Electron microscopic study of the median structure of the posterior column of the spinal cord of the adult rat with a special reference to the posterior median septum.

In neuroanatomy textbooks on humans, the posterior median septum is commonly depicted along the midline of the posterior column of the spinal cord. For intramedullary spinal cord tumors, the standard surgical treatment is posterior midline myelotomy. However, its anatomical basis is still unclear. Therefore, in this study we focused on the ultrastructural characterization of the median structure of the posterior column in an adult rat. In the median part of the fasciculi gracilis, a fine lineal tissue continued from the posterior median sulcus to the 3/4th depth of the fasciculi. At higher magnification, this fine lineal tissue consisted of bundles of astrocytes, which are often disrupted and eventually disappeared. At the junction of the ventral part of the fasciculi cuneatus and the gray commissure, short lineal figures of glial tissues extended dorsally. These lineal figures of glial tissues were morphologically similar to other lineal figures of glial tissues found in the posterior column; bundles of astrocytes extending along the axons that entered the gray commissure and the perivascular lineal figures of glial tissues. In conclusion, this study revealed that the posterior median septum is composed of very fine lineal figures of glial tissues that are often disrupted and eventually disappear. We consider these basic structures to be similar in humans. Therefore, during posterior midline myelotomy, accurately separating along the posterior median septum in the posterior column is extremely difficult.

Ultrastructural and Molecular Characterization of Platelet-derived growth factor Beta-Positive Leptomeningeal Cells in the Adult Rat Brain.

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-ß)-expressing cells in the leptomeninges in the adult rat brain using light and electron microscopy. PDGFR-ß+ 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-ß+ 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-ß+ cells ensheathing the vasculature in the subarachnoid space joined with pial PDGFR-ß+ cells upon entering the cortical parenchyma, yet perivascular PDGFR-ß+ 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-ß immunoreactivity was almost exclusively localized to larger caliber vessels, and significantly decreased in capillary-like microvessels. Collectively, our data identify PDGFR-ß as a novel cellular marker for leptomeningeal fibroblasts comprising the leptomeninges and perivascular adventitial cells of the subarachnoid and penetrating large-sized cortical vasculatures.

The perivascular pathways for influx of cerebrospinal fluid are most efficient in the midbrain.

Although there are no conventional lymphatic vessels in the brain, fluid and solutes drain along basement membranes (BMs) of cerebral capillaries and arteries towards the subarachnoid space and cervical lymph nodes. Convective influx/glymphatic entry of the cerebrospinal fluid (CSF) into the brain parenchyma occurs along the pial-glial BMs of arteries. This project tested the hypotheses that pial-glial BM of arteries are thicker in the midbrain, allowing more glymphatic entry of CSF. The in vivo MRI and PET images were obtained from a 4.2-year-old dog, whereas the post-mortem electron microscopy was performed in a 12-year-old dog. We demonstrated a significant increase in the thickness of the pial-glial BM in the midbrain compared with the same BM in different regions of the brain and an increase in the convective influx of fluid from the subarachnoid space. These results are highly significant for the intrathecal drug delivery into the brain, indicating that the midbrain is better equipped for convective influx/glymphatic entry of the CSF.

The vascular stem cell niches and their significance in the brain.

