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Traditionally, the primary role of the meninges is thought to be structural, i.e., to act as a surrounding membrane that contains and cushions the brain with cerebrospinal fluid. During development, the meninges is formed by both mesenchymal and neural crest cells. There is now emerging evidence that subsets of undifferentiated stem cells might persist in the adult meninges. In this mini-review, we survey representative studies of brain-meningeal interactions and discuss the hypothesis that the meninges are not just protective membranes, but instead contain multiplex stem cell subsets that may contribute to central nervous system (CNS) homeostasis. Further investigations into meningeal multipotent cells may reveal a "hidden" target for promoting neurovascular remodeling and repair after CNS injury and disease., (© 2021. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Certain driver mutations and pathological diagnoses are associated with the anatomical site of meningioma, based on which the meninges have different embryological origins. We hypothesized that mutations and pathological diagnoses of meningiomas are associated with different embryological origins. We comprehensively evaluated associations among tumor location, pathological diagnosis (histological type), and genetic alterations including AKT1, KLF4, SMO, POLR2A, and NF2 mutations and 22q deletion in 269 meningioma cases. Based on the embryological origin of meninges, the tumor locations were as follows: neural crest, paraxial mesodermal, and dorsal mesodermal origins. Tumors originating from the dura of certain embryologic origin displayed a significantly different pathological diagnoses and genetic abnormality ratio. For instance, driver genetic mutations with AKT1, KLF4, SMO, and POLR2A, were significantly associated with the paraxial mesodermal origin (p = 1.7 × 10 -10 ). However, meningiomas with NF2-associated mutations were significantly associated with neural crest origin (p = 3.9 × 10 -12 ). On analysis of recurrence, no difference was observed in embryological origin. However, POLR2A mutation was a risk factor for the tumor recurrence (p = 1.7 × 10 -2 , Hazard Ratio 4.08, 95% Confidence Interval 1.28-13.0). Assessment of the embryological origin of the meninges may provide novel insights into the pathomechanism of meningiomas.

Meningiomas are tumors arising from the meninges and represent the most frequent central nervous system tumors in adults. Recent large-scale genetic studies and preclinical meningioma mouse modelling led to a better comprehension of meningioma development and suggested evidences of close relationships between meningeal embryology and tumorigenesis. In this non-systematic review, we summarize the current knowledge on meningeal embryology and developmental biology, and illustrate how meningioma tumorigenesis is deeply related to meningeal embryology, concerning the potential cell of origin, the role of reactivation of embryonic stem cells, the influence of the embryonic tissue of origin, and the parallelism between topography-dependant molecular pathways involved in normal meninges and in meningioma development. Our study emphasizes why future studies on meningeal embryology are mandatory to affine our comprehension of mechanisms underlying meningioma initiation and development.

The meninges are a multilayered structure composed of fibroblasts, blood and lymphatic vessels, and immune cells. Meningeal fibroblasts secrete a variety of factors that control CNS development, yet strikingly little is known about their heterogeneity or development. Using single-cell sequencing, we report distinct transcriptional signatures for fibroblasts in the embryonic dura, arachnoid, and pia. We define new markers for meningeal layers and show conservation in human meninges. We find that embryonic meningeal fibroblasts are transcriptionally distinct between brain regions and identify a regionally localized pial subpopulation marked by the expression of μ-crystallin. Developmental analysis reveals a progressive, ventral-to-dorsal maturation of telencephalic meninges. Our studies have generated an unparalleled view of meningeal fibroblasts, providing molecular profiles of embryonic meningeal fibroblasts by layer and yielding insights into the mechanisms of meninges development and function., Competing Interests: Declaration of Interests The authors declare no competing interests., (Copyright © 2020 Elsevier Inc. All rights reserved.)

The meninges are membranous layers surrounding the central nervous system. In the head, the meninges lie between the brain and the skull, and interact closely with both during development. The cranial meninges originate from a mesenchymal sheath on the surface of the developing brain, called primary meninx, and undergo differentiation into three layers with distinct histological characteristics: the dura mater, the arachnoid mater, and the pia mater. While genetic regulation of meningeal development is still poorly understood, mouse mutants and other models with meningeal defects have demonstrated the importance of the meninges to normal development of the calvaria and the brain. For the calvaria, the interactions with the meninges are necessary for the progression of calvarial osteogenesis during early development. In later stages, the meninges control the patterning of the skull and the fate of the sutures. For the brain, the meninges regulate diverse processes including cell survival, cell migration, generation of neurons from progenitors, and vascularization. Also, the meninges serve as a stem cell niche for the brain in the postnatal life. Given these important roles of the meninges, further investigation into the molecular mechanisms underlying meningeal development can provide novel insights into the coordinated development of the head., (© 2019 Wiley Periodicals, Inc.)

Cranial neural crest cells (CNCCs) give rise to cranial mesenchyme (CM) that differentiates into the forebrain meningeal progenitors in the basolateral and apical regions of the head. This occurs in close proximity to the other CNCC-CM-derivatives, such as calvarial bone and dermal progenitors. We found active Wnt signaling transduction in the forebrain meningeal progenitors in basolateral and apical populations and in the non-meningeal CM preceding meningeal differentiation. Here, we dissect the source of Wnt ligand secretion and requirement of Wnt/β-catenin signaling for the lineage selection and early differentiation of the forebrain meninges. We find persistent canonical Wnt/β-catenin signal transduction in the meningeal progenitors in the absence of Wnt ligand secretion in the CM or surface ectoderm, suggesting additional sources of Wnts. Conditional mutants for Wntless and β-catenin in the CM showed that Wnt ligand secretion and Wnt/β-catenin signaling were dispensable for specification and proliferation of early meningeal progenitors. In the absence of β-catenin in the CM, we found diminished laminin matrix and meningeal hypoplasia, indicating a structural and trophic role of mesenchymal β-catenin signaling. This study shows that β-catenin signaling is required in the CM for maintenance and organization of the differentiated meningeal layers in the basolateral and apical populations of embryonic meninges., (© 2019 Wiley Periodicals, Inc.)

