Evidence for intraventricular secretion of angiotensinogen and angiotensin by the subfornical organ using transgenic mice.

Direct intracerebroventricular injection of angiotensin II (ANG II) causes increases in blood pressure and salt and water intake, presumably mimicking an effect mediated by an endogenous mechanism. The subfornical organ (SFO) is a potential source of cerebrospinal fluid (CSF), ANG I, and ANG II, and thus we hypothesized that the SFO has a secretory function. Endogenous levels of angiotensinogen (AGT) and renin are very low in the brain. We therefore examined the immunohistochemical localization of angiotensin peptides and AGT in the SFO, and AGT in the CSF in two transgenic models that overexpress either human AGT (A+ mice), or both human AGT (hAGT) and human renin (SRA mice) in the brain. Measurements were made at baseline and following volumetric depletion of CSF. Ultrastructural analysis with immunoelectron microscopy revealed that superficially located ANG I/ANG II and AGT immunoreactive cells in the SFO were vacuolated and opened directly into the ventricle. Withdrawal of CSF produced an increase in AGT in the CSF that was accompanied by a large decline in AGT immunoreactivity within SFO cells. Our data provide support for the hypothesis that the SFO is a secretory organ that releases AGT and possibly ANG I/ANG II into the ventricle at least under conditions when genes that control the renin-angiotensin system are overexpressed in mice.

Characterization of Extensive Microstructural Variations Associated with Punctate White Matter Lesions in Preterm Neonates.

BACKGROUND AND PURPOSE: Punctate white matter lesions are common in preterm neonates. Neurodevelopmental outcomes of the neonates are related to the degree of extension. This study aimed to characterize the extent of microstructural variations for different punctate white matter lesion grades. MATERIALS AND METHODS: Preterm neonates with punctate white matter lesions were divided into 3 grades (from mild to severe: grades I-III). DTI-derived fractional anisotropy, axial diffusivity, and radial diffusivity between patients with punctate white matter lesions and controls were compared with Tract-Based Spatial Statistics and tract-quantification methods. RESULTS: Thirty-three preterm neonates with punctate white matter lesions and 33 matched controls were enrolled. There were 15, 9, and 9 patients, respectively, in grades I, II, and III. Punctate white matter lesions were mainly located in white matter adjacent to the lateral ventricles, especially regions lateral to the trigone, posterior horns, and centrum semiovale and/or corona radiata. Extensive microstructural changes were observed in neonates with grade III punctate white matter lesions, while no significant changes in DTI metrics were found for grades I and II. A pattern of increased axial diffusivity, increased radial diffusivity, and reduced/unchanged fractional anisotropy was found in regions adjacent to punctate white matter lesion sites seen on T1WI and T2WI. Unchanged axial diffusivity, increased radial diffusivity, and reduced/unchanged fractional anisotropy were observed in regions distant from punctate white matter lesion sites. CONCLUSIONS: White matter microstructural variations were different across punctate white matter lesion grades. Extensive change patterns varied according to the distance to the lesion sites in neonates with severe punctate white matter lesions. These findings may help in determining the outcomes of punctate white matter lesions and selecting treatment strategies.

Transitory cystic cavities in the developing mammalian brain - normal or anomalous?

Transitory cavities associated with the ventricular system represent probably one of the most unique features in the developing mammalian brain. In rodents, the cavities exist transiently in the developing brain and do not appear to be associated with any pathological events. Among the various cavities, the pyramidal-shaped cavum septum pellucidum (CSP) located beneath the corpus callosum and between the lateral ventricles is most well defined. In addition to the CSP are the bilateral subependymal cysts that are consistently associated with the third and fourth ventricles as well as the aqueduct. The cavities/cysts contain a large number of amoeboid microglia expressing surface receptors and hydrolytic enzymes common to tissue macrophages. The significance of these cavities in the developing brain remains a conjecture. Firstly, the cavity walls are free of an apparent epithelial lining; hence, it is speculated that the cavities that appear to communicate with the widened neighboring interstitial tissue spaces may have resulted from physical traction due to the rapid growth of the perinatal brain. Secondly, the cavities contain prominent clusters of amoeboid microglia that may be involved in clearing the debris of degenerating axons and cells resulting from the early brain tissue remodeling. With the increase in brain tissue compactness following the beginning of myelination in the second postnatal week, all cavities are obliterated; concomitantly, the number of amoeboid microglia in them diminishes and all this might signal further maturation of the brain.

