistry, and transmission and scanning electron microscopy. A pial layer made by flat overlapping cells and subpial tissue was identified. We found fenestrations in samples from human spinal pia mater at the thoracic-lumbar junction, conus medullaris, and nerve root levels, but these fenestrations did not appear at the thoracic level. We speculate whether the presence of fenestrations in human spinal pia mater at the level of the lumbar spinal cord and at the nerve root levels has any influence on the transfer of local anesthetics across this membrane., Implications: The ultrastructural anatomy of the human pia mater, such as pial cells, membrane thickness, and subpial tissue at different levels of the thoracic and lumbar spinal cord and nerve roots, was studied by special staining techniques, immunohistochemistry, and transmission and scanning electron microscopy. Fenestrations were found in samples at the thoracic-lumbar junction, conus medullaris, and nerve root levels. No fenestrations were found in samples at the thoracic level. At present, we cannot determine the significance of these findings.

The presence of sodium-potassium-adenosine triphosphatase (Na+,K+-ATPase) on the surface of arachnoid cells indicates that active transport of electrolytes and water occurs there. Previously, we accidentally found that intraventricular administration of TGF-beta1 impaired rat spatial learning. Levels of Na+,K+ -ATPase were decreased in arachnoid cells with fibrosis. To characterize the role of the Na+,K+ -ATPase, Wistar rats were intraventricularly administered a total of 200 microl of ouabain, at concentrations of 10(-5), 10(-4) and 10(-3) M, for one week with an osmotic pump, and were examined with a Morris water maze. Latency for reaching the platform did not significantly differ between ouabain-administered rats and controls. Spatial learning was impaired in a dose-dependent manner. Na+,K+ -ATPase activity of arachnoid cells ceased during ouabain administration, and recovered completely three weeks after the end of ouabain administration. The present results suggest that the Na+,K+ -ATPase on the surface of arachnoid cells contributes to maintenance of rat spatial learning.

The present investigation provides novel information on the topographical distribution of macrophages and dendritic cells (DCs) in normal meninges and choroid plexus of the rat central nervous system (CNS). Whole-mounts of meninges and choroid plexus of Lewis rats were incubated with various anti-leucocyte monoclonal antibodies and either visualised with gold-conjugated secondary antibody followed by silver enhancement and subsequent examination by environmental scanning electron microscopy or by the use of fluorochromes and confocal microscopy. Large numbers of MHC class II(+) putative DCs were identified on the internal or subarachnoid aspect of dural whole-mounts, on the surface of the cortex (pia/arachnoid) and on the surface of the choroid plexus. Occupation of these sites would allow DCs access to cerebrospinal fluid (CSF) and therefore allow antigens into the subarachnoid space and ventricles. By contrast, macrophages were less evident at sites exposed to CSF and were more frequently located within the connective tissue of the dura/arachnoid and choroid plexus stroma and also in a sub-pial location. The present data suggest that DC may be strategically located within the CNS to sample CSF-borne antigens. Furthermore, the data suggest that CNS tissue samples collected without careful removal of the meninges may inadvertently be contaminated by DCs and meningeal macrophages.

The vascular architecture of the human cerebellar meninges was investigated. The surface meninges were poor in vasculature. In the sulci, the meninges were highly vascular but had few capillaries. The venous blood vessels gave long side branches at right angles to the parent vessels in a cruciform pattern, running horizontally along the cerebellar sulci. They were situated at the origin of the secondary or tertiary sulci. Anastomoses between these horizontal branches gave a crosshatched appearance. Short branches often extended to the bases of the sulci, terminating in T-shaped bifurcations with numerous tiny branches, like the roots of a tree. The arteries ran perpendicular to venous branches which were parallel to each other exclusively along the sagittal plane. These arteries bifurcated to straddle the horizontally running veins at the origin of the secondary or tertiary sulci. They gave off many small branches like teeth of a fork from each artery in the secondary or tertiary sulci after they bifurcated to straddle the venous branches and penetrated the cerebellar cortex at the bases of sulci. These fork-like ramifications in the bases of the sulci were most likely responsible for the ready development of pronounced ischemic state. They might also play an important role in the occurrence of ischemic damage at the bases of sulci in cases of severe generalized ischemia.