Nervous system development is a precisely orchestrated series of events requiring a multitude of intrinsic and extrinsic cues. Sortilin and SorCS2 are members of the Vps10p receptor family with complementary influence on some of these cues including the neurotrophins (NTs). However, the developmental time points where sortilin and SorCS2 exert their activities in conjunction or independently still remain unclear. In this study we present the characterization of the spatiotemporal expression pattern of sortilin and SorCS2 in the developing murine nervous system. Sortilin is highly expressed in the fetal nervous system with expression localized to distinct cell populations. Expression was high in neurons of the cortical plate and developing allocortex, as well as subpallial structures. Furthermore, the neuroepithelium lining the ventricles and the choroid plexus showed high expression of sortilin, together with the developing retina, spinal ganglia, and sympathetic ganglia. In contrast, SorCS2 was confined in a marked degree to the thalamus and, at E13.5, the floor plate from midbrain rostrally to spinal cord caudally. SorCS2 was also found in the ventricular zones of the ventral hippocampus and nucleus accumbens areas, in the meninges and in Schwann cells. Hence, sortilin and SorCS2 are extensively present in several distinct anatomical areas in the developing nervous system and are rarely co-expressed. Possible functions of sortilin and SorCS2 pertain to NT signaling, axon guidance and beyond. The present data will form the basis for hypotheses and study designs for unravelling the functions of sortilin and SorCS2 during the establishment of neuronal structures and connections., (© 2018 Wiley Periodicals, Inc.)

Background: Traditionally, the central nervous system (CNS) has been viewed as an immune-privileged environment with no lymphatic vessels. This view was partially overturned by the discovery of lymphatic vessels in the dural membrane that surrounds the brain, in contact with the interior surface of the skull. We here examine the distribution and developmental timing of these lymphatic vessels., Results: Using the Prox1-GFP BAC transgenic reporter and immunostaining with antibodies to lymphatic markers LYVE-1, Prox1, and Podoplanin, we have carried out whole-mount imaging of dural lymphatic vasculature at postnatal stages. We have found that between birth and postnatal day (P) 13, lymphatic vessels extend alongside dural blood vessels from the side of the skull toward the midline. Between P13 and P20, lymphatic vessels along the transverse sinuses reach the superior sagittal sinus (SSS) and extend along the SSS toward the olfactory bulb., Conclusions: Compared with the embryonic developmental timing of lymphatic vessels in other tissues, e.g. skin, dural lymphatic vessel development is dramatically delayed. This study provides useful anatomical data for continuing investigations of the fundamental mechanisms that underlie dural lymphatic vessel development. Developmental Dynamics 247:741-753, 2018. © 2018 Wiley Periodicals, Inc., (© 2018 Wiley Periodicals, Inc.)

Sturge-Weber syndrome is a sporadic congenital neurocutaneous disorder caused by a somatic activating mutation in GNAQ; it affects 1 in every 20,000 to 50,000 newborns. It is characterized by a facial Port-wine stain, leptomeningeal angiomatosis, and glaucoma. Seizures are the most common neurological manifestation and typically present in the first months of life. Glaucoma may be present at birth or develop later. Neuroimaging studies show leptomeningeal angiomatosis, supporting diagnosis. Standard treatment for Sturge-Weber syndrome includes laser treatment for the Port-wine stain, anticonvulsants, and medical or surgical treatment for the glaucoma. Prognosis depends on the extent of leptomeningeal involvement and the severity of the glaucoma., (Copyright © 2016 AEDV. Publicado por Elsevier España, S.L.U. All rights reserved.)

During embryonic development oligodendrocyte precursor cells (OPCs) are generated first in the ventral forebrain and migrate dorsally to occupy the cortex. The molecular cues that guide this migratory route are currently completely unknown. Here, we show that bone morphogenetic protein-4 (Bmp4), Bmp7, and Tgfβ1 produced by the meninges and pericytes repelled ventral OPCs into the cortex at mouse embryonic stages. Ectopic activation of Bmp or Tgfβ1 signaling before the entrance of OPCs into the cortex hindered OPC migration into the cortical areas. OPCs without Smad4 signaling molecules also failed to migrate into the cortex efficiently and formed heterotopia in ventral areas. OPC migration into the cortex was also dramatically reduced by conditional inhibition of Tgfβ1 or Bmp expression from mesenchymal cells. The data suggest that mesenchymal Tgfβ family proteins promote migration of ventral OPCs into the cortex during corticogenesis., (Copyright © 2014 the authors 0270-6474/14/3414973-11$15.00/0.)

Background: Nestin is a type VI intermediate filament protein known as a marker for progenitor cells that can be mostly found in tissues during the embryonic and fetal periods. In our study, we aimed to determine the expression of nestin in meninges covering the brain tissue at different developmental stages and in the new born., Methods: In this study 10 human fetuses in different development stages between developmental weeks 9-34 and a newborn brain tissue were used. Fetuses in paraffin section were stained with H+E and nestin immunohistochemical staining protocol was performed., Results: In this study, in the human meninges intense nestin expression was detected as early as in the 9th week of development. Intensity of this expression gradually decreased in later stages of development and nestin expression still persisted in a small population of newborn meningeal cells., Conclusion: In the present study, nestin positive cells gradually diminished in the developing and maturing meninges during the fetal period. This probably depends on initiation of a decrease in nestin expression and replacement with other tissue-specific intermediate filaments while the differentiation process continues. These differences can make significant contributions to the investigation and diagnosis of various pathological disorders (Tab. 1, Fig. 3, Ref. 36).

We examined the role of the orphan nuclear hormone receptor CoupTFI in mediating cortical development downstream of meningeal retinoic acid signaling. CoupTFI is a regulator of cortical development known to collaborate with retinoic acid (RA) signaling in other systems. To examine the interaction of CoupTFI and cortical RA signaling we utilized Foxc1-mutant mice in which defects in meningeal development lead to alterations in cortical development due to a reduction of RA signaling. By analyzing CoupTFI(-/-);Foxc1(H/L) double mutant mice we provide evidence that CoupTFI is required for RA rescue of the ventricular zone and the neurogenic phenotypes in Foxc1-mutants. We also found that overexpression of CoupTFI in Foxc1-mutants is sufficient to rescue the Foxc1-mutant cortical phenotype in part. These results suggest that CoupTFI collaborates with RA signaling to regulate both cortical ventricular zone progenitor cell behavior and cortical neurogenesis.