Paravascular channels, cisterns, and the subarachnoid space in the rat brain: A single compartment with preferential pathways.

Recent evidence suggests an extensive exchange of fluid and solutes between the subarachnoid space and the brain interstitium, involving preferential pathways along blood vessels. We studied the anatomical relations between brain vasculature, cerebrospinal fluid compartments, and paravascular spaces in male Wistar rats. A fluorescent tracer was infused into the cisterna magna, without affecting intracranial pressure. Tracer distribution was analyzed using a 3D imaging cryomicrotome, confocal microscopy, and correlative light and electron microscopy. We found a strong 3D colocalization of tracer with major arteries and veins in the subarachnoid space and large cisterns, attributed to relatively large subarachnoid space volumes around the vessels. Confocal imaging confirmed this colocalization and also revealed novel cisternal connections between the subarachnoid space and ventricles. Unlike the vessels in the subarachnoid space, penetrating arteries but not veins were surrounded by tracer. Correlative light and electron microscopy images indicated that this paravascular space was located outside of the endothelial layer in capillaries and just outside of the smooth muscle cells in arteries. In conclusion, the cerebrospinal fluid compartment, consisting of the subarachnoid space, cisterns, ventricles, and para-arteriolar spaces, forms a continuous and extensive network that surrounds and penetrates the rat brain, in which mixing may facilitate exchange between interstitial fluid and cerebrospinal fluid.

AMPA-Kainate Receptor Inhibition Promotes Neurologic Recovery in Premature Rabbits with Intraventricular Hemorrhage.

Intraventricular hemorrhage (IVH) in preterm infants leads to cerebral inflammation, reduced myelination of the white matter, and neurological deficits. No therapeutic strategy exists against the IVH-induced white matter injury. AMPA-kainate receptor induced excitotoxicity contributes to oligodendrocyte precursor cell (OPC) damage and hypomyelination in both neonatal and adult models of brain injury. Here, we hypothesized that IVH damages white matter via AMPA receptor activation, and that AMPA-kainate receptor inhibition suppresses inflammation and restores OPC maturation, myelination, and neurologic recovery in preterm newborns with IVH. We tested these hypotheses in a rabbit model of glycerol-induced IVH and evaluated the expression of AMPA receptors in autopsy samples from human preterm infants. GluR1-GluR4 expressions were comparable between preterm humans and rabbits with and without IVH. However, GluR1 and GluR2 levels were significantly lower in the embryonic white matter and germinal matrix relative to the neocortex in both infants with and without IVH. Pharmacological blockade of AMPA-kainate receptors with systemic NBQX, or selective AMPA receptor inhibition by intramuscular perampanel restored myelination and neurologic recovery in rabbits with IVH. NBQX administration also reduced the population of apoptotic OPCs, levels of several cytokines (TNFα, IL-ß, IL-6, LIF), and the density of Iba1(+) microglia in pups with IVH. Additionally, NBQX treatment inhibited STAT-3 phosphorylation, but not astrogliosis or transcription factors regulating gliosis. Our data suggest that AMPA-kainate receptor inhibition alleviates OPC loss and IVH-induced inflammation and restores myelination and neurologic recovery in preterm rabbits with IVH. Therapeutic use of FDA-approved perampanel treatment might enhance neurologic outcome in premature infants with IVH. SIGNIFICANCE STATEMENT: Intraventricular hemorrhage (IVH) is a major complication of prematurity and a large number of survivors with IVH develop cerebral palsy and cognitive deficits. The development of IVH leads to inflammation of the periventricular white matter, apoptosis and arrested maturation of oligodendrocyte precursor cells, and hypomyelination. Here, we show that AMPA-kainate receptor inhibition by NBQX suppresses inflammation, attenuates apoptosis of oligodendrocyte precursor cells, and promotes myelination as well as clinical recovery in preterm rabbits with IVH. Importantly, AMPA-specific inhibition by the FDA-approved perampanel, which unlike NBQX has a low side-effect profile, also enhances myelination and neurological recovery in rabbits with IVH. Hence, the present study highlights the role of AMPA-kainate receptor in IVH-induced white matter injury and identifies a novel strategy of neuroprotection, which might improve the neurological outcome for premature infants with IVH.