The lumbosacral spinal cord of birds is unique among vertebrates in that segmentally organized accessory lobes protrude from the ventrolateral spinal cord into the vertebral canal. Recently, it has been suggested that these lobes may be part of an extralabyrinthine sense organ of equilibrium. For a better understanding of such a function, a complete analysis of the structural basis was performed by means of electron microscopy. The lobes consist of multipolar neurons, myelinated and unmyelinated axons, glia-derived glycogen cells, glial cells, and capillaries. The dorsal part of the lobe is covered by a loose mesh of pia mater. Ventrolaterally, an arachnoidal trabecle is in close contact with the lobe. Extracellular lacunae extend from the periphery deep into each lobe. The lacunae are separated from the subarachnoidal space by a loose mesh of processes of the glycogen cells with its basal lamina. The lacunae are filled by a network of processes of glycogen cells, glial cell, dendrites, and small axons. Both neuronal somata and dendrites are contacted by numerous axon terminals that form rather uniform synapses. Finger-like processes emerge from both the somata and the dendrites. The dendrites branch deeply into the extracellular lacunae and form lateral ramifications, which consist of narrow stalks with serially arranged bulbous portions, from which finger-like processes emerge. Finger-like processes are well-known elements in mechanotransduction. Glycogen cells and lacunae may contribute to transmission of hydrostatic pressure changes during movements of the body., (Copyright 2002 Wiley-Liss, Inc.)

Unlabelled: Previous studies of samples from cranial meninges have created doubts about the existence of a virtual subdural space. We examined the ultrastructure of spinal meninges from three human cadavers immediately after death to see whether there is a virtual subdural space at this level. The arachnoid mater had two portions: a compact laminar portion covering the dural sac internal surface and a trabecular portion extending like a spider web around the pia mater. There was a cellular interface between the laminar arachnoid and the internal layer of the dura that we called the dura-arachnoid interface. There was no subdural space in those specimens where the dura mater was macroscopically in continuity with the arachnoid trabecules. In the specimens where the dura mater was separated from the arachnoid, we found fissures in between the neurothelial cells that extended throughout the interface. We hypothesize that the subdural space would have its origin within the dura-arachnoid interface when the neurothelial cells break up, creating in this way a real subdural space., Implications: The subdural space was not seen under transmission electron microscopy in samples of human spinal meninges where surgical manipulation was avoided. Scanning electron microscopy in other samples showed the presence of broken neurothelial cells giving up fissures that extended along the dura-arachnoid interface. These findings may explain the origin of a real subdural space.

The connexin family of proteins (Cx) that form intercellular gap junctions in vertebrates is well represented in the mammalian central nervous system. Among these, Cx30 and Cx43 are present in gap junctions of astrocytes. Cx32 is expressed by oligodendrocytes and is present in heterologous gap junctions between oligodendrocytes and astrocytes as well as at autologous gap junctions between successive myelin layers. Cx36 mRNA has been identified in neurons, and Cx36 protein has been localized at ultrastructurally defined interneuronal gap junctions. Cx26 is also expressed in the CNS, primarily in the leptomeningeal linings, but is also reported in astrocytes and in neurons of developing brain and spinal cord. To establish further the regional, cellular, and subcellular localization of Cx26 in neural tissue, we investigated this connexin in adult mouse brain and in rat brain and spinal cord using biochemical and immunocytochemical methods. Northern blotting, western blotting, and immunofluorescence studies indicated widespread and heterogeneous Cx26 expression in numerous subcortical areas of both species. By confocal microscopy, Cx26 was colocalized with both Cx30 and Cx43 in leptomeninges as well as along blood vessels in cortical and subcortical structures. It was also localized at the surface of oligodendrocyte cell bodies, where it was coassociated with Cx32. Freeze-fracture replica immunogold labeling (FRIL) demonstrated Cx26 in most gap junctions between cells of the pia mater by postnatal day 4. By postnatal day 18 and thereafter, Cx26 was present at gap junctions between astrocytes and in the astrocyte side of most gap junctions between astrocytes and oligodendrocytes. In perinatal spinal cord and in five regions of adult brain and spinal cord examined by FRIL, no evidence was obtained for the presence of Cx26 in neuronal gap junctions. In addition to its established localization in leptomeningeal gap junctions, these results identify Cx26 as a third connexin (together with Cx30 and Cx43) within astrocytic gap junctions and suggest a further level of complexity to the heterotypic connexin channel combinations formed at these junctions., (Copyright 2001 Wiley-Liss, Inc.)