Embryonic development is controlled by a small set of signal transduction pathways, with vastly different phenotypic outcomes depending on the time and place of their recruitment. How the same molecular machinery can elicit such specific and distinct responses, remains one of the outstanding questions in developmental biology. Part of the answer may lie in the high inherent genetic complexity of these signaling cascades, as observed for the Wnt-pathway. The mammalian genome encodes multiple Wnt proteins and receptors, each of which show dynamic and tightly controlled expression patterns in the embryo. Yet how these components interact in the context of the whole organism remains unknown. Here we report the generation of a novel, inducible transgenic mouse model that allows spatiotemporal control over the expression of Wnt5a, a protein implicated in many developmental processes and multiple Wnt-signaling responses. We show that ectopic Wnt5a expression from E10.5 onwards results in a variety of developmental defects, including loss of hair follicles and reduced bone formation in the skull. Moreover, we find that Wnt5a can have dual signaling activities during mouse embryonic development. Specifically, Wnt5a is capable of both inducing and repressing β-catenin/TCF signaling in vivo, depending on the time and site of expression and the receptors expressed by receiving cells. These experiments show for the first time that a single mammalian Wnt protein can have multiple signaling activities in vivo, thereby furthering our understanding of how signaling specificity is achieved in a complex developmental context., (Copyright © 2012 Elsevier Inc. All rights reserved.)

The meninges have traditionally been viewed as specialized membranes surrounding and protecting the adult brain from injury. However, there is increasing evidence that the fetal meninges play important roles during brain development. Through the release of diffusible factors, the meninges influence the proliferative and migratory behaviors of neural progenitors and neurons in the forebrain and hindbrain. Meningeal cells also secrete and organize the pial basement membrane (BM), a critical anchor point for the radially oriented fibers of neuroepithelial stem cells. With its emerging role in brain development, the potential that defects in meningeal development may underlie certain congenital brain abnormalities in humans should be considered. In this review, we will discuss what is known about assembly of the fetal meninges and review the role of meningeal-derived proteins in mouse and human brain development., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

Development of the meninges in and around the plica ventralis encephali has not been well documented. A distinct mesenchymal structure, the so-called plica ventralis encephali, is sandwiched by the fetal mesencephalic flexure. We histologically examined paraffin-embedded sections from 18 human embryos and fetuses at 6-12 weeks of gestation. In the loose tissues of the plica, the first meninx appeared as a narrow membrane along the oculomotor nerve at 7-8 weeks. Subsequently, the plica ventralis evolved into 3 parts: bilateral lateral mesenchymal condensations and a primitive membranous meninx extending between. Notably, the topographical anatomy of the oculomotor, trochlear and trigeminal nerves did not change: the oculomotor nerve ran along the rostral aspect of the membranous meninx, the trigeminal nerve ran along the caudal side of the lateral mesenchymal condensation, and the trochlear nerve remained embedded in the lateral condensation. Up to 9-10 weeks, the lateral mesenchymal condensations became tongue-like folds; i.e., the primitive form of the tentorium cerebelli, while the membranous meninx became the diaphragma sellae. The falx cerebri seemed to develop from the tongue-like folds. Overall, the final tentorium cerebelli corresponded to the regressed plica ventralis, while the parasellar area originated from the base of the plica and other tissues along the ventral aspects of the basisphenoid and basioccipital., (Copyright © 2010 Elsevier GmbH. All rights reserved.)

The development of the epidural space was studied on 51 series of sections from embryos and early fetuses aged from four to 13 weeks using histological and embryological methods for plastic and graphic reconstruction. We found that three stages can be discriminated in the development of the epidural space: I) the primary epidural space (embryos of 16-31 mm crown-rump length (CRL); II) reduction of the primary epidural space (embryos of 35-55 mm CRL); and III) the secondary epidural space (embryos of 60-70 mm CRL and fetuses of 80-90 mm CRL). The morphogenesis of the primary epidural space is determined by the formative influence of the spinal cord and its dura mater, while that of the secondary epidural space is determined by the walls of the vertebral canal. In the spinal cord-dura mater of the spinal cord-vertebral canal correlation system, the latter two components, subjected to the inducing influence of the first, determine the morphogenesis of the epidural space via a system of morphogenetic correlations. The correlational relationships are apparent as time-linked connections between the rudiment of the dura mater of the spinal cord and the vertebral canal, resulting in the stage-by-stage formation of the epidural space in the ventrodorsal and craniocaudal directions. These same morphogenetic correlations also determine the staging of the development of the epidural space.

The meninges are an important connective tissue envelope investing the brain. Their function is to provide a protective coating to the brain and also participate in the formation of blood-brain barrier. Understanding their anatomy is fundamental to understanding the location and spread of pathologies in relation to the layers. It also provides an insight into the characteristics of such pathologies when imaging them. This review aims to describe the anatomy of the meninges, and to demonstrate the imaging findings of specific features.

The dura is traditionally viewed as a supportive fibrous covering of the brain containing the dural venous sinuses but otherwise devoid of vessels and lacking any specific function. However, review of the embryology and anatomy reveals the dura to be a complex, vascularized and innervated structure, not a simple fibrous covering. The dura contains an inner vascular plexus that is larger in the infant than in the adult, and this plexus likely plays a role in CSF absorption. This role could be particularly important in the infant whose arachnoid granulations are not completely developed. Although subdural hemorrhage is frequently traumatic, there are nontraumatic conditions associated with subdural hemorrhage, and the inner dural plexus is a likely source of bleeding in these nontraumatic circumstances. This review outlines the development and age-specific vascularity of the dura and offers an alternative perspective on the role of the dura in homeostasis of the central nervous system.