AIM: This cytohistopathological study was performed for a better knowledge of phenotypes derived from pluripotent stem cells, as well as for precise location of stem cells within the vascular niche in the brain. METHODS: We used light and transmission electron microscopy to demonstrate the presence of stem cells in the vascular wall of microvessels in the cerebral and cerebellar cortex, pia mater (considered by us a cordocytic-vascular tissue), adventitia of larger cortical arteries and veins, and around vessels. We investigated multiple vascular segments and brain tissue in a variety of clinical cases, such as cerebral tumors, cerebrovascular malformations, thromboses in the carotid system, and direct laceration. RESULTS: Our morphological and ultrastructural observations pointed out many changing phenotypes, as well as cell interrelationships within the vascular niche, both for repair processes when cordocytes cooperate with mesenchymal stem cells, and pathological processes such as atherogenesis, tumorigenesis, and neurotrauma. Our results underlie the important roles of cordocytes in their interrelations with precursor/stem cells in the arterial adventitia. CONCLUSION: The cells derived from pluripotent stem cells along different lineages have had different phenotypes as they derived from hematopoietic stem cells or mesenchymal stem cells, with or without epigenetic disregulations or depending on different microenvironments. Cell interactions, phenotypes, and underlying mechanisms, as well as biological responses to different small molecules or compounds, remain to be determined by future molecular insights within the vascular niche.

Cyclical appearance of African trypanosomes in the cerebrospinal fluid: new insights in how trypanosomes enter the CNS.

It is textbook knowledge that human infective forms of Trypanosoma brucei, the causative agent of sleeping sickness, enter the brain across the blood-brain barrier after an initial phase of weeks (rhodesiense) or months (gambiense) in blood. Based on our results using an animal model, both statements seem questionable. As we and others have shown, the first infection relevant crossing of the blood brain border occurs via the choroid plexus, i.e. via the blood-CSF barrier. In addition, counting trypanosomes in blood-free CSF obtained by an atlanto-occipital access revealed a cyclical infection in CSF that was directly correlated to the trypanosome density in blood infection. We also obtained conclusive evidence of organ infiltration, since parasites were detected in tissues outside the blood vessels in heart, spleen, liver, eye, testis, epididymis, and especially between the cell layers of the pia mater including the Virchow-Robin space. Interestingly, in all organs except pia mater, heart and testis, trypanosomes showed either a more or less degraded appearance of cell integrity by loss of the surface coat (VSG), loss of the microtubular cytoskeleton and loss of the intracellular content, or where taken up by phagocytes and degraded intracellularly within lysosomes. This is also true for trypanosomes placed intrathecally into the brain parenchyma using a stereotactic device. We propose a different model of brain infection that is in accordance with our observations and with well-established facts about the development of sleeping sickness.

[The cerebral perivascular spaces: review of the literature on diffuse or focal expansion]. / Gli spazi perivascolari encefalici: revisione della letteratura sulle dilatazioni focali o diffuse.

Virchow-Robin spaces (VRS) are pial-lined, interstitial fluid-filled structures that do not directly communicate with the subarachnoid space, accompany penetrating arteries and veins and can be visualized on magnetic resonance imaging. This article reviews the imageology characteristics, the functions, the causes and the relation with neurological disorders of VRS.

From pluripotent stem cells to multifunctional cordocytic phenotypes in the human brain: an ultrastructural study.

Light microscopy and transmission electron microscopy were used to investigate surgical cases in a variety of pathological conditions (thromboses, tumors, cerebrovascular malformations, Moyamoya disease) to identify and characterize different phenotypes belonging to a new interstitial cell recently described ultrastructurally in the brain and here named "cordocyte." Also, this work is an attempt to identify and characterize precursor/stem cells for cordocytic lineage in the perivascular areas, within perivascular nerves and pia mater (now considered a cordocytic-vascular tissue). Unexpected relationships and functions emerge from observations concerning these phenotypes, almost ubiquitous, but not yet fully studied in the brain.

Breaches of the pial basement membrane are associated with defective dentate gyrus development in mouse models of congenital muscular dystrophies.

A subset of congenital muscular dystrophies (CMDs) has central nervous system manifestations. There are good mouse models for these CMDs that include POMGnT1 knockout, POMT2 knockout and Large(myd) mice with all exhibiting defects in dentate gyrus. It is not known how the abnormal dentate gyrus is formed during the development. In this study, we conducted a detailed morphological examination of the dentate gyrus in adult and newborn POMGnT1 knockout, POMT2 knockout, and Large(myd) mice by immunofluorescence staining and electron microscopic analyses. We observed that the pial basement membrane overlying the dentate gyrus was disrupted and there was ectopia of granule cell precursors through the breached pial basement membrane. Besides these, the knockout dentate gyrus exhibited reactive gliosis in these mouse models. Thus, breaches in the pial basement membrane are associated with defective dentate gyrus development in mouse models of congenital muscular dystrophies.