Defective motile cilia in Prickle2-deficient mice.

Motile cilia play diverse roles across phyla and cell types, and abnormalities in motile cilia lead to numerous disease states, including hydrocephalus. Although motile ciliary abnormalities in Prickle2 mutants have not yet been described, the planar cell polarity genes, including Prickle2, are implicated in the development and function of motile cilia. This report evaluates Prickle2-deficient mice for dysfunction in processes known to depend on functioning motile cilia. Prickle2-deficient mice do not develop hydrocephalus, but do display abnormal morphology and motility in the motile cilia of the ependyma. The morphology of tracheal motile cilia is also abnormal. Taken together, these results demonstrate that Prickle2 is required for normal ependymal motile cilia development and function.

[Effect of single or combined application of UDP-glucose, GDNF and memantine on improvement of white matter injury in neonatal rats assessed with light and electron microscopy pathologically].

OBJECTIVE: To evaluate pathologically the effect of the single or combined application of UDP-glucose, GDNF and memantine on the improvement of white matter injury in neonatal rats with periventricular leukomalacia (PVL) under light and electron microscopy. METHODS: A five-day-old neonatal rat model for PVL was established by ligation of the lateral common carotid artery following 120-minute hypoxia. Rats were randomly divided into six groups (30 rats in each group): sham-operated, PVL, UDP-glucose (UDP-glucose 2000 mg/kg intraperitoneally after PVL), GDNF (GDNF 100 µg/kg intracerebrally after PVL), tmemantine (memantine 20 mg/kg intraperitoneally after PVL), and a combination administration of three drugs (UDP-glucose, GDNF and memantine). The rats were sacrificed 7 or 21 days after PVL for assessment of pathological changes in the white matter under both light and electron microscopy. The number and thickness of the myelin sheath in the white matter were measured under electron microscopy, and both of pathological grading and scoring were undertaken under light microscopy. RESULTS: There was rare and sparse myelinogenesis with a loose arrangement of nerve fibers in the white matter under electron microscopy in the PVL group at 7 and 21 days after PVL. The number and thickness of the myelin sheath in the PVL group were significantly less than in the sham-operated, UDP-glucose, GDNF, memantine and combination administration groups (P<0.01). The results of pathological grading of white matter under light microscopy showed that all rats in the PVL group manifested either mild injury (38%-50%) or severe injury (50%-62%) at 7 and 21 days after PVL. The majority of rats (50%-88%) in the four drug administration groups had normal white matter at 7 and 21 days after PVL. The pathological scores at 7 and 21 days after PVL in the PVL group were the highest, and they were significantly higher than in the other five groups (P<0.05). CONCLUSIONS: The single or combined application of UDP-glucose, GDNF and memantine may significantly improve pathological changes in the white matter of rats with PVL. The favorable effect is inferred to be closely correlated with the improvement of brain microenvironment and the enhancement of nerve regeneration promoted by the three drugs.

Morphological and behavioral changes in the pathogenesis of a novel mouse model of communicating hydrocephalus.