Pachymeninx and leptomeninx of cranial cavity and spine are considerably different in their collagenous fiber texture, cellular composition, vascularization, and innervation. The majority of meningeal nerve fibers terminate as free nerve endings whereas encapsulated and lamellated nerve terminals additionally occur in higher vertebrates including man. With respect to nerve fiber classification, arborization pattern, topography, and organization of the microenvironment at the termination site afferent and efferent nerve terminals are differentiated. Only the dura mater and the pial subcompartment of the leptomeninx possess the morphological prerequisites for neurogenic inflammation. In the current review, the results of morphological studies regarding the meningeal innervation including the sites of CSF (cerebrospinal fluid) production and absorption are discussed with emphasis on their structure-function relationships., (Copyright 2001 Wiley-Liss, Inc.)

We applied high-resolution laser-scanning microscopy, electron microscopy, and non-radioactive in situ hybridization histochemistry to determine the cellular and intracellular localization of lipocalin-type prostaglandin D synthase, the major brain-derived protein component of cerebrospinal fluid, and its mRNA in leptomeninges, choroid plexus, and parenchyma of the adult rat brain. Both immunoreactivity and mRNA for prostaglandin D synthase were located in arachnoid barrier cells, arachnoid trabecular cells, and arachnoid pia mater cells. Furthermore, meningeal macrophages and perivascular microglial cells, identified by use of ED2 antibody, were immunopositive for prostaglandin D synthase. In the arachnoid trabecular cells, the immunoreactivity for prostaglandin D synthase was located in the nuclear envelope, Golgi apparatus, and secretory vesicles, indicating the active production and secretion of prostaglandin D synthase. In the meningeal macrophages, prostaglandin D synthase was not found around the nucleus but in lysosomes in the cytoplasm, pointing to an uptake of the protein from the cerebrospinal fluid. Furthermore, the existence of meningeal cyclooxygenase (COX) -1 and COX-2 was investigated by Western blot, Northern blot, and reverse transcriptase-polymerase chain reaction (RT-PCR), and the colocalization of COX-2 and prostaglandin D synthase was demonstrated in virtually all cells of the leptomeninges, choroid plexus epithelial cells, and perivascular microglial cells, suggesting that these cells synthesize prostaglandin D(2) actively. Alternatively, oligodendrocytes showed prostaglandin D synthase immunoreactivity without detectable COX-2. The localization of lipocalin-type prostaglandin D synthase in meningeal cells and its colocalization with COX-2 provide evidence for its function as a prostaglandin D(2)-producing enzyme., (Copyright 2000 Wiley-Liss, Inc.)

To our knowledge, five cases of Castleman's disease involving only the central nervous system have been reported previously. We report a sixth case, which occurred in a 47-year-old woman with a 3-month history of headaches and a large superior frontal lobe mass on neuroimaging. Excisional biopsy revealed confluent lymphoid nodular areas with multiple well-developed germinal centers surrounded by concentrically layered proliferations of small B lymphocytes typical of Castleman's disease. Ultrastructural study found 100-nm virallike particles within follicular dendritic cells as well as intercellular spaces. These particles were suggestive of a D-type retrovirus. The patient underwent postoperative radiotherapy and was neurologically normal 3 months after surgery.

Previously, immunohistochemical methods were primarily used to detect and provide indirect evidence on the composition of Lewy bodies, the pathological hallmark of Parkinson's disease. This was chiefly because there are very few procedures that describe the isolation of these structures. We report here a relatively simple method that we have developed for the exclusive isolation of Lewy bodies from brain tissue. The isolation of the Lewy bodies and subsequent evaluation of their components may furnish an insight into their role in the neurodegenerative mechanism(s) operating in the spectrum of Lewy body disorders.