Three-dimensional organization of the perivascular glial limiting membrane and its relationship with the vasculature: a scanning electron microscope study.

To examine the three-dimensional structure of the perivascular glial limiting membrane (Glm) and its relationship with the vasculature in rat/mouse cerebral cortices, serial ion-etched plastic sections were observed under the scanning electron microscope and their images were reconstructed. In the case of arterioles and venules close to the pial surface, cord-like principal processes predominantly formed the endfeet; whereas in the case of capillaries and venules, sheet-like secondary processes chiefly formed Glm. Moreover, it was found that several plate-like structures protruded from the basement membrane surrounding the arterioles to penetrate into the astrocytic somata. The perivascular Glm was formed by monolayers of astrocytic processes and/or somata irrespective of the types of blood vessel. However, the thickness of the perivascular Glm, varied greatly according to the type of blood vessel. The thickness of Glm decreased in the order of arterioles, venules and capillaries. The outer surface of the perivascular Glm was extremely irregular, and sheet-like processes arising from this Glm infiltrated into the surrounding neuropil.

Photochemically initiated intracellular astrocytic calcium waves in living mice using two-photon uncaging of IP(3).

We have developed a caged IP(3) analogue for two-photon photolysis in living animals. This probe is a cell permeable version and was coloaded with a fluorescent Ca(2+) dye into astrocytes in layer 1 of the somatosensory cortex of anesthetized mice. Two-photon irradiation of single cells at 720 nm produced rapid and robust increases in intracellular Ca(2+) concentrations monitored using two-photon microscopy at 950 nm. The photoevoked intracellular Ca(2+) waves were similar in magnitude to intrinsic signals in wild type mice. These waves did not propagate to other cells beyond the targeted astrocyte. In contrast, we observed intercellular astrocytic Ca(2+) waves in two mouse models of familial Alzheimer's disease. These data suggest that Alzheimer's might perturb gliotransmission but not IP(3) signaling per se in mouse models of the disease.

Aracnoides trabecular, piamadre espinal humana y anestesia subaracnoidea / Trabecular arachnoid membrane, human spinal piamater and subarachnoid anesthesia

En los textos de anestesiología se aportan pocos detalles sobre la aracnoides trabecular y la piamadre espinal humana, a pesar de ser estructuras íntimamente relacionadas con los anestésicos locales administrados en una anestesia subaracnoidea. Complicaciones tales como el síndrome de cauda equina y el síndrome de irritación radicular transitorio posterior a la realización de bloqueos subaracnoideos, sumado a la alta permeabilidad que ha sido asociada con la piamadre, nos ha motivado a investigar sobre la ultraestructura de estas meninges. Método. Las muestras estudiadas se tomaron de cadáveres recientes y fueron examinadas por microscopía electrónica de transmisión y de barrido. Resultados. El trabeculado aracnoideo rodeaba a las raíces nerviosas, a la médula y a los vasos que se encontraban dentro del espacio subaracnoideo, limitando zonas. La piamadre estaba formada por un plano celular y por un compartimiento subpial. En el plano celular existían perforaciones naturales, especialmente en la región del cono medular y en las raíces nerviosas. Conclusiones. La inyección accidental de anestésicos locales dentro de las fundas que formaban el trabeculado aracnoideo podría justificar una dilución inadecuada de estas soluciones y el origen de síndromes neurotóxicos transitorios o permanentes. La alta permeabilidad de la piamadre podría deberse, en parte, a la existencia de perforaciones naturales, las cuales facilitarían un pasaje rápido de las sustancias introducidas en el líquido cefalorraquídeo hacia las raíces nerviosas y la médula espinal. En este caso, la membrana basal ubicada por debajo de las fibras colágenas del compartimiento subpial sería la única estructura limitante previa al tejido glio-neuronal de la médula.

