Processes Of Radial Glial Cells Research Articles

The role of oligodendrocyte progenitor cells in the spatiotemporal vascularization of the human and mouse neocortex.

Brain vasculature formation begins with vessel invasion from the perineural vascular plexus, which expands through vessel sprouting and growth. Recent studies have indicated the existence of oligodendrocyte-vascular crosstalk during development. However, the relationship between oligodendrocyte progenitor cells (OPCs) and the ordered spatiotemporal vascularization of the neocortex has not been elucidated. Our findings suggest that OPCs play a complex role in the vessel density of the embryonic and postnatal neocortex. Analyses of normal human and mouse embryonic cerebral cortex show that vascularization and OPC distribution are tightly controlled in a spatially and temporally restricted manner, exhibiting a positive correlation. Loss of OPCs at both embryonic and postnatal stages led to a reduction in vascular density, suggesting that OPC populations play a role in vascular density. Nonetheless, dynamic observation on cultured brain slices and staining of tissue sections indicate that OPC migration is unassociated with the proximity to blood vessels, primarily occurring along radial glial cell processes. Additionally, in vitro experiments demonstrate that OPC secretions promote vascular endothelial cell (VEC) growth. Together, these observations suggest that vessel density is influenced by OPC secretions.

ATF5 deficiency causes abnormal cortical development

Activating transcription factor 5 (ATF5) is a member of the cAMP response element binding protein (CREB)/ATF family of basic leucine zipper transcription factors. We previously reported that ATF5-deficient (ATF5−/−) mice exhibited behavioural abnormalities, including abnormal social interactions, reduced behavioural flexibility, increased anxiety-like behaviours, and hyperactivity in novel environments. ATF5−/− mice may therefore be a useful animal model for psychiatric disorders. ATF5 is highly expressed in the ventricular zone and subventricular zone during cortical development, but its physiological role in higher-order brain structures remains unknown. To investigate the cause of abnormal behaviours exhibited by ATF5−/− mice, we analysed the embryonic cerebral cortex of ATF5−/− mice. The ATF5−/− embryonic cerebral cortex was slightly thinner and had reduced numbers of radial glial cells and neural progenitor cells, compared to a wild-type cerebral cortex. ATF5 deficiency also affected the basal processes of radial glial cells, which serve as a scaffold for radial migration during cortical development. Further, the radial migration of cortical upper layer neurons was impaired in ATF5−/− mice. These results suggest that ATF5 deficiency affects cortical development and radial migration, which may partly contribute to the observed abnormal behaviours.

Talpid3-Mediated Centrosome Integrity Restrains Neural Progenitor Delamination to Sustain Neurogenesis by Stabilizing Adherens Junctions

Neurogenesis in the developing neocortex relies on extensive mitosis of radial glial cells (RGCs) in the apical surface. The nuclear migration of epithelial-like RGCs is fundamentally important for proper mitosis, but how the apical processes of RGCs are anchored to ensure the nucleokinetic behavior of RGCs remains unclear. Here we find that Talpid3, related to Joubert syndrome, is localized to the mother centriole of RGCs and is required for their apical mitosis. Genetic silencing of Talpid3 causes abnormal RGC delamination and thereby impairs their interkinetic nuclear migration in both cell-autonomous and non-autonomous manners. Further analyses reveal that Talpid3 associates with Ninein to regulate microtubule organization and maintain the integrity of adherens junctions to anchor RGCs. Moreover, genetic ablation of Talpid3 results in synchronized, ectopic mitosis of neural progenitors and dysregulated neurogenesis. Our study provides an intriguing perspective for the non-ciliogenic role of centriolar proteins in mediating cortical neurogenesis.

The association of microcephaly protein WDR62 with CPAP/IFT88 is required for cilia formation and neocortical development.