Unlabelled: Although there are various published descriptions of the dura mater spinalis [4, 7, 9, 11, 16], some points relating to the texture of the collagen fibres in the dura have still not been adequately explained. In this study the orientation of the collagen fibril bundles was revealed with the aid of scanning electron microscopy, and our observations have yielded new insights into the three-dimensional structure of the human dura mater spinalis., Materials and Methods: The preparations used were taken from the bodies of four persons who had died of acute cardiac infarct at the ages of 70-78 years. The histories of these patients gave no indications of earlier neurological, endocrine or septic illnesses. The tissue examined was taken 8-12 h after death in all cases; it was immediately fixed in glutaraldehyde and then processed for scanning electron microscopy in the usual way., Results: In the outermost (epidural) layer of the dura mater spinalis the collagen fibres are bunched together in bands that run in all directions. Elastic fibres 2 mm thick are woven into this three-dimensional network of collagen systems. On the inside (the arachnoid side) thin collagen fibres are fused into layers in such a way that the innermost layer resting on the arachnoid has a smooth, shiny appearance comparable to that of a serosa. It is attached to the actual dura with a supporting band of connective tissue. Rests of the subdural neuroepithelium could contribute to the smooth appearance of the superficial aspect., Conclusions: The outermost layer of the dura is made up mainly of collagen fibres, which run in all three directions--longitudinal, horizontal and transverse--both singly and in groups. These findings are at odds with "classic" descriptions, according to which the fibres in the dura mater spinalis all have a parallel course with a longitudinal orientation in tangential sections.

There is evidence for lymphatic drainage of interstitial fluid from the brain along perivascular spaces in a number of mammalian species. Ultrastructural studies suggest that there are similar drainage pathways in the human cerebral cortex. Perivascular spaces in the basal ganglia, however, differ from those in the cortex in that they dilate to form lacunes and rarely accumulate beta-amyloid (amyloid angiopathy) in Alzheimer's disease; in the cortex, lacunes are rare but amyloid angiopathy is common. The aim of the present study is to compare the structure of perivascular spaces in the basal ganglia and at the anterior perforated substance with perivascular spaces in the cerebral cortex. Eight postmortem brains from patients aged 23-80 years (mean 68 y) were examined by light microscopy, by scanning and transmission electron microscopy and by direct visualisation of etched paraffin blocks. The results show that arteries in the basal ganglia are surrounded by 2 distinct coats of leptomeninges separated by a perivascular space which is continuous with the perivascular space around arteries in the subarachnoid space. The inner layer of leptomeninges closely invests the adventitia of the vessel wall and the outer layer is continuous with the pia mater on the surface of the brain at the anterior perforated substance. Veins in the basal ganglia have no outer layer of leptomeninges and thus the perivascular space is continuous with the subpial space. The anatomy of the periarterial spaces in the basal ganglia differs significantly from that in the cerebral cortex where there is only a single periarterial layer of leptomeninges. Differences in structure of perivascular spaces around arteries may reflect relative efficiencies in the drainage of interstitial fluid from different sites in the brain. Furthermore, the structure of the perivascular spaces may contribute to the relatively high frequency of lacunes in the basal ganglia, and the low frequency of amyloid angiopathy at this site in Alzheimer's disease.

Purpose: The intraorbital optic nerve sheath meninges contain a perineural subarachnoid space lined by meningeal cell layers and intercellular fibrous tissue. We sought to determine whether functional or structural characteristics, or both, of the optic nerve sheath are influenced by the increased intracranial pressure after the rupture of cerebral aneurysms., Methods: We infused the great cisterns of cats with either x-ray contrast medium or autologous blood. The cisternal infusions were done under the experimental condition of a sudden 2.5-minute increase in intracranial pressure similar to that recorded after the rupture of cerebral aneurysms in humans., Results: Digital subtraction radiographs of the optic nerves taken during the cisternal infusion of contrast medium at the start showed the opacification of the optic nerve subarachnoid space. After 2 minutes, the contrast medium leaked into the orbit, indicating the breakdown of the meningeal fluid barrier. Ultrastructural investigation of the optic nerve sheath after high-pressure cisternal infusions showed the arachnoid cell layers scattered. The flattened arachnoid cells displayed mainly intracellular and some intercellular, porelike openings. After infusion of blood into the great cistern, erythrocytes were found within porelike openings of the arachnoid cells., Conclusions: The meningeal fluid barrier of the optic nerve sheath can be destroyed by pressure changes associated with subarachnoid hemorrhage. This disruption might be regarded as a natural optic nerve sheath fenestration that allows outflow of cerebrospinal fluid into the orbit to protect the optic nerve from increased intracranial pressure after aneurysmal rupture.