Few details are to be found in anesthesiology texts concerning the trabecular arachnoid membrane and the human spinal pia mater in spite of being structures that are intimately related to local anesthetics administered in subarachnoid anesthesia. We were driven to investigate the ultrastructure of these meninges by complications such as the cauda equina syndrome and the transitory radicular irritation syndrome following subarachnoid blocks, added to the high permeability associated to the pia mater. Method. The samples analyzed were taken from recently deceased cadavers and were examined under transmission and scanning electron microscopy. Results. The arachnoid trabeculation surrounded the nerve roots, the spinal cord and the vessels within the subarachnoid space, limiting areas. The pia mater was formed by a cellular plane and by a sub-pial compartment. There were natural perforations in the cellular plane, particularly in the medullar cone region and the nerve roots. Conclusions. Accidental injection of local anesthetics into the sheaths formed by arachnoid trabeculation could be the cause of inadequate dilution of these solutions and the source of transitory or permanent neurotoxic syndromes. The high permeability of the pia mater could be partly due to the existence of natural perforations, which enable the quick passage of the substances introduced in the cerebrospinal fluid into the nerve roots and spinal cord. ln this case, the basal membrane located underneath the collagen fibers of the subpial compartment would be the only limiting structure before the glioneural tissue of the spinal cord.

Os textos de anestesiologia fornecem poucos detalhes sobre a aracnóide trabecular e a pia-máter espinhal humana, apesar delas serem estruturas intimamente relacionadas com os anestésicos locais administrados em uma anestesia subaracnóidea. Complicações tais como a síndrome de cauda eqüina e a síndrome de irritação radicular transitória posterior a bloqueios subaracnóideosas quais se soma a alta permeabilidade, que tem sido associada à pia-máter -levou-nos a pesquisar a ultraestrutura dessas meninges. Método. As amostras estudadas foram coletadas de cadáveres recentes e examinadas por microscopia eletrónica de transmissão e de varredura. Resultados. A trabeculação aracnóidea rodea va as raízes nervosas, a medula e os vasos no interior do espaço subaracnóide, limitando zonas. A pia-máter estava formada por um plano celular e um espaço subpial. No plano celular existiam perfurações naturais, especialmente na regiáo do cone medular e nas raízes nervosas. Conclusóes. A injeção acidental de anestésicos locais no interior das coberturas que formavam a trabeculação aracnoidea poderia justificar uma diluição inadequada das soluções e a origem de síndromes neurotóxicas transitórias ou permanentes. A causa da alta permeabilidade da pia-máter seria, em parte, a existencia de perfurações naturais que facilitariam a rápida passagem das substancias introduzidas no líquido cefalorraquiano para as raizes nervosas e a medula espinhal. Neste caso, a membrana basal localizada abaixo das fibras colágenas do espaço subpial seria a única estrutura limitante anterior ao tecido glio-neuronal da medula.

Comparison of metastatic brain tumour models using three different methods: the morphological role of the pia mater.

As methods of cancer diagnosis and treatment progress, interest in metastatic brain tumours continues to increase. There are many studies using various methods of animal model and we considered that each model reflects different pathological processes because of the unique composition of the brain. We prepared metastatic brain tumour models using three different methods. In this study, we attempted to elucidate the roles of the pia mater in brain metastasis. The metastatic foci showed an angiocentric pattern, forming collars of neoplastic cells, and were designated 'perivascular proliferations'. Furthermore, we observed neoplastic cells that infiltrated the brain parenchyma, the border of which had become indistinct. These were labelled 'invasive proliferations'. The internal carotid artery injection model reflects haematogenous metastasis. In this model, both perivascular and invasive proliferations were observed. The intrathecal injection model reflects leptomeningeal carcinomatosis. In this model, metastasis to the meninges was observed. In the stereotactic injection model, the tumour proliferation at the injection site and the infiltration into the brain parenchyma were observed. The pia-glial membrane serves as a scaffold when neoplastic cells spread to the perivascular space forming angiocentric pattern. The pia-glial membrane is found between the brain parenchyma and blood vessels. Blood vessels penetrate the brain through tunnels known as perivascular spaces that are covered by pia mater. Three different methods which we prepared reflect three different pathological processes. Our findings suggest that the pia mater is a critical factor in brain metastasis.