WDR62 mutations that result in protein loss, truncation or single amino-acid substitutions are causative for human microcephaly, indicating critical roles in cell expansion required for brain development. WDR62 missense mutations that retain protein expression represent partial loss-of-function mutants that may therefore provide specific insights into radial glial cell processes critical for brain growth. Here we utilized CRISPR/Cas9 approaches to generate three strains of WDR62 mutant mice; WDR62 V66M/V66M and WDR62R439H/R439H mice recapitulate conserved missense mutations found in humans with microcephaly, with the third strain being a null allele (WDR62stop/stop). Each of these mutations resulted in embryonic lethality to varying degrees and gross morphological defects consistent with ciliopathies (dwarfism, anophthalmia and microcephaly). We find that WDR62 mutant proteins (V66M and R439H) localize to the basal body but fail to recruit CPAP. As a consequence, we observe deficient recruitment of IFT88, a protein that is required for cilia formation. This underpins the maintenance of radial glia as WDR62 mutations caused premature differentiation of radial glia resulting in reduced generation of neurons and cortical thinning. These findings highlight the important role of the primary cilium in neocortical expansion and implicate ciliary dysfunction as underlying the pathology of MCPH2 patients.

Expression of the RNA methyltransferase Nsun5 is essential for developing cerebral cortex

Nsun5 gene, encoding a cytosine-5 RNA methyltransferase, is deleted in about 95% patients with Williams-Beuren syndrome (WBS). WBS is a neurodevelopmental disorder and characterized by cognitive disorder. We generated single-gene Nsun5 knockout (Nsun5-KO) mice and reported that the Nsun5 deletion leads to deficit in spatial cognition. This study focused on investigating the influence of Nsun5 deficiency in the development of cerebral cortex. In comparison with wild-type littermates, the cortical thickness in postnatal day 10 Nsun5-KO mice was obviously reduced with an abnormal laminar organization, and the processes of pyramidal cells were shorter and finer. Nsun5 was selectively expressed in radial glial cells (RGCs) of cerebral cortex from embryonic day (E) 12.5 to E16.5, but not in intermediate progenitor cells (IPCs) or neocortical neurons. The Nsun5 deletion did not alter proliferation of RGCs or differentiation of RGCs into IPCs. Notably, the ablation of Nsun5 disrupted the growth of radial glial scaffolds, thus numerous basal processes of RGCs failed to reach pial basement membrane. Level of cell polarity regulator Cdc42 protein in radial glial scaffolds of E14.5 Nsun5-KO mice was reduced, but the level of Cdc42 mRNA was unchanged. The dysfunction of glial scaffolds impeded the radial migration of upper-layer and deeper-layer neurons to cause their subcortical accumulation and apoptosis, resulting in an obvious thinness of the cortical plate in E18.5 Nsun5-KO mice. These findings establish a critical role of Nsun5 in development of cerebral cortex through regulating radial glial scaffolds of RGCs to control migration of neocortical neurons.

Proteomic profiling reveals dopaminergic regulation of progenitor cell functions of goldfish radial glial cells in vitro