The Ro1 model of hydrocephalus represents an excellent model for studying the pathogenesis of hydrocephalus due to its complete penetrance and inducibility, enabling the investigation of the earliest cellular and histological changes in hydrocephalus prior to overt pathology. Hematoxylin and eosin staining, immunofluorescence and electron microscopy were used to characterize the histopathological events of hydrocephalus in this model. Additionally, a broad battery of behavioral tests was used to investigate behavioral changes in the Ro1 model of hydrocephalus. The earliest histological changes observed in this model were ventriculomegaly and disorganization of the ependymal lining of the aqueduct of Sylvius, which occurred concomitantly. Ventriculomegaly led to thinning of the ependyma, which was associated with periventricular edema and areas of the ventricular wall void of cilia and microvilli. Ependymal denudation was subsequent to severe ventriculomegaly, suggesting that it is an effect, rather than a cause, of hydrocephalus in the Ro1 model. Additionally, there was no closure of the aqueduct of Sylvius or any blockages within the ventricular system, even with severe ventriculomegaly, suggesting that the Ro1 model represents a model of communicating hydrocephalus. Interestingly, even with severe ventriculomegaly, there were no behavioral changes, suggesting that the brain is able to compensate for the structural changes that occur in the pathogenesis of hydrocephalus if the disorder progresses at a sufficiently slow rate.

Molecular heterogeneity in the choroid plexus epithelium: the 22-member γ-protocadherin family is differentially expressed, apically localized, and implicated in CSF regulation.

The choroid plexus (CP) epithelium develops from the ependyma that lines the ventricular system, and plays a critical role in the development and function of the brain. In addition to being the primary site of CSF production, the CP maintains the blood-CSF barrier via apical tight junctions between epithelial cells. Here we show that the 22-member γ-protocadherin (γ-Pcdh) family of cell adhesion molecules, which we have implicated previously in synaptogenesis and neuronal survival, is highly expressed by both CP epithelial and ependymal cells, in which γ-Pcdh protein localization is, surprisingly, tightly restricted to the apical membrane. Multi-label immunostaining demonstrates that γ-Pcdhs are excluded from tight junctions, basolateral adherens junctions, and apical cilia tufts. RT-PCR analysis indicates that, as a whole, the CP expresses most members of the Pcdh-γ gene family. Immunostaining using novel monoclonal antibodies specific for single γ-Pcdh proteins shows that individual epithelial cells differ in their apically localized γ-Pcdh repertoire. Restricted mutation of the Pcdh-γ locus in the choroid plexus and ependyma leads to significant reductions in ventricular volume, without obvious disruptions of epithelial apical-basal polarity. Together, these results suggest an unsuspected role for the γ-Pcdhs in CSF production and demonstrate a surprising molecular heterogeneity in the CP epithelium.

Sustained activation of mTOR pathway in embryonic neural stem cells leads to development of tuberous sclerosis complex-associated lesions.

Tuberous Sclerosis Complex (TSC) is a multisystem genetic disorder characterized by hamartomatous neurological lesions that exhibit abnormal cell proliferation and differentiation. Hyperactivation of mTOR pathway by mutations in either the Tsc1 or Tsc2 gene underlies TSC pathogenesis, but involvement of specific neural cell populations in the formation of TSC-associated neurological lesions remains unclear. We deleted Tsc1 in Emx1-expressing embryonic telencephalic neural stem cells (NSCs) and found that mutant mice faithfully recapitulated TSC neuropathological lesions, such as cortical lamination defects and subependymal nodules (SENs). These alterations were caused by enhanced generation of SVZ neural progeny, followed by their premature differentiation and impaired maturation during both embryonic and postnatal development. Notably, mTORC1-dependent Akt inhibition and STAT3 activation were involved in the reduced self-renewal and earlier neuronal and astroglial differentiation of mutant NSCs. Thus, finely tuned mTOR activation in embryonic NSCs may be critical to prevent development of TSC-associated brain lesions.

Oleic acid synthesized in the periventricular zone promotes axonogenesis in the striatum during brain development.