An experimental Staphylococcus aureus meningitis model for investigating induced leptomeningeal and subpial inflammation in rats: a transmission electron microscopy study.

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.

A double three-step theory of brain metastasis in mice: the role of the pia mater and matrix metalloproteinases.

The brain is frequently affected by the spread of lung cancer, and haematogenous metastasis is a common route to brain metastasis. We therefore developed an isogenic brain metastasis model of lung cancer to use the Lewis lung carcinoma cell line and analysed dynamics of neoplastic cells after extravasation. Histological analysis revealed two characteristic patterns: metastatic foci exhibiting an angiocentric pattern were designated 'perivascular proliferations'; neoplastic cells infiltrating the brain parenchyma were designated 'invasive proliferations'. Electron microscopic observation of perivascular proliferations showed that neoplastic cells were confined to the perivascular space. In invasive proliferations, however, fragments of collagen fibre were observed in the gaps between neoplastic cells, indicating that the neoplastic cells had disintegrated the pia-glial membrane. We analysed the expressions of matrix metalloproteinase-2 (MMP-2) and MMP-9 by using both immunohistochemical analysis and real-time polymerase chain reaction analysis. MMP-2 expression was significantly higher in invasive proliferations. MMP-9 expression was significantly higher in day 7, but there was no significant difference in day 11. The pia-glial membrane and perivascular space are the barriers that neoplastic cells must overcome to infiltrate the brain. In conclusion, our findings suggest that brain metastasis requires two distinct processes.

Breaches of the pial basement membrane and disappearance of the glia limitans during development underlie the cortical lamination defect in the mouse model of muscle-eye-brain disease.

Neuronal overmigration is the underlying cellular mechanism of cerebral cortical malformations in syndromes of congenital muscular dystrophies caused by defects in O-mannosyl glycosylation. Overmigration involves multiple developmental abnormalities in the brain surface basement membrane, Cajal-Retzius cells, and radial glia. We tested the hypothesis that breaches in basement membrane and the underlying glia limitans are the key initial events of the cellular pathomechanisms by carrying out a detailed developmental study with a mouse model of muscle-eye-brain disease, mice deficient in O-mannose beta1,2-N-acetylglucosaminyltransferase 1 (POMGnT1). The pial basement membrane was normal in the knockout mouse at E11.5. It was breached during rapid cerebral cortical expansion at E13.5. Radial glial endfeet, which comprise glia limitans, grew out of the neural boundary. Neurons moved out of the neural boundary through these breaches. The overgrown radial glia and emigrated neurons disrupted the overlying pia mater. The overmigrated neurons did not participate in cortical plate (CP) development; rather they formed a diffuse cell zone (DCZ) outside the original cortical boundary. Together, the DCZ and the CP formed the knockout cerebral cortex, with disappearance of the basement membrane and the glia limitans. These results suggest that disappearance of the basement membrane and the glia limitans at the cerebral cortical surface during development underlies cortical lamination defects in congenital muscular dystrophies and a cellular mechanism of cortical malformation distinct from that of the reeler mouse, double cortex syndrome, and periventricular heterotopia.

Adult human subventricular, subgranular, and subpial zones contain astrocytes with a specialized intermediate filament cytoskeleton.