Radial glial cells (RGCs) are stem-like cells found in the developing and adult central nervous system. They function as both a scaffold to guide neuron migration and as progenitor cells that support neurogenesis. Our previous study revealed a close anatomical relationship between dopamine neurons and RGCs in the telencephalon of female goldfish. In this study, label-free proteomics was used to identify the proteins in a primary RGC culture and to determine the proteome response to the selective dopamine D1 receptor agonist SKF 38393 (10μM), in order to better understand dopaminergic regulation of RGCs. A total of 689 unique proteins were identified in the RGCs and these were classified into biological and pathological pathways. Proteins such as nucleolin (6.9-fold) and ependymin related protein 1 (4.9-fold) were increased in abundance while proteins triosephosphate isomerase (10-fold) and phosphoglycerate dehydrogenase (5-fold) were decreased in abundance. Pathway analysis revealed that proteins that consistently changed in abundance across biological replicates were related to small molecules such as ATP, lipids and steroids, hormones, glucose, cyclic AMP and Ca2+. Sub-network enrichment analysis suggested that estrogen receptor signaling, among other transcription factors, is regulated by D1 receptor activation. This suggests that these signaling pathways are correlated to dopaminergic regulation of radial glial cell functions. Most proteins down-regulated by SKF 38393 were involved in cell cycle/proliferation, growth, death, and survival, which suggests that dopamine inhibits the progenitor-related processes of radial glial cells. Examples of differently expressed proteins including triosephosphate isomerase, nucleolin, phosphoglycerate dehydrogenase and capping protein (actin filament) muscle Z-line beta were validated by qPCR and western blot, which were consistent with MS/MS data in the direction of change. This is the first study to characterize the RGC proteome on a large scale in a vertebrate species. These data provide novel insight into glial protein networks that are associated with neuroendocrine function and neurogenesis in the teleost brain. Biological significanceWhile the role of radial glial cells in organizing brain structure and neurogenesis has been well studied, protein profiling experiments in this unique cell type has not been conducted. This study is the first to profile the proteome of goldfish radial glial cells in culture and to study the regulation of progenitor functions of radial glial cells by the neurotransmitter dopamine. This study provides the foundation for molecular network analysis in fish radial glial cells, and identifies cellular processes and signaling pathways in these cells with roles in neurogenesis and neuroendocrine function. Lastly, this study begins to characterize signatures and biomarkers for specific neuroendocrine and neurogenesis disruptors.

Vasculature-guided neural migration in mouse cerebellum

Neural migration is regarded as a key step for cortical development and cortical lamination. While classical theory holds that immature neurons migrate to their destinations along radial glia, there is some preliminary evidence showing that vasculature is probably involved in that process as well. In this study, we compared vascular development with that of radial glia and the migration of neurons, so that the relationships among them could be elucidated. We found that the radial glial cells and vasculature were highly similar in their distribution and development. For instance, in the external granular layer (EGL, the putative molecular layer), the processes of radial glial cells were arranged in an orderly radial pattern, whereas in the deep areas of the cerebellum, their arrangements were in relative disorder. On the other hand, the vasculatures in EGL were usually orientated radially, which paralleled the projections of radial glia; however, the distribution of vasculature in the internal granular layer (IGL, the putative granular layer) and white matter was in relative disorder as well, similar to the pattern exhibited by radial glial processes. This high harmonization between vasculature and radial glia suggests their interconnected biological relationships during cerebellar development and neural migration. We also observed that a large number of newborn neurons were migrating along blood vessels, suggesting that the vasculature could serve as a scaffold for cell migration. In conclusion, cerebellar vasculature could guide neural migration not only as a platform for biological interaction with radial glia, but also serve scaffolding functions for neural migration.

Reelin Induces Branching of Neurons and Radial Glial Cells during Corticogenesis.

Newborn neurons migrate along the processes of radial glial cells (RGCs) to reach their final positions in the cortex. Here, we visualized individual migrating neurons and RGCs using in utero electroporation. We show that branching of migrating neurons and RGCs is closely correlated spatiotemporally with the distribution of Reelin. Time-lapse imaging revealed that the leading processes of migrating neurons gave rise to increasingly more branches once their growth cones contacted the Reelin-containing marginal zone. This was accompanied by translocation of the nucleus and gradual shortening of the leading process. Absence of Reelin in reeler mice altered these processes resulting in misorientation, loss of bipolarity, and aberrant migration of cortical neurons. Moreover, in reeler, the branching of the basal processes of RGCs in the marginal zone was severely disrupted. Consistent with previous reports, we show that in dissociated reeler cortical cultures, exposure to recombinant Reelin enhanced dendritic complexity and glial branching. Our results suggest that Reelin induces branching of the leading processes of migrating neurons and that of basal processes of RGCs when they arrive at the Reelin-containing marginal zone. Branching of these processes may be crucial for the termination of nuclear translocation during the migratory process and for correct neuronal positioning.