The mucopolysaccharidoses (MPS) are lysosomal storage diseases in which a specific enzyme defect causes glycosaminoglycans storage in tissues. The authors present a necropsy case of a 10 years old boy with clinical and laboratorial diagnosis of MPS. The necropsy revealed thickening of meninges, cardiac valves and hepatomegaly. The microscopical examination of the brain showed finely vacuolated histiocytes around blood vessels and meninges. Systemic deposits of vacuolated histiocytes in cardiac valves and liver were also detected. The ultrastructural examination of the brain, liver and spleen showed filamentous material accumulated in vacuolated histiocytes and hepatocytes and features neuronal storage disease.

The outer blood-cerebrospinal fluid barrier is formed by leptomeningeal cells of the arachnoidea. The structures underlying this barrier are tight junctions. In contrast to the tight junctions of endo- and epithelial cells, which have been investigated by means of ultrastructural as well as by molecular methodology, equivalent studies on meningeal cells are lacking. In the present study, therefore, the ultrathin section and freeze-fracture morphology of cranial leptomeningeal cells of carp, frog, chicken and rat was investigated by quantitative morphometry. In addition, the developmental features of the meningeal barrier in chicken and rat were compared. The parameters determined were the complexity of the tight junctions, the density of strands and branchpoints and the degree of association of tight junction particles with the protoplasmatic and exoplasmatic leaflets of cellular membranes. The complexity of tight junctions was highest in chicken and lowest in frog meningeal cells, whereas intermediate values were reached in the carp and the rat. E- and P-face associations were similar in carp and frog, whereas in chicken, the P-face association and in rat the E-face association dominated. During development, tight junction complexity continuously increased up to adult stages in the chicken, whereas in the rat, the adult value was already reached at postnatal day 2. At early embryonic stages, particle insertion into tight junctions was incomplete but occurred equally into both membranous leaflets; if completed at E19 in the chicken, a redistribution of particles toward a higher P-face association was observed. In the rat, tight junction particles were redistributed toward a higher E-face association. These results are discussed in the context of blood-brain barrier induction, maintenance and regulation.

Human spinal dura and arachnoid, obtained during neurosurgical operations, were studied by transmission electron microscopy. The ultrastructure of spinal meninges largely conformed to the morphology of the cranial meninges, but some minor differences were detected. The dura was composed of an outermost loosely arranged fibroelastic layer, a middle basically fibrous portion and an innermost cellular layer (dural border cell layer). The dural border cell layer was characterised by multiple interdigitating cell processes, no extracellular collagen, significant extracellular spaces and few cell junctions. Paravascular vesiculated nerve profiles were encountered within the fibroadipose epidural tissue. The arachnoid was composed of an outermost portion (arachnoid barrier cell layer), presenting tightly packed cells, numerous tight junctions and no extracellular collagen. In view of its numerous tight junctions, the arachnoid barrier cell layer is considered to represent an effective morphological and physiological meningeal barrier between the cerebrospinal fluid in the subarachnoid space and the blood circulation in the dura. The arachnoid barrier layer was always characterised by a distinct continuous basal lamina on its inner surface towards the innermost collagenous portion of the arachnoid (arachnoid reticular cell layer). The interweaving arachnoid trabecular cells within this layer possessed numerous mitochondria and were anchored to the inner surface of the arachnoid barrier cell layer by desmosomes. An additional layer of flattened branching cells was demonstrated along the inner surface of the arachnoid reticular cell layer and assumed to be an "arachnoid border cell layer'. Morphological data suggest that the dura and arachnoid closely adhere at spinal levels in man without any naturally occurring "subdural space'. However, structurally, the dural border cell layer forms a weak cell layer at the dura-arachnoid continuum that is easily disrupted. The creation of an artifactual subdural space at spinal levels is discussed.