Our previous works showed that oleic acid synthesized in vitro by astrocytes in response to albumin behaves as a neurotrophic factor in neurons, up-regulating several proteins, such as the axonal growth marker growth-associated protein 43(GAP-43). Although the molecular mechanism of this process is fairly known, there is no evidence pinpointing the region/s in which oleic acid is synthesized. In this study, we show that the rate-limiting enzyme in oleic acid synthesis, stearoyl-CoA desaturase (SCD-1), is located in the periventricular zone of the brain of newborn rats, simultaneously to an increase in the amount of free oleic acid in the forebrain. In addition, the spatio-temporal presence of albumin - the signal that promotes oleic acid synthesis - and that of GAP-43 are correlated with that of SCD-1. Using organotypic slice cultures, we found that albumin up-regulates SCD-1 and stimulates the growth of GAP-43-positive axons in the striatum. The effect of albumin on GAP-43 was reduced when SCD-1 was silenced by siRNA. In conclusion, our results suggest that albumin up-regulates axonogenesis in the striatum by increasing the amount of the neurotrophic factor oleic acid synthesized by SCD-1 in the periventricular zone of the newborn brain.

Characteristic features of a nerve primo-vessel suspended in rabbit brain ventricle and central canal.

Bonghan theory was proposed by Bonghan Kim to illustrate the anatomy and physiology of the acupuncture meridian system. One of his astonishing claims was the physical presence of the nerve primo-vessel, which can be involved with a regenerating system of nerves. Our previous work has shown that there is a nerve primo-vessel in brain ventricles and the central canal of the spine of a rabbit. In this study, confocal laser scanning microscopy, transmission electron microscopy, and high voltage electron microscopy demonstrated that a nerve primo-vessel comprised DNA particles, other microparticles, and rod-shaped nuclei encircled by helix-shaped actins. The nerve primo-vessel had acridine orange-stained DNA particles that varied in size and were in parallel. These characteristics of the nerve primo-vessel are crucial for a comprehensive understanding of their function in the central nervous system, which may be associated with nerve regeneration.

Embryonic exposure to ethanol disturbs regulation of mitotic spindle orientation via GABA(A) receptors in neural progenitors in ventricular zone of developing neocortex.

Neural progenitors in the ventricular zone of the developing neocortex divide oriented either parallel or perpendicular to the ventricular surface based on their mitotic spindle orientation. It has been shown that the cleavage plane orientation is developmentally regulated and plays a crucial role in cell fate determination of neural progenitors or the maintenance of the proliferative ventricular zone during neocortical development. We tested if fetal exposure to ethanol, the most widely used psychoactive agent and a potent teratogen that may cause malformation in the central nervous system, alters mitotic cleavage orientation of the neural progenitors at the apical surface of the ventricular zone in the developing neocortex. Fetal exposure to ethanol on E10.5 and 11.5 increased the occurrence frequency of a horizontal cleavage plane that is parallel to the ventricular surface on E 12.5. Administration of picrotoxin, a GABA(A) receptor antagonist, prior to ethanol administration canceled the effect of ethanol with the frequency of horizontal division similar to the control level, although picrotoxin itself did not show any effect on cleavage plane orientation. Phenobarbital, a GABA(A) receptor agonist, induced horizontal cleavage to an extent similar to that induced by ethanol administration. (+)MK801, an antagonist of NMDA receptor that is another major target of ethanol in neural cells, did not affect the cleavage plane of dividing progenitors. These results suggest that fetal ethanol exposure induced alterations in the cleavage plane orientation of neural progenitors in the ventricular zone of the neocortex via the enhancement of the function of GABA(A) receptors.

Proliferation in the human ipsilateral subventricular zone after ischemic stroke.