The spatial and temporal arrangement of the radial glial scaffold suggests a role in axon tract formation in the developing spinal cord

Radial glial cells serve diverse roles during the development of the central nervous system (CNS). In the embryonic brain, they are recognised as guidance conduits for migrating neuroblasts and as multipotent stem cells, generating both neurons and glia. While their stem cell capacities in the developing spinal cord are as yet not fully clarified, they are classically seen as a population of astrocytes precursors, before gradually disappearing as the spinal cord matures. Although the origins and lineages of CNS radial glial cells are being more clearly understood, the relationships between radial glial cells and growing white matter (WM) tracts are largely unknown. Here, we provide an in-depth description of the distribution and organisation of radial glial cell processes during the peak periods of axonogenesis in the rat spinal cord. We show that radial glial cell distribution is highly ordered in the WM from E14 to E18, when the initial patterning of axon tracts is taking place. We report that the density of radial glial cell processes is tightly conserved throughout development in the dorsal, lateral and ventral WM funiculi along the rostrocaudal axis of the spinal cord. We provide evidence that from E16 the dorsal funiculi grow within and are segregated by fascicles of processes emanating from the dorsomedial septum. The density of radial glial cells declines with the maturation of axon tracts and coincides with the onset of the radial glial cell-astrocyte transformation. As such, we propose that radial glial cells act as structural scaffolds by compartmentalising and supporting WM patterning in the spinal cord during embryonic development.

The origin of cortical neurons.

Neurons of the mammalian cerebral cortex comprise two broad classes: pyramidal neurons, which project to distant targets, and the inhibitory nonpyramidal cells, the cortical interneurons. Pyramidal neurons are generated in the germinal ventricular zone, which lines the lateral ventricles, and migrate along the processes of radial glial cells to their positions in the developing cortex in an 'inside-out' sequence. The GABA-containing nonpyramidal cells originate for the most part in the ganglionic eminence, the primordium of the basal ganglia in the ventral telencephalon. These cells follow tangential migratory routes to enter the cortex and are in close association with the corticofugal axonal system. Once they enter the cortex, they move towards the ventricular zone, possibly to obtain positional information, before they migrate radially in the direction of the pial surface to take up their positions in the developing cortex. The mechanisms that guide interneurons throughout these long and complex migratory routes are currently under investigation.

Modes of neuronal migration in the developing cerebral cortex.

The conventional scheme of cortical formation shows that postmitotic neurons migrate away from the germinal ventricular zone to their positions in the developing cortex, guided by the processes of radial glial cells. However, recent studies indicate that different neuronal types adopt distinct modes of migration in the developing cortex. Here, we review evidence for two modes of radial movement: somal translocation, which is adopted by the early-generated neurons; and glia-guided locomotion, which is used predominantly by pyramidal cells. Cortical interneurons, which originate in the ventral telencephalon, use a third mode of migration. They migrate tangentially into the cortex, then seek the ventricular zone before moving radially to take up their positions in the cortical anlage.

Glial‐defined rhombomere boundaries in developing Xenopus hindbrain

The vertebrate central nervous system is characterized by regional specialization, which arises during early development and contributes to patterning the emerging central nervous system (CNS). In the hindbrain, rhombomeres demarcate nonoverlapping regions of the CNS that give rise to distinct neural structures. The cellular structures that define boundaries between adjacent rhombomeres are as yet unclear. However, in certain species the boundary regions between discrete CNS regions appear to be defined by specialized glial cells. Here, we show that in developing Xenopus, DMγ, a membrane protein of the proteolipid protein family, is expressed in a subset of radial glia. During development, DMγ transcripts are first expressed in presumptive glial cells throughout the hindbrain, but later become confined to the ventricular zone at rhombomere centers, whereas the protein is exclusively expressed in radial glial cell processes that occupy the rhombomere boundary regions. Likewise, early in development vimentin and glial fibrillary acidic protein are extensively coexpressed in hindbrain radial glia but subsequently define distinct rhombomere domains: vimentin remains localized in radial glia at the rhombomere boundary regions, whereas expression of glial fibrillary acidic protein becomes restricted to the centers. Moreover, radial glial processes at the boundary region are distinguishable from those at the center region; the processes of the boundary region radial glia extend upward in a fan-shaped arrangement and are encircled by the processes from the center glia. These data suggest that an early event in determining rhombomere topology is the specification of both morphologically and biochemically distinct subsets of radial glia. J. Comp. Neurol. 424:47–57, 2000. © 2000 Wiley-Liss, Inc.