Loss of function of the mouse forkhead/winged helix transcription factor Foxc1 induces congenital hydrocephalus and impaired skull bone development due to failure of apical expansion of the bone. In this study we investigated meningeal development in the congenital hydrocephalus (ch) mouse with spontaneous loss of function mutant of Foxc1, around the period of initiation of skull bone apical expansion. In situ hybridization of Runx2 revealed active apical expansion of the frontal bone begins between embryonic day 13.5 and embryonic day 14.5 in the wild type, whereas expansion was inhibited in the mutant. Ultrastructural analysis revealed that three layers of the meninges begin to develop at E13.5 in the basolateral site of the head and subsequently progress to the apex in wild type. In ch homozygotes, although three layers were recognized at first at the basolateral site, cell morphology and structure of the layers became abnormal except for the pia mater, and arachnoidal and dural cells never differentiated in the apex. We identified meningeal markers for each layer and found that their expression was down-regulated in the mutant arachnoid and dura maters. These results suggest that there is a close association between meningeal development and the apical growth of the skull bones.

Object: This study was undertaken to test a hypothesis that meningiomas of the midline skull base and spine are predominantly of the meningothelial histological subtype., Methods: The cases of 794 consecutive patients who underwent resection for meningioma at the Cleveland Clinic between January 1991 and March 2004 were reviewed retrospectively. The authors analyzed the relationship between the tumors' histological subtypes and sites of origin in the 731 patients from this group who harbored tumors that were determined to be benign histologically (World Health Organization Grade I). Meningothelial meningiomas (MMs) accounted for 63.5% (464/731) of the Grade I tumors. The incidence of MM according to the site of origin was as follows: 84.9% (186/219) in the midline skull base, 58.3% (35/60) in the lateral skull base, 48.5% (183/377) in a non-skull base location, and 80% (60/75) in spinal locations. The incidence of MM in the midline skull base and spinal locations were significantly higher than in non-skull base or lateral skull base locations., Conclusions: Meningiomas of the midline neuraxis are predominantly meningotheliomas. Analysis of the increasingly available data on genetic and topographic characteristics of MMs suggests that they may represent a unique entity, contrary to the prevailing belief that all benign meningiomas are identical tumors.

The aim of this study was to describe the embryologic and foetal development of the anterior paraclinoid region and more precisely the relationship of the internal carotid artery to the dura mater. This has been done by examining a collection of histological sections, representing a continuous series of 48 embryologic and foetal specimens, covering the period of the first 6 months of intra-uterine life. Neurological and vascular elements develop during the embryologic period; the internal carotid artery is recognizable in the various sections of its course and acquires a histological adult parietal constitution. The foetal period corresponds to the development of the meningeal structures. The superior, medial and lateral walls appear on the fifteenth week of amenorrhoea and do not change after that. The internal carotid artery enters subarachnoid space accompanied by a sleeve of mesenchymatous cells, which fixes it to the anterior clinoid process. The constitution of this sleeve, arising from the superior wall of the lateral sellar compartment, remained independent of the principle vascular part, which allows the formation of a plan of cleavage. The foetal relations of the dura mater and the internal carotid artery were seen to be different from those of adult subjects described in the literature, suggesting an existence of period of maturation postnatally.

The avian lumbosacral vertebral column and spinal cord show a number of specializations which have recently been interpreted as a sense organ of equilibrium. This sense organ is thought to support balanced walking on the ground. Although most of the peculiar structures have been described previously, there was a need to reevaluate the specializations with regard to the possible function as a sense organ. Specializations were studied in detail in the adult pigeon. The development of the system was studied both in the pigeon (semiprecocial at hatching) and in the chicken (precocial). Specializations in the vertebral canal consist of a considerable enlargement, which is not due to an increase in the size of the spinal nervous tissue, but to a large glycogen body embedded in a dorsal rhomboid sinus. The dorsal wall of the vertebral canal shows segmented bilateral dorsal grooves, which are covered by the meninges towards the lumen of the vertebral canal leaving openings in the midline and laterally. This results in a system of lumbosacral canals which look and may function similar to the semicircular canals in the inner ear. Laterally these canals open above ventrolateral protrusions or accessory lobes of the spinal cord which contain neurons. There are large subarachnoidal cerebrospinal fluid spaces, lateral and ventral to the accessory lobes. Movement of this fluid is thought to stimulate the lobes mechanically. As to the development of avian lumbosacral specializations, main attention was given to the organization of the lobes and the adjacent fluid spaces including the dorsal canals. In the pigeon the system is far from being adult-like at hatching but maturates rapidly after hatching. In the chicken the system looks already adult-like at hatching. The implications derived from the structural findings are discussed with regard to a possible function of the lumbosacral specializations as a sense organ of equilibrium. The adult-like organization in the newly hatched chickens, which walk around immediately after hatching, supports the assumed function as a sense organ involved in the control of locomotion on the ground.

The meninges correspond to an anatomical concept. For the morphologist, the microscopic organization, the hypothetical presence of a subdural space, the nature of the interface between the deep meningeal layer and the nervous parenchyma in the perivascular spaces are the central issues. For the clinician, dynamic aspects of cerebrospinal fluid flow, secretion, and resorption are essential factors with practical consequences in terms of disease and patient management. Comparative anatomy, embryology, and organogenesis provide an interesting perspective for the descriptive and functional anatomy of the meninges. Usually considered as protective membranes, the meninges play a prominent role in the development and maintenance of the central nervous system. The meninges are in constant evolution, from their formation to senescence. The meninges present three layers in children and adults: the dura mater, the arachnoid and the pia mater. The cerebrospinal fluid is secreted by the choroid plexuses, flows through the ventricles and the subarachnoid space, and is absorbed by arachnoid granulations. Other sites of secretion and resorption are suggested by comparative anatomy and human embryology and organogenesis.

The formation of endothelial tight junctions (TJs) is crucial in blood-brain barrier (BBB) differentiation, and the expression and targeting of TJ-associated proteins mark the beginning of BBB functions. Using confocal microscopy, this study analyzed endothelial TJs in adult human cerebral cortex and the fetal telencephalon and leptomeninges in order to compare the localization of two TJ-associated transmembrane proteins, occludin and claudin-5. In the arterioles and microvessels of adult brain, occludin and claudin-5 form continuous bands of endothelial immunoreactivity. During fetal development, occludin and claudin-5 immunoreactivity is first detected as a diffuse labeling of endothelial cytoplasm. Later, at 14 weeks, the immunosignal for both proteins shifts from the cytoplasm to the interface of adjacent endothelial cells, forming a linear, widely discontinuous pattern of immunoreactivity that achieves an adult-like appearance within a few weeks. These results demonstrate that occludin and claudin-5 expression is an early event in human brain development, followed shortly by assembly of both proteins at the junctional areas. This incremental process suggests more rapid establishment of the human BBB, consistent with its specific function of creating a suitable environment for neuron differentiation and neurite outgrowth during neocortical histogenesis.