BACKGROUND: It is uncertain whether neurogenesis occurs in humans after stroke. We studied the morphologic changes that occurred in the subventricular zone (SVZ) in patients who died following an acute ischemic stroke. METHODS: We examined coronal brain slices from patients who died after a first-ever cerebral nonlacunar infarction in the middle cerebral artery territory. We evaluated the morphologic changes in the ipsilateral and contralateral SVZ by light and electron microscopy. Using immunochemistry with Ki-67 and PCNA, we detected cell proliferation. We used Tuj-1 for immature neurons and PSA-NCAM for migrating cells. RESULTS: The study included 7 patients with a mean age of 82 +/- 5 (mean +/- SD) years; 4 were men. They died a mean of 10 +/- 5 days after the ischemic stroke. Brain samples were obtained a mean of 4 +/- 2 hours after death. In comparison with the contralateral SVZ, the following changes were observed in the ipsilateral SVZ: an increase in the width of the gap and ribbon layers, as well as in the cell density of the ribbon layer, an enlargement of the cytoplasmic volume of astrocytes, and an increase of Ki-67-positive cells. In the ipsilateral SVZ, mitoses and cells that stained for either Tuj-1 or PSA-NCAM markers were observed more frequently than in the contralateral SVZ. CONCLUSION: We found unequivocal evidence of active cell proliferation in the ipsilateral subventricular zone following an acute ischemic stroke in our patients.

Strain differences of cerebral ventricles in mice: can the MRL/MpJ mouse be a model for hydrocephalus?

Hydrocephalus is an intractable disease characterized by the excessive accumulation of cerebrospinal fluid (CSF) in the cerebral ventricles. There are many cases in both human and animals; however, the cause and mechanism of it's development is not clearly understood. In this study, differences of cerebral ventricles in 5 inbred mice strains (MRL/MpJ, C57BL/6, C3H/He, DBA/2 and BALB/c) were investigated by histological techniques to determine the possibility of a new animal model for hydrocephalus. Our analysis showed that significant differences in the volume and the surface area of lateral ventricles in the 5 inbred strains, with MRL/MpJ mice having the largest lateral, third, aqueduct and fourth ventricles. In addition, when MRL/MpJ mice were compared to BALB/c mice on 0 day after birth, the former already had larger lateral ventricles than the latter. Although there were no significant difference in the ratios of ependymal cell types in MRL/MpJ mice and BALB/c mice, the number and the diameter of lipid droplets in MRL/MpJ mice were, interestingly, smaller than those in BALB/c mice. It is well known that ependymal cells absorb nutritional substances in CSF by endocytosis, suggesting the possibility that their decrease may relate to the larger cerebral ventricles in MRL/MpJ. In conclusion, MRL/ MpJ mice have greater volumes in cerebral ventricles than other strains and may be useful for a model showing high susceptibility to hydrocephalus.

Periventricular white matter damage in the hypoxic neonatal brain: role of microglial cells.

Periventricular white matter damage (PWMD) also known as periventricular white matter injury, is one of the major causes of neurological impairment in premature newborns. The etiology of white matter injury is multifaceted with hypoxia-ischemia being an important underlying factor. The developing oligodendrocytes are susceptible to damage resulting in myelination deficits. Excess release of glutamate, free radical production, release of cytokines and iron accumulation are factors thought to mediate damage to the developing white matter. Recent studies have also suggested a role for vascular endothelial growth factor and nitric oxide in the pathogenesis of PWMD. Although the role of microglial cells in the development of PWMD is still debatable, our recent investigations have shed some light on their involvement in the pathogenesis of PWMD. Challenges for the future include in-depth investigation of crosstalk between microglia and immature oligodendrocytes as well as other glial cells and vascular endothelial cells.

A comparison of epithelial and neural properties in progenitor cells derived from the adult human ciliary body and brain.