The contribution of the ganglionic eminence to the neuronal cell types of the cerebral cortex.

The principal neuronal types of the mammalian cerebral cortex are the excitatory pyramidal cells and the inhibitory interneurons, the non-pyramidal cells. It is thought that these neurons arise in the ventricular zone surrounding the telencephalic ventricles. From there, newly generated neurons migrate outward along the processes of radial glial cells to reach the cortical plate where they accumulate in an 'inside-out' sequence to form the six-layered structure of the neocortex. Here we review emerging evidence that pyramidal neurons are generated in the cortical ventricular zone, whereas the majority of the non-pyramidal cells arise in the ganglionic eminences of the ventral telencephalon. These neurons follow tangential migratory routes to reach their positions in the developing cortex.

Glial-defined rhombomere boundaries in developingXenopus hindbrain

The vertebrate central nervous system is characterized by regional specialization, which arises during early development and contributes to patterning the emerging central nervous system (CNS). In the hindbrain, rhombomeres demarcate nonoverlapping regions of the CNS that give rise to distinct neural structures. The cellular structures that define boundaries between adjacent rhombomeres are as yet unclear. However, in certain species the boundary regions between discrete CNS regions appear to be defined by specialized glial cells. Here, we show that in developing Xenopus, DM gamma, a membrane protein of the proteolipid protein family, is expressed in a subset of radial glia. During development, DM gamma transcripts are first expressed in presumptive glial cells throughout the hindbrain, but later become confined to the ventricular zone at rhombomere centers, whereas the protein is exclusively expressed in radial glial cell processes that occupy the rhombomere boundary regions. Likewise, early in development vimentin and glial fibrillary acidic protein are extensively coexpressed in hindbrain radial glia but subsequently define distinct rhombomere domains: vimentin remains localized in radial glia at the rhombomere boundary regions, whereas expression of glial fibrillary acidic protein becomes restricted to the centers. Moreover, radial glial processes at the boundary region are distinguishable from those at the center region; the processes of the boundary region radial glia extend upward in a fan-shaped arrangement and are encircled by the processes from the center glia. These data suggest that an early event in determining rhombomere topology is the specification of both morphologically and biochemically distinct subsets of radial glia.

Developmental expression, pattern of distribution, and effect on cell aggregation implicate a neuron‐glial junctional domain protein in neuronal migration

We developed a panel of monoclonal antibodies to cerebellar astroglial cells and selected for study those that revealed microdomain structures on the cell surface of neocortical and cerebellar astrocytes. One antibody, 15D7-AD7, recognized the ∼72 kDa polypeptide doublet that was identified previously by the polyclonal antibody D4 as a component of the microdomain structure formed between migrating neurons and radial glial cell processes (Cameron and Rakic [1994] J. Neurosci. 14:3139–3155). Immunofluorescent localization studies reveal a spatial and temporal pattern of 15D7 immunoreactivity in multiple brain regions that correlates well with time periods when neuronal cell migration is a prominent morphogenetic event. In areas where the process of migration is underway, 15D7 immunoreactivity is detected simultaneously in both radial glial cells and cells that have the positional and morphologic features characteristic of migrating neurons. Subsequent to the completion of migration, immunoreactivity is detected in the transitional forms of radial glial cells and mature astrocytes, but not in neurons. Cell aggregation analyses reveal that 15D7 antibodies perturb the rate of aggregation for astrocyte-astrocyte, neuron-neuron, and mixed cell-cell combinations. Taken together, the present studies suggest that the polypeptides recognized by the 15D7 antibodies likely participate in an adhesive process, principally within the ventricular and subventricular zones, that is essential at the onset of the cell migration process. J. Comp. Neurol. 387:467–488, 1997. © 1997 Wiley-Liss, Inc.