The formation of the meninges around the new spinal cord of the regenerating tail of the lizards Anolis and Lampropholis has been studied after injection of 3H-thymidine and 3H-proline. An optical and electron microscopic autoradiographical study revealed that many irregular 3H-thymidine labeled meningeal fibroblasts surround the apical regenerating spinal cord. Meningeal fibroblasts become flat at about 0.5-1 mm from the tail tip while they actively uptake 3H-proline involved in the synthesis of collagen and elastin. Elastic fibres are in intimate contact with the meningeal fibroblasts. One hour after injection of 3H-proline trace grains were seen most in the Golgi apparatus and secretory elastin-like vesicles. The importance of the elastic fibrils around the caudal spinal cord of lizard in relation to the body movements is discussed.

The glial-limiting membrane at the border of the central nervous system (CNS) consists of glial endfeet covered by a basal lamina. The formation of the glia limitans seems to be controlled by adjacent meninges but only little is known about this interaction. In the present study astrocytes and meningeal cells were investigated in vitro to see if cocultures of these cells can serve as a suitable model for the differentiation of the glial-limiting membrane and can be used to define the conditions under which the glial-limiting membrane develops. The following observations were made in cocultures of meningeal and astrocytic cells of two-day-old rats: (i) epithelioid astrocytes were transformed into stellate cells; (ii) single colonies of proliferating epithelioid astrocytes were generated; (iii) the area around these colonies becomes devoid of meningeal cells, which seem to form a circular border around the astroglial islands; (iv) from the glial colonies long thin glial processes grow towards the surrounding meningeal cells, terminating at the site of contact; (v) in the contact zone between meningeal cells and astrocytes irregular shaped deposits of electron dense material resembling a basal lamina were seen. These observations indicate that indeed a structure resembling a glial-limiting membrane develops in cocultures of meningeal and astrocytic cells. Its formation depends on the balance of growth promoting effects of meningeal cells on astrocytes and growth inhibiting effects of astrocytes on meningeal cells. Both activities can be enriched from conditioned media of pure astrocytic or meningeal cell culture. The proposed model of meningo-astrocytic cocultures may be a helpful instrument for further investigations on the formation of the glia limitans.

In the optic tectum of goldfish, the outer, middle and inner layers of the endomeninx were evident in animals ranging in age from 1 month to several years. The outer layer in young animals consisted of closely overlapping cells with intertwined processes, whereas in the older animals it contained large extracellular spaces. The intermediate layer cells were always arranged in a single continuous layer, but in young animals they overlapped extensively with one another toward their edges whereas in the oldest animals they became extremely flat and non-overlapping. The inner layer included an outer tier of cells with their bases adhering to the intermediate layer, and an inner tier of cells detached from both the intermediate layer and the basal lamina overlying the brain parenchyma. Inner layer cells contained many large vacuoles that were in continuity with the extracellular space. With age, the extracellular space and the vacuolar system expanded, and the inner layer evolved into a meshwork of attenuated cytoplasmic processes embedded in the granular extracellular matrix. Another age-related feature was the accumulation adjacent to the basal lamina of uniform disc-shaped membranous structures, resembling multilamellar bodies of lung surfactant. These "disc bodies" were apparently generated by the coalescence of vesicles formed at the surface of the inner layer cells, possibly as a by-product of protein secretion by these cells.