Cells isolated from the ciliary body (CB) of the adult human eye possess properties of retinal stem/progenitor cells and can be propagated as spheres in culture. As these cells are isolated from a non-neural epithelium which has neuroepithelial origin, they may have both epithelial and neural lineages. Since it is the properties of neural progenitor cells that are sought after in a future scenario of autotransplantation, we wanted to directly compare human CB spheres with neurospheres derived from the human subventricular zone (SVZ), which is the best characterized neural stem cell niche in the CNS of adults. The CB epithelium was dissected from donor eyes (n = 8). Biopsies from the ventricular wall were harvested during neurosurgery due to epilepsy (n = 7). CB and SVZ tissue were also isolated from Brown Norwegian rats. Dissociated single cells were cultivated in a sphere-promoting medium and passaged every 10-30 days. Fixed spheres were studied by immunohistochemistry, quantitative RT-PCR and scanning/transmission electron microscopy. We found that both CB and SVZ spheres contained a mixed population of cells embedded in extracellular matrix. CB spheres, in contrast to SVZ neurospheres, contained pigmented cells with epithelial morphology that stained for cytokeratins (3/12 + 19), were connected through desmosomes and tight-junctions and produced PEDF. Markers of neural progenitors (nestin, Sox-2, GFAP) were significantly lower expressed in human CB compared to SVZ spheres, and nestin positive cells in the CB spheres also contained pigment. There was higher expression of EGF and TGF-beta receptors in human CB spheres, and a comparative greater activation of the canonical Wnt pathway. These results indicate that adult human CB spheres contain progenitor cells with epithelial properties and limited expression of neural progenitor markers compared to CNS neurospheres. Further studies mapping the regulation between epithelial and neural properties in the adult human CB spheres are vital to fully utilize them as a clinical source of retinal progenitor cells in the future.

Distribution of beta-dystroglycan immunopositive globules in the subventricular zone of rat brain.

Present study demonstrates a novel localization of beta-dystroglycan in rat brain. Beside the meningeal surface and the cerebral vessels where beta-dystroglycan immunopositivity has been described, now immunopositive solid bodies were found at the basis of ependymocytes. Since they proved to be round in either coronal, sagittal, and horizontal section, revealing their globular shape, they received the term 'globules.' Double immunofluorescent labeling of GFAP (glial fibrillary acidic protein) and beta-dystroglycan revealed that 'globules' are within the 'cordon' of subventricular astrocytes. The 'globules' were found throughout the ventricular system, except the ventral part and floor of the third ventricle. The latter one comprised the median eminence and extended caudally to the inframamillary recess. The area where the 'globules' were missing, corresponded to that where ependymocytes were replaced by GFAP-immunopositive tanycytes. Utrophin and alpha-dystrobrevin were co-localized with dystroglycan, whereas alpha1-syntrophin was detected along the borders of the ependymal cells. Comparing our results to former data, the 'globules' seem to be the anchoring places of the 'fractons' formed by laminin [Mercier et al. (2003) J Comp Neurol 455:324-340], which interconnect the vascular basal lamina and the ependymal layer. The ependymal localization of the 'globules' was proved by pre-embedding electron microscopic immunohistochemistry. The 'globules' proved to be a labyrinthine structure, which seemed to be identical with the 'basal membrane labyrinth' described by Leonhardt [Leonhardt (1970) Z Zellforsch Mikrosk Anat 105:395-404]. The description of the beta-dystroglycan-immunopositive 'globules' contributes to the better understanding of the structure of the subventricular zone where neurogenesis can occur during adulthood and provides an important link to the meningeo-glial network.

Mutations in Hydin impair ciliary motility in mice.

Chlamydomonas reinhardtii hydin is a central pair protein required for flagellar motility, and mice with Hydin defects develop lethal hydrocephalus. To determine if defects in Hydin cause hydrocephalus through a mechanism involving cilia, we compared the morphology, ultrastructure, and activity of cilia in wild-type and hydin mutant mice strains. The length and density of cilia in the brains of mutant animals is normal. The ciliary axoneme is normal with respect to the 9 + 2 microtubules, dynein arms, and radial spokes but one of the two central microtubules lacks a specific projection. The hydin mutant cilia are unable to bend normally, ciliary beat frequency is reduced, and the cilia tend to stall. As a result, these cilia are incapable of generating fluid flow. Similar defects are observed for cilia in trachea. We conclude that hydrocephalus in hydin mutants is caused by a central pair defect impairing ciliary motility and fluid transport in the brain.