A variety of connexins are expressed in the diverse cell types of the central nervous system and are thought to regulate some of the functional properties exhibited by immature and mature cells. A proper understanding of the role of specific connexins in these processes requires an unambiguous characterization of their spatial and temporal pattern of expression. In order to define the cellular distribution of connexin 26 (Cx26) in the mouse we have generated a reporter allele (Cx26lacZ) by genetically manipulating the locus so that the beta-galactosidase (lacZ) gene is expressed from the endogenous Cx26 promoter. This modification decreased expression from the allele and resulted in embryonic lethality for the Cx26lacZ/lacZ genotype in accordance with previous studies on Cx26 knock-out animals indicating that Cx26-containing gap junctions are necessary for embryonic development. Despite the lower than expected transcript levels, the amount of lacZ protein produced in heterozygous mice was sufficient to label tissues known to contain Cx26, such as liver, kidney, skin, cochlea, small intestine, placenta and thyroid gland. In the embryonic and mature central nervous system, however, lacZ was restricted to meningeal cells and could not be detected in either neurons or glia. The absence of Cx26 mRNA in these cells could also be confirmed by reverse transcription-polymerase chain reaction and in situ hybridization. Our experiments indicate that the Cx26lacZ mouse line can be used as a reporter of Cx26 gene expression and suggest that Cx26, contrary to previous reports, is restricted to the meninges in both embryonic and adult brain.

The neural scar that forms after injury to the mammalian central nervous system is a barrier to sprouting and regenerating axons. In addition to reactive astrocytes that are present throughout the lesion site, leptomeningeal fibroblasts invade the lesion core. When isolated in vitro, these cells form a very poor substrate for growing neurites, even more so than reactive astrocytes. Nevertheless the molecular mechanisms involved in this growth inhibition are not well understood. Semaphorins have been reported to be upregulated in meningeal cells (MCs) on mechanical injury to the brain and spinal cord. In the present study, we show that Sema3A mRNA and active protein are produced by cultured meningeal cells. A protein extract from these cells induces the collapse of embryonic dorsal root ganglion (DRG) growth cones. This collapsing activity is partially blocked by neuropilin-1 antibodies and is absent in meningeal cells derived from Sema3A-knockout mice. In addition to growth cone collapse, recombinant Sema3A but not Sema3C inhibits neurite outgrowth of embryonic DRGs. Consistent with this result we find that the inhibitory effect of meningeal cells on neurite outgrowth is partially overcome on Sema3A-deficient MCs. Furthermore we show that the inhibitory effect of MC-derived Sema3A on neurite outgrowth is modulated by nerve growth factor. Our results show that Sema3A, a chemorepellent during nervous system development, is a major neurite growth-inhibitory molecule in meningeal fibroblasts and is therefore likely to contribute to the inhibitory properties of the neural scar.

One general function for retinoic acid (RA) is pattern organization in the CNS. This regulatory factor has an essential role in spinal cord motor neuron and early posterior hindbrain development. In the anterior CNS, however, there is only a limited number of foci of RA synthesis, and less attention has been placed on regions such as the anterior hindbrain where RA synthesizing enzymes are absent. This study shows that a rich source of RA lies around the hindbrain from the RA synthetic enzyme retinaldehyde dehydrogenase-2 (RALDH2) present in the surrounding meninges and mesenchyme by embryonic day 13. RALDH2 is not distributed uniformly throughout the meninges but is restricted to territories over the developing hindbrain, suggesting that RA signaling may be localized to those regions. Further regulation of RA signaling is provided by the presence of a RA sink in the form of the CYP26B1 RA catabolic enzyme expressed in deeper regions of the brain. As a guide to the neural anatomy of hindbrain RA signaling, we used a mouse transgenic for a lacZ reporter gene driven by a RA response element (RAREhsplacZ) to identify regions of RA signaling. This reporter mouse provides evidence that RA signaling in the hindbrain after embryonic day 13 occurs in the regions of the cerebellum and precerebellar system adjacent to sources of RA, including the inferior olive and the pontine nuclei.

We report two cases of leptomeningeal angiomatosis in atypical frontoparietotemporal locations without an associated facial port-wine stain. Evidence of a leptomeningeal angioma was found in each when they were evaluated for headaches and seizures. The diagnosis of a leptomeningeal angioma was suggested by calcifications noted on computed tomographic scan of the head and confirmed with contrast-enhanced magnetic resonance images of the brain. We hypothesize that given the lack of occipital involvement with the angioma, and therefore the noncontiguous nature of this lesion with the developing upper facial ectoderm, the failure to develop a facial angioma would be expected. We found that the useof an anticonvulsant along with a migraine prophylactic medication appeared to have the greatest efficacy in these two cases, whereas anticonvulsants alone were less helpful. This diagnosis should be considered in any child presenting with seizures or complicated migraines and intracranial calcifications.

Migration of neuronal precursor cells from the external germinal layer (EGL) to the internal granular layer (IGL) is a crucial process in the development of the mammalian cerebellar cortex. These cells make up the only precursor population known to migrate away from the surface of the brain. We studied the role of the chemokine stromal-derived factor 1 (SDF-1) in the cerebellar tissue of rats and knockout mice and found (i) that it functions as an attractive guidance cue for neuronal migration and (ii) that its secretion from non-neuronal meningeal tissue is important for controlling the migration of embryonic EGL cells.