It has been proposed that either the Igf-2 gene or the H19 gene--but not both--can be expressed from a given chromosome. Igf-2 is known to be biallelically expressed in the choroid plexus and leptomeninges, however, raising the question of whether H19 is down-regulated or absent there. We found by in situ hybridization that H19 is indeed expressed in the choroid plexus and leptomeninges of the developing mouse foetus. Comparison with the expression pattern of Igf-2 showed that the genes are coexpressed in all areas, with the exception of the choroid plexus epithelium. To evaluate whether H19 is also biallelically expressed in these tissues, we microdissected embryos from interspecific crosses and performed RNAse protection analysis on the isolated RNA. This revealed that H19 maintains its imprint in the choroid plexus/leptomeninges, being transcribed from the maternal allele at a level comparable to that in normal liver. We discuss the significance of these results for current models of Igf-2 and H19 imprinting.

We diagnosed idiopathic granulomatous angiitis of the central nervous system in a 51-year-old man by leptomeningeal and cortical biopsy. The patient's disease was prolonged, with symptoms recurring over a period of 15 years. Treatment with prednisone and cyclophosphamide produced total remission after a follow-up of 22 months. Computerized tomography was less sensitive and revealed fewer lesions than did magnetic resonance imaging. Angiography was not sensitive and leptomeningeal and cortical biopsy were essential for diagnosis in this patient.

The morphological changes in the intermediate endomeningeal layer of the goldfish brain during light and dark adaptation were studied by freeze-fracture electron microscopy. During the different stages of adaptation no significant changes were found in the density of intramembrane particles and nuclear pores in these cells. The density of plasmalemmal vesicles in the meningocyte surface increased in the groups maintained in the dark for 48 and 72 h (maximum) and then decreased in the group maintained for 96 h in the dark to a basal level. There were also morphological changes in the junctional complexes. At the upper cell membranes (in contact with the outer layer) of meningocytes in the group maintained in the dark for 48 h, we found an increase in the surface occupied by gap junctions. In addition, gap junctions were absent in the lateral membranes of meningocytes from animals maintained in the dark for 72 h. The morphology of gap junctions in the group maintained in the dark for 96 h was similar to that of the control group. These results suggest that the cells of the teleost intermediate endomeningeal layer undergo important changes in activity during adaptative experiments.

Transmission and scanning electron microscopical observations in the rat indicate a considerable capacity of the spinal meninges to reabsorb cerebrospinal fluid. The density of blood vessels and lymphatics in the duramater is extremely high, particularly in the areas of meningeal funnels and spinal nerve root sleeves. Arterioles with closely related unmyelinated nerve fibres, many fenestrated capillaries and venules predetermine these areas as sites where absorption processes could take place. At certain sites of the meningeal angle region, the arachnoid membrane, mostly multilayered, is reduced to only three or four layers. Intercellular discontinuities and cytoplasmic fenestrations occurring in the arachnoid lining cell layer result in direct communications between the subarachnoid space and cisterns of the arachnoid "reticular layer". These cisterns are partly fluid-filled, partly occupied by a net of collagen fibre bundles. Some cisterns harbour macrophages that often project filiform processes through the lining cell layer into the subarachnoid space, contacting cerebrospinal fluid. Desmosomes and gap junctions are present in all layers of the arachnoid. However, tight junctions and the continuous electrondense intercellular gap, known to occur normally within the "arachnoid barrier layer", were not seen in many sites of the meningeal angle region. Numerous arachnoid cells display a high degree of vesiculation. Cationized ferritin, introduced in vivo into the rat subarachnoid space, passes inter- and intracellularly from the cerebrospinal fluid compartment through the arachnoid membrane, reaching dural blood vessels and lymphatics. Tracer could be visualized both in the cytoplasm of the endothelium and on the luminal surface of the cells. Tracer also passed through pial cell layers into pial vessels, through leptomeningeal sheaths into vessels crossing the subarachnoid space, into the connective tissue compartment and into vessels of spinal dorsal root ganglia. In the angle region, a particularly large number of macrophages can be found on the surface of leptomeninges, within the arachnoid reticular layers, and in close relation to dural and epidural capillaries, venules and lymphatics. Their possible role in the process of cerebrospinal fluid reabsorption is discussed.