Human glial fibrillary acidic protein-delta (GFAP-delta) is a GFAP protein isoform that is encoded by an alternative splice variant of the GFAP-gene. As a result, GFAP-delta protein differs from the predominant splice form, GFAP-alpha, by its C-terminal protein sequence. In this study, we show that GFAP-delta protein is not expressed by all GFAP-expressing astrocytes but specifically by a subpopulation located in the subpial zone of the cerebral cortex, the subgranular zone of the hippocampus, and, most intensely, by a ribbon of astrocytes following the ependymal layer of the cerebral ventricles. Therefore, at least in the sub ventricular zone (SVZ), GFAP-delta specifically marks the population of astrocytes that contain the neural stem cells in the adult human brain. Interestingly, the SVZ astrocytes actively splice GFAP-delta transcripts, in contrast to astrocytes adjacent to this layer. Furthermore, we show that GFAP-delta protein, unlike GFAP-alpha, is not upregulated in astrogliosis. Our data therefore indicate a different functional role for GFAP-delta in astrocyte physiology. Finally, transfection studies showed that GFAP-delta protein expression has a negative effect on GFAP filament formation, and therefore could be important for modulating intermediate filament cytoskeletal properties, possibly facilitating astrocyte motility. Further studies on GFAP-delta and the cells that express it are important for gaining insights into its function during differentiation, migration and during health and disease.

Abeta42 is essential for parenchymal and vascular amyloid deposition in mice.

Considerable circumstantial evidence suggests that Abeta42 is the initiating molecule in Alzheimer's disease (AD) pathogenesis. However, the absolute requirement for Abeta42 for amyloid deposition has never been demonstrated in vivo. We have addressed this by developing transgenic models that express Abeta1-40 or Abeta1-42 in the absence of human amyloid beta protein precursor (APP) overexpression. Mice expressing high levels of Abeta1-40 do not develop overt amyloid pathology. In contrast, mice expressing lower levels of Abeta1-42 accumulate insoluble Abeta1-42 and develop compact amyloid plaques, congophilic amyloid angiopathy (CAA), and diffuse Abeta deposits. When mice expressing Abeta1-42 are crossed with mutant APP (Tg2576) mice, there is also a massive increase in amyloid deposition. These data establish that Abeta1-42 is essential for amyloid deposition in the parenchyma and also in vessels.

Localization and functional analyses of the MLC1 protein involved in megalencephalic leukoencephalopathy with subcortical cysts.

Mutations in the MLC1 gene are responsible for one form of the neurological disorder megalencephalic leukoencephalopathy with subcortical cysts (MLC). The disease is a type of vacuolating myelinopathy. The biochemical properties and the function of the MLC1 protein are unknown. To characterize MLC1, we generated polyclonal antibodies. The MLC1 protein was detected in the brain, assembled into higher molecular complexes, as assessed by assembly-dependent trafficking assays. In situ hybridization and immunohistochemistry were used to determine MLC1 localization within the adult mouse brain. MLC1 was expressed in neurons, detected preferentially in particular axonal tracts. This expression pattern correlates with the major phenotype observed in the disease. In addition, it was expressed in some astrocytes, concentrating in Bergmann glia, the astrocyte end-feet membranes adjacent to blood vessels and in astrocyte-astrocyte membrane contact regions. Other neuronal barriers, such as the ependyma and the pia mater, were also positive for MLC1 expression. MLC1 was detected in vivo and in heterologous systems at the plasma membrane. MLC mutations impaired folding, and the defect was corrected in vitro by addition of curcumin, a Ca(2+)-ATPase inhibitor. In summary, this study provides an explanation as to why mutations in MLC1 provoke the disease and points to a possible therapy for some patients.

Ultrastructural findings in human spinal pia mater in relation to subarachnoid anesthesia.

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.

Macrophages and dendritic cells in the rat meninges and choroid plexus: three-dimensional localisation by environmental scanning electron microscopy and confocal microscopy.

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.