During mammalian evolution, expansion of the cerebral hemispheres was accompanied by expansion of the frontal and parietal bones of the skull vault and deployment of the coronal (fronto-parietal) and sagittal (parietal-parietal) sutures as major growth centres. Using a transgenic mouse with a permanent neural crest cell lineage marker, Wnt1-Cre/R26R, we show that both sutures are formed at a neural crest-mesoderm interface: the frontal bones are neural crest-derived and the parietal bones mesodermal, with a tongue of neural crest between the two parietal bones. By detailed analysis of neural crest migration pathways using X-gal staining, and mesodermal tracing by DiI labelling, we show that the neural crest-mesodermal tissue juxtaposition that later forms the coronal suture is established at E9.5 as the caudal boundary of the frontonasal mesenchyme. As the cerebral hemispheres expand, they extend caudally, passing beneath the neural crest-mesodermal interface within the dermis, carrying with them a layer of neural crest cells that forms their meningeal covering. Exposure of embryos to retinoic acid at E10.0 reduces this meningeal neural crest and inhibits parietal ossification, suggesting that intramembranous ossification of this mesodermal bone requires interaction with neural crest-derived meninges, whereas ossification of the neural crest-derived frontal bone is autonomous. These observations provide new perspectives on skull evolution and on human genetic abnormalities of skull growth and ossification.

The small lipid retinoic acid is known to promote neuronal differentiation in vitro and to act as a teratogen in the embryonic brain, but very little is known about the natural role of endogenously synthesized retinoic acid in forebrain development. Retinoic acid is synthesized mainly by three retinaldehyde dehydrogenases. We show here where the retinaldehyde dehydrogenases for the developing telencephalon are expressed and how their expression patterns change over developmental time. Retinoic acid diffusing from the retinaldehyde dehydrogenase sites is likely to influence the early telencephalon before the beginning of neurogenesis, as well as differentiation and radial migration of neurons into the cerebral cortex. Because of its diffusible character, retinoic acid represents a unique tool for the coordination of growth processes over an intermediate distance range in the developing telencephalon.

The pathogenesis of cranial and spinal dysraphia has been controversial. Studies of spinal dysraphia have shown that the relationships of the pia and dura to the cutaneous layers were best understood as the result of a primary abnormality of mesenchymal structures, with the nervous system lesions occurring as a result of exposure of the bare spinal cord on the body surface. This study was undertaken to determine if the relationship of the cutaneous layers in anencephaly were similar to those found in spinal dysraphia. We reviewed serial histologic sections of the cranial structures of 10 anencephalic fetuses autopsied at The Johns Hopkins Hospital. We found the dura to be continuous with the deep dermis and the pia continuous with the superficial dermis and epidermis, the same arrangement observed in myelomeningocele. The development of eyes and cranial nerves, the absence of a bony calvarium, and the meningeal-cutaneous relationships found in this study support the idea that anencephaly can originate as an abnormality of mesenchymal structures and that the brain is secondarily lost to injury in utero because of its exposed position.

We collected human fetal and adult normal meninges to relate the age of the tissue with the presence of collagenous and non-collagenous components of Extra Cellular Matrix (ECM). Immunohistochemistry led us to observe some differences in the amount and in the distribution of these proteins between the two sets of specimens. In particular, laminin and tenascin seem to be expressed more intensely in fetal meninges when compared to adult ones. In order to investigate whether the morphofunctional characteristics of fetal meninges may be represented in pathological conditions we also studied meningeal specimens from human meningiomas. Our attention was particularly focused on the expression of those non-collagenous proteins involved in nervous cell migration and neuronal morphogenesis as laminin and tenascin, which were present in lesser amount in normal adult specimens. Microscopical evidences led us to hipothesize that these proteins which are synthesized in a good amount during the fetal development of meninges can be newly produced in tumors. On the contrary, the role of tenascin and laminin in adult meninges is probably only interesting for their biophysical characteristics.

The origin of the ophthalmic artery exhibits several variants which can be explained by a vascular morphogenesis study. The particular relations of the ophthalmic artery with the optic nerve and the cranial meninges are examined using serial histological sections.

Serial examination of the cerebral hemispheres of 20 sudden infant death syndrome victims revealed high incidence of leukomalacia (40%), leptomeningeal glioneuronal heterotopias (70%) at the base of the cerebrum, and astrogliosis (65%) in the white matter and medulla reticular formation compared with 20 age-matched controls. These results suggest that an antepartum insult may become an important predisposing risk factor in some patients for sudden infant death syndrome.

Objective: The major morbidity of glioma lies in its infiltrative growth. One of the major patterns of this invasive growth is the formation of Scherer's secondary structures associated with the blood vessels and the leptomeninges. To better understand the role of extracellular matrix (ECM) in glioma invasion, we investigated in vitro the interaction between glioma cells and the meningeal mesenchymal tissue from the brain. As an aid to this study, ECM in glioma cell line spheroids was compared with that in primary fetal brain aggregates., Methods: To study the expression of ECM, four glioma cell lines (U-87 MG, U-251 MG, AN1/lac-z, and HF-66) and primary cells from fetal rat brain were grown as spheroids and monolayers. To sudy the role of ECM in glioma invasion, spheroids from the glioma cell lines were grown over established cultures of fetal meningeal and mesenchymal tissue. Expression of fibronectin, laminin, tenascin, collagen VI, and chondroitin sulfate proteoglycan was studied by immunofluorescence., Results: Expression of ECM by the spheroids was variable. U-87 MG expressed most of the ECM components robustly, whereas AN1/lac-z expressed them all weakly. Fetal rat brain aggregates produced minimal ECM. In cocultures of glioma spheroids and fetal meningeal mesenchymal tissue, individual cells from the glioma spheroids that expressed least fibronectin (AN1/lac-z and U-251 MG) migrated along the fibronectin-positive mesenchymal cells in the culture dish. Cells from the other two lines (U-87 MG and HF-66) that expressed fibronectin strongly did not demonstrate such behavior. None of the other ECM components showed a similar association; mesenchymal cells did not express laminin as strongly as fibronectin, and glioma cells were not observed to align with the laminin-positive structures., Conclusion: This study suggests that fibronectin may play a key role in intracerebral invasion of glioma cells.