Developmental expression, pattern of distribution, and effect on cell aggregation implicate a neuron-glial junctional domain protein in neuronal migration

We developed a panel of monoclonal antibodies to cerebellar astroglial cells and selected for study those that revealed microdomain structures on the cell surface of neocortical and cerebellar astrocytes. One antibody, 15D7-AD7, recognized the approximately 72 kDa polypeptide doublet that was identified previously by the polyclonal antibody D4 as a component of the microdomain structure formed between migrating neurons and radial glial cell processes (Cameron and Rakic [1994] J. Neurosci. 14:3139-3155). Immunofluorescent localization studies reveal a spatial and temporal pattern of 15D7 immunoreactivity in multiple brain regions that correlates well with time periods when neuronal cell migration is a prominent morphogenetic event. In areas where the process of migration is underway, 15D7 immunoreactivity is detected simultaneously in both radial glial cells and cells that have the positional and morphologic features characteristic of migrating neurons. Subsequent to the completion of migration, immunoreactivity is detected in the transitional forms of radial glial cells and mature astrocytes, but not in neurons. Cell aggregation analyses reveal that 15D7 antibodies perturb the rate of aggregation for astrocyte-astrocyte, neuron-neuron, and mixed cell-cell combinations. Taken together, the present studies suggest that the polypeptides recognized by the 15D7 antibodies likely participate in an adhesive process, principally within the ventricular and subventricular zones, that is essential at the onset of the cell migration process.

Reelin mRNA expression during mouse brain development.

Using in situ hybridization, expression of the mRNA for reelin, the gene most probably responsible for the reeler trait, was studied during mouse brain development, from embryonic day 13 to maturity. The highest level of expression was found in Cajal-Retzius neurons, while a high signal was also seen in the olfactory bulb, the external granular layer of the cerebellum and, particularly at early developmental stages, in hypothalamic differentiation fields, tectum and spinal cord. A moderate to low level of expression was found in the septal area, striatal fields, habenular nuclei, some thalamic nuclei, particularly the lateral geniculate, the retina and some nuclei of the reticular formation in the central field of the medulla. Paradoxically, no reelin expression was detected in radial glial cells, the cortical plate, Purkinje cells, inferior olivary neurons and many other areas that are characteristically abnormal in reeler mutant mice. Together with other preliminary studies, the present observations suggest that the action of reelin is indirect, possibly mediated by the extracellular matrix. Most of the data can be explained by supposing that reelin is a cell-repulsive molecule which prevents migrating neurons from invading reelin-rich areas, and thus facilitates the deployment of radial glial cell processes and the formation of early architectonic patterns.

Role of neuron-glial junctional domain proteins in the maintenance and termination of neuronal migration across the embryonic cerebral wall

To identify glial membrane proteins that contribute to the process of neuronal migration in the developing brain, we developed a polyclonal antiserum (D4) and a monoclonal antibody (NJPA1: neuron-glial junctional polypeptide antibody) that recognize membrane proteins localized to the plasmalemmal junction between migrating neurons and adjacent radial glial fibers (Cameron and Rakic, 1994). Here, we show that in the developing cerebral cortex, immunoreactivity for these junctional polypeptides is present throughout the neuronal migratory pathway but becomes minimal or absent where radial glial cell processes enter the marginal zone region, the barrier at which newly arrived neurons normally stop their migration and detach from their glial fiber substrate. We thus tested, using imprints of embryonic cerebral wall and slice preparations, whether the junctional membrane proteins detected by our antibodies contribute to the regulation of neuronal migration in the cerebral cortex. The rate of neuronal migration on glial cell substrates was reduced significantly in the presence of D4 or NJPA1 antibodies. Antibody exposure typically led to the withdrawal of leading processes, changes in microtubular organization and, in some instances, to detachment of neurons from their glial cell substrates. These results suggest that the polypeptides recognized by the D4 and NJPA1 antibodies are essential for the maintenance of normal neuronal migration. Dismantling of neuron-glial cell junctional domains formed by these membrane proteins may underlie neuronal cell detachment from glial migratory substrates at the interface between cortical plate and marginal zone in the developing cerebral wall.