Human leptomeningeal (LM) cells were grown in serum-free medium supplemented with insulin, transferrin and bovine serum albumin. The cultured cells maintained the polygonal morphology characteristic of leptomeningeal cells. Electronmicroscopic and immunofluorescence examinations revealed interdigitation of cell membranes, invagination of cytoplasm into the nucleus, specialized intercellular junctions and the presence of cytokeratin. These characteristics are consistent with previous reports characterizing LM cells. This serum-free culture system may facilitate the in vitro study of the leptomeninges.

Ultrastructural, immunocytochemical, and immunoelectron microscopical examinations are reported that describe the morphology of putative sensory nerve endings in the dura mater encephali of the rat and the cat. Morphometrical measurements and reconstructions showed that in the cat the mean diameter of axons, the bare area of axolemma, and the content of mitochondria and vesicles are highly variable in dural nerve endings. Nerve fibers with a high volume density of mitochondria are thought to be sensory, while nerve fibers containing many small vesicles are considered autonomic. There is, however, a broad overlap of mitochondria-rich and vesicle-rich nerve fibers in the dura, so that discrimination between sensory and autonomic endings by these characteristics frequently fails. Whole-mount preparations treated cytochemically for detection of substance P- and calcitonin gene-related peptide-like immunoreactivity in the rat and the cat showed a network of immunopositive nerve fibers in the vicinity of dural blood vessels. Most of these peptidergic and probably sensory nerve fibers were found terminating in the dural connective tissue far from vessels. Calcitonin gene-related peptide-positive nerve fibers were much more abundant than substance P-positive fibers. Immunoelectron microscopic preparations revealed that calcitonin gene-related peptide- and substance P-like immunoreactivity is found in a small proportion of generally thin unmyelinated nerve fibers. These proportions were very similar in the rat and the cat. Summarizing the recent literature, the morphological characteristics of putative sensory nerve fibers in the dura mater are discussed in relation to their possible functional significance for neurogenic inflammation and nociception.

This report deals with the distribution, morphology and specific topical relationships of bone-marrow-derived cells (free cells) in the spinal meninges and dorsal root ganglia of the normal rat. The morphology of these cells has been studied by transmission and scanning electron microscopy. Cells expressing the major histocompatibility complex (MHC) class II gene product have been recognized by immunofluorescence. At the level of the transmission electron microscope, free cells are found in all layers of the meninges. Many of them display characteristic ultrastructural features of macrophages, whereas others show a highly vacuolated cytoplasm and are endowed with many processes. These elements lack a conspicuous lysosomal system and might represent dendritic cells. Scanning electron microscopy has revealed that free cells contact the cerebrospinal fluid via abundant cytoplasmic processes that cross the cell layers of the pia mater and of the arachnoid. Cells expressing the MHC class II antigen are also found in all layers of the meninges. They are particularly abundant in the layers immediately adjacent to the subarachnoid space, in the neighbourhood of dural vessels, along the spinal roots and in the dural funnels. In addition to the meninges, strong immunoreactivity for MHC class II antigen is observed in the dorsal root ganglia. The ultrastructural and immunohistochemical findings of this study suggest the existence of a well-developed system of immunological surveillance of the subarachnoid space and of the dorsal root ganglia.

We describe the morphology of the cells in the three layers of the teleost endomeninx, as viewed by freeze-fracturing. The cells of the outer endomeningeal layer are fusiform and closely packed, with interdigitations that hold the cells together, gap junctions and a few strands of particles resembling tight junctions, but no desmosomes. The intermediate layer is formed by a single layer of flattened and elongated cells with rectangular shape, and well developed junctional complexes (gap junctions, tight junctions and desmosomes). These cells also show numerous plasmalemmal vesicles (6.5 +/- 1.3/microns 2) in the upper (in contact with the outer layer) and lower (in contact with the inner layer) membranes. Cross fracture of these cells shows many membrane-bound and free vesicles. The inner layer is formed by spindle shaped cells with wide intercellular spaces filled with a granular matrix and collagen fibers. The density of intramembrane particles is higher than that in the meningocytes of the outer and intermediate layers. The morphology of the teleost endomeninx appears similar to that of the leptomeninges of birds and mammals, but the outer endomeningeal layer of teleosts (which resembles the arachnoid of elasmobranchs and amphibians) appears different from any cell layer in the meninges of amniotic vertebrates.