Little is known about the development of the central nervous system (CNS) in humans. Ethical considerations preclude experimental studies in this field, and as a result most available data on human ontogenesis are descriptive. Comparative anatomic and embryologic studies have demonstrated that the main developmental milestones are conserved across species, and their results can be used to suggest a likely scenario for human development. The development of the ventricles, meninges, and choroid plexuses are discussed in this article. The central cavity of the neural tube is formed during neurulation, which occurs during the fourth gestational week. The first milestone is occlusion of the spinal neurocele (the central canal in the neural tube) shortly after neurulation. This prevents free communication between the ventricular system and the amniotic cavity. The second milestone is development of the meninges, which separate the central nervous system from the rest of the body. The embryonic origin of the meninges varies across species. In birds (and probably in mammals), the spinal meninges are derived from the somitic mesoderm, the brainstem meninges from the cephalic mesoderm, and the telencephalic meninges from the neural crest. Differentiation of the meninges, which involves formation of the subarachnoid space, occurs early, before the cerebrospinal fluid (CSF) begins to flow around the CNS. During ontogenesis, the meninges play a key role in regulating the growth of underlying nervous structures. They induce the formation of the superficial glial limiting layer and stimulate the growth of precursors located in the superficial blastemas of the cerebellum and hippocampus. The choroid plexuses are complex specialized structures that produce most of the CSF. Their epithelium derives from the neural tube epithelium and their mesenchyma from the meninges. Of the many enzymes produced in the choroid plexuses, some reflect the pivotal metabolic role of these structures (alkaline and acid phosphatases, magnesium-dependent ATPase, glucose-6-phosphatase, thiamine pyrophosphatase, adenylate cyclase, oxidoreductase, esterases, hydrolases, cathepsin D, and glutathion S-transferase). The two enzymes that are crucial to the production of CSF are Na+/K+ ATPase and carbonic anhydrase. Inactivation of catecholamines is mediated by catechol-O-methyltransferase and by the monoamine oxidases A and B. The morphology and synthesis profile of the choroid plexuses changes during development, although little is known about these changes in humans.

Recent in vivo investigations indicate that the mycotoxin ochratoxin A (OTA) is a neurotoxicant during prenatal stages. In line with in vivo data, in our embryonic chick brain and neural retina cell cultures the markers for neuritic outgrowth and differentiation (NF68 and 160 kDa, MAP2 and MAP5) were especially negatively affected. In vivo OTA is nearly completely bound to serum constituents. In our culture system binding of OTA to BSA evoked a significant shift of the concentration-effect relationships in meningeal and brain cell cultures. As a result of the albumin binding the OTA IC5 and IC50 values of all parameters increased by nearly the same value (about 15-fold in brain and 32-fold in meningeal cell cultures). One of the mechanisms responsible for OTA toxicity is thought to be the competitive inhibition versus Phe of Phe-dependent enzymes. Therefore, in addition, we investigated the effects of l-phenylalanine (Phe) and its influence on OTA toxicity in brain and neural retina cell cultures. Phe itself was found to differently affect brain and neural retina cell cultures. However, in both cultures OTA toxicity is not diminished by Phe. Therefore, our data indicate that at least in our cultures competition with Phe-dependent processes does not play a role in OTA toxicity., (Copyright 1997 Academic Press.)

Melatonin binding sites were identified over the leptomeninges surrounding the human fetal brain using quantitative in vitro autoradiography and the melatonin agonist, 2-[125I]iodomelatonin. Binding was found to be saturable and of high affinity (dissociation constant (Kd) = 54 pM and maximal theoretical binding (Bmax) = 13 fmol/mg protein), and inhibited by guanosine-5'-o-(3-thiotriphosphate) (GTPgammaS) suggesting that these binding sites represent G protein-coupled melatonin receptors. RT-PCR performed on mRNA isolated from the human fetal leptomeninges detected expression of the G protein-coupled melatonin receptor Mel1a, but not Mel1b. In situ hybridisation confirmed the localisation of Mel1a mRNA transcripts over the leptomeninges of the fetal brain. The identification of 2-[125I]iodomelatonin and Mel1a melatonin receptor expression in the fetal leptomeninges implies that melatonin may play a role in the early growth and development of the human brain.

We have studied the derivatives of the first somite using the quail-chick marking technique. After transplantation of the somite, the chick embryos were reincubated for periods ranging from 4 h to 11 days. Coronal and sagittal sections of the embryos were prepared for parallel staining with Feulgen-reaction, anti-quail antibody, anti-desmin antibody and QH-1 antibody. The first somite consists of an epithelial envelope surrounding somitocoele cells. Like other somites, it forms sclerotome, dermatome and myotome. Cells contribute to the occipital and parasphenoid bone, the meninges, the dermis in the occipital region and the pharyngeal connective tissue. The contribution of the first somite to bones, meninges, dermis and pharyngeal connective tissue is characterised by sharp anterior and posterior boundaries. In contrast, other derivatives such as connective tissue surrounding the vagus nerve, the carotid artery, and jugular vein exceed 10 to 18 segments. This is also true for myogenic cells participating in the formation of the cucullaris capitis muscle that extends from the temporal bone to the shoulder. In one third of the embryos, myocytes of the intrinsic laryngeal muscles are derived from the grafted first somite. Moreover, endothelial cells originate from this somite and migrate into the head (hind-brain, meninges, dermis), neck (pharynx, connective tissue surrounding the vagus nerve, carotid artery and jugular vein) and thorax. With respect to differentiation and derivatives the first somite is similar to other somites.