Migration of dopaminergic neurons in the embryonic mesencephalon of mice

Migration of dopamine (DA)-containing neurons and its guiding cues were histologically examined in the embryonic mesencephalon of normal mice. Cells immunoreactive (ir) for tyrosine hydroxylase (TH), a DA-synthesizing enzyme, were first detected on embryonic day 10 (E10) in the medio-basal part of the mesencephalon and were distributed throughout the entire length of the ventral mesencephalic wall at E12. By E14, TH-ir cells were located laterally along the ventral pial surface to form the primordia of the substantia nigra. Experiments with a single injection of bromodeoxyuridine, a thymidine analog, demonstrated that cells generated in the ventricular surface of the ventral mesencephalon at E11 migrated ventrally and then moved laterally to form the substantia nigra and the ventral tegmental area. Electron microscopic examination of the ventral mesencephalon of E12 mice disclosed that in the dorsal part ventrally migrating immature neurons made close contacts with the processes of radial glial cells. The expression of tenascin was transiently seen on radial glial processes between E10 and E13 coincident with the period of the ventral migration of mesencephalic DA neurons. By double immunostaining of E13 mesencephalon, ventrally migrating TH-ir cells were seen to be apposed to tenascin-bearing radial glial processes. On the other hand, laterally migrating neurons in the basal part of the mesencephalon were observed by electron microscopy to contact with tangentially arranged nerve fibers which were immunopositive for the 160 kDa neurofilament polypeptide at the light microscopic level from E10. Double immunostaining of E13 mesencephalon demonstrated that laterally migrating TH-ir cells were intermingled among neurofilament-ir fiber bundles. The cells of origin of the tangential nerve fibers were detected in the lateral part of the mesencephalon, when a fluorescent dye, 1,1′-dioctadecyl-3,3,3′,3′-tetramethyl-indocarbocyanine perchlorate (DiI) was injected into the basal part of the mesencephalon of fixed E12 mice. The present results suggest that guiding cues of the radial migration of mesencephalic DA neurons represent processes of radial glial cells which express tenascin. On the other hand, tangentially arranged nerve fibers originating from the lateral part of the mesencephalon may provide a scaffolding along which the mesencephalic DA neurons subsequently migrate laterally to form the ventral tegmental area and the substantia nigra.

Identification of membrane proteins that comprise the plasmalemmal junction between migrating neurons and radial glial cells

We have initiated studies to identify membrane polypeptides of radial glial cells that contribute to the selective cell-cell recognition and migration events in developing brain. Of several polyclonal antisera evaluated, one (D4), developed against formaldehyde fixed type 1 cerebellar glial cells, immunolabels the free surface of cortical and cerebellar astroglial and radial glial cells in a patchy pattern. In dissociated glial-neuronal cell cocultures, microdomains of immunoreactivity are detected at the site where the somal region of cells with the morphology of migrating neurons is in contact with an elongated glial cell fiber. Microdomains are absent from oligodendrocytes, process-bearing astrocytes, and neurons. The surface microdomains do not colocalize with components that compose focal adhesion plaques--integrin subunits, vinculin, or actin--and their integrity appears to require an intact microtubule rather than actin cytoskeleton. Furthermore, microdomain structure is maintained in the absence of extracellular Ca and Mg ions. Immunoblot analyses using antibodies affinity purified to individual proteins indicate that the microdomains are composed of two antigens with apparent molecular mass of approximately 48 kDa and approximately 72 kDa. The 48 kDa antigen is not observed in non-neural epithelial tissues and is detected in cortical and cerebellar tissues only at a developmental period that coincides with the stage of active neuronal cell migration. In contrast, the 72 kDa antigen is expressed in many neural and non-neural tissues at late developmental and adult stages. Our data suggest that the identified membrane proteins may contribute to the formation of the junction between migrating neurons and radial glial cell processes and that this junctional complex is linked to the microtubule cytoskeleton.