To identify regions involved in tissue specific regulation of transcription of the alpha1(VI) collagen chain, transgenic mice were generated carrying various portions of the gene's 5'-flanking sequence fused to the E. coli beta-galactosidase gene. Analysis of the transgene expression pattern by X-gal staining of embryos revealed that: (a) The proximal 0.6 kb of promoter sequence activated transcription in mesenchymal cells at sites of insertion of superficial muscular aponeurosis into the skin; tendons were also faintly positive. (b) The region between -4.0 and -5.4 kb from the transcription start site was required for activation of the transgene in nerves. It also drove expression in joints, in intervertebral disks, and in subepidermal and vibrissae mesenchyme. (c) The fragment comprised within -6.2 and -7.5 kb was necessary for high level transcription in skeletal muscle and meninges. Positive cells in muscle were mostly mononuclear and probably included connective tissue elements, although staining of myoblasts was not ruled out. This fragment also activated expression in joints, in intervertebral disks, and in subepidermal and vibrissae mesenchyme. (d) beta-Galactosidase staining in vibrissae induced by the sequences -4.0 to -5.4 and -6.2 to -7.5 was not coincident: with the latter sequence labeled nuclei were found mainly in the ventral and posterior quadrant, and, histologically, in the outer layers of mesenchyme surrounding and between the follicles, whereas with the former the remaining quadrants were positive and expressing cells were mostly in the inner layers of the dermal sheath. (e) Other tissues, notably lung, adrenal gland, digestive tract, which produce high amounts of collagen type VI, did not stain for beta-galactosidase. (f) Central nervous system and retina, in which the endogenous gene is inactive, expressed the lacZ transgene in most lines. The data suggest that transcription of alpha1(VI) in different tissues is regulated by distinct sequence elements in a modular arrangement, a mechanism which confers high flexibility in the temporal and spatial pattern of expression during development.

Meningeal fibroblasts express the proenkephalin gene during embryonal development but terminate the expression shortly before birth. When brought into primary culture at postnatal day 1, the fibroblasts again express the gene. Activation of protein kinase A reduces this expression and thus may contribute to its prenatal termination. Since the noradrenergic innervation of the meninges begins around the time of birth, it was investigated in the present study, how adrenergic agonists affected the levels of proenkephalin mRNA in cultured fibroblasts. The beta 2-adrenoceptor agonists salbutamol and procaterol increased the levels of endogenous cAMP and diminished the concentration of proenkephalin mRNA indicating that the cultured fibroblasts possessed this beta-subtype. In contrast, noradrenaline increased the level of proenkephalin mRNA in a concentration-dependent manner. This effect was independent of endogenous cAMP and was mediated by alpha 1-adrenoceptors. The data indicate that the noradrenergic innervation of the meninges at the time of birth is not responsible for the termination of the proenkephalin gene expression.

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.

LacZ reporter gene constructs were used to analyze the murine retinoic acid receptor beta (mRAR beta) gene promoter in transgenic mice. LacZ expression in transgenic mouse embryos with 250 bp of promoter sequences closely parallels that of RAR beta between embryonic days 8.5 and 12.5. This indicates that the -1 to -250 promoter region contains most regulatory elements required for tissue specific expression. Additional elements in the -250 to -625 region are required for high expression levels after day 12.5. Elements in the -625 to -3100 region are necessary to reproduce the RAR beta expression in the meninges and the eye mesenchyme. The expression pattern of the transgene and the endogenous RAR beta, as revealed by in situ hybridization, suggests an important role of the RAR beta in the developing nervous system.

Primary cultures of meningeal cells from embryonic rat cerebra secrete neurite growth-inducing components into serum-free culture medium. This conditioned medium (CM) was analyzed by FPLC and immunochemical and enzymatic treatments and tested for neurite promoting activity (NPA) in a quantitative bioassay using hippocampal neurons from embryonic rat. By immunoprecipitation or specific adsorption we identified laminin (LN)-proteoglycan complexes and fibronectin (FN), respectively, as the major neurite promoting components within meningeal cell CM. The LN-proteoglycan complexes and their NPA were sensitive to chondroitinase (chondroitin ABC lyase, EC 4.2.2.4) and to a smaller extent to heparitinase (heparitin sulfate lyase, EC 4.2.2.8). Minor fractions of the total NPA in CM correlated with free LN and a putative but not yet characterized FN-proteoglycan complex.

The expression of intermediate filament protein in human spinal cord arachnoid cells and ependyma was studied by immunohistochemistry and immunoblotting. Monoclonal antibodies specific for individual cytokeratin polypeptides indicated a developmental change in the presence of cytokeratin 8 and 18 in spinal leptomeninx and tanycytes of the spinal cord ependyma. While in fetal material cytokeratin 8 and 18 were abundant in arachnoid cells, in adults immunoreactivity was restricted to a few cells. Immunoblots prepared from adult as well as fetal arachnoid membranes showed significant amounts of cytokeratin 8. These findings indicate that although cytokeratin is represented in both fetal and adult arachnoid cells there is development regulation of its specific localization.

The development of the rat meninx from the viewpoint of cell proliferation was studied microscopically and immunohistochemically using bromodeoxyuridine (BUdR). A compact cell layer around the neural tube, the meninx primitiva, was observed in 12- and 13-day fetuses. A reticular structure resembling the subarachnoid space appeared in the 14-day fetus. The ectomeninx, consisting of a collagen fiber layer, part of which became the dura mater, appeared in 15-day fetuses, allowing discrimination of the endomeninx, the arachnoid cell layer. The primordium of the choroid plexus also appeared in the lateral ventricle on the same day. Bone appeared in the primitive dura mater, and stratification of the meninx was almost complete in 21-day fetuses. BUdR-positive cells were confirmed in the meninx from day 12 of gestation to day 15 postpartum. The number of BUdR-positive cells was greatest in fetuses aged about 12 or 13 days, reaching nearly 50%, but decreased gradually toward the neonatal period. The findings of this study suggest that, after the migration of neural crest cells, marked cell proliferation in the meninx begins. Differentiation into various layers then follows and is almost complete before birth, whereas the proliferation of arachnoid cells continues even in the early neonatal period.

The origin of cells covering the nervous system and the cutaneous receptors was studied using the quail-chick marking technique and light and electron microscopy. In the first experimental series the brachial neural tube of the quail was grafted in place of a corresponding neural tube segment of the chick embryo at HH-stages 10 to 14. In the second series the leg bud of quail embryos at HH-stages 18-20 was grafted in place of the leg bud of the chick embryos of the same stages and vice versa. It was found that all meningeal layers of the spinal cord, the perineurium and the endoneurium of peripheral nerves, as well as the capsular and inner space cells of Herbst sensory corpuscles, develop from the local mesenchymal cells. Schwann cells and cells of the inner core of sensory corpuscles are of neural crest origin. The precursors of Merkel cells migrate similarly to the Schwann cells into the limb bud where they later differentiate. This means that in addition to the Schwann cells and the melanocytes a further neural crest-derived subpopulation of cells enters the limb.