Glypican-4 is an FGF2-binding heparan sulfate proteoglycan expressed in neural precursor cells.

FGF2 is a crucial mitogen for neural precursor cells in the developing cerebral cortex. Heparan sulfate proteoglycans (HSPGs) are thought to play a role in cortical neurogenesis by regulating the action of FGF2 on neural precursor cells. In this article, we present data indicating that glypican-4 (K-glypican), a GPI-anchored cell surface HSPG, is involved in these processes. In the developing mouse brain, glypican-4 mRNA is expressed predominantly in the ventricular zone of the telencephalon. Neither the outer layers of the telencephalic wall nor the ventricular zone of other parts of the developing brain express significant levels of glypican-4, with the exception of the ventricular zone of the tectum. In cultures of E13 rat cortical precursor cells, glypican-4 is expressed in cells immunoreactive for nestin and the D1.1 antigen, markers of neural precursor cells. Glypican-4 expression was not detected in early postmitotic or fully differentiated neurons. Recombinant glypican-4 produced in immortalized neural precursor cells binds FGF2 through its heparan sulfate chains and suppressed the mitogenic effect of FGF2 on E13 cortical precursor cells. The spatiotemporal expression pattern of glypican-4 in the developing cerebral wall significantly overlaps with that of FGF2. These results suggest that glypican-4 plays a critical role in the regulation of FGF2 action during cortical neurogenesis.     

Histogenesis of the inferior colliculus in rat

Summary The early histogenesis of the inferior colliculus from embryonic day 11 to 18 (E11 to E18) has been studied in the rat by analysis of Golgi impregnated material and plastic sections.This analysis has shown that the pseudo-stratified columnar neuroepithelium observed at E11 is followed by the appearance at E12 of three zones: marginal, intermediate and ventricular. Signs of cell differentiation are first observed in the intermediate zone. Secondary rearrangements occur within this zone, and by E16 a thin cortical plate (the cortex of the inferior colliculus) develops at the junction of the intermediate and marginal zones giving rise to the external and pericentral nuclei of this structure, which has a cortical organization in adults. The remainder of the intermediate zone (the nucleus of the inferior colliculus), invaded by axons, expands dramatically by E16–E17 and gives rise to the central and dorso-medial nuclei of the inferior colliculus which have a nuclear organization in adults. The morphogenetic events which take place in these two regions differ and can be identified by the study of cell migration and differentiation.In the nucleus of the inferior colliculus, neuronal migration begins with detachment of the ventricularly directed process, or trailing process, of the primitive epithelial cell from the ventricular surface. This is followed by the ascent of the cell nucleus through the pially directed, or leading, process by a mechanism identical to the perikaryal translocation already described in other regions of the nervous system. This mechanism of cell migration is characteristic of a first type of migratory young neuron (type I). Axons initiate from the leading process of these cells during migration and dendrites grow out in various directions giving these cells a bipolar or a multipolar appearance. Dendritic differentiation occurs first in the outermost cells of the nucleus and proceeds inwards.In the cortex of the inferior colliculus, neuronal migration also begins with detachment of the ventricular process, which occurs by E12, immediately followed by the detachment and retraction of the apical or leading process. Within the intermediate zone, migratory cells become rounded and sprout numerous processes. One of these processes is tipped by a growth cone and displays all the characteristics of an axon. It is directed tangentially in the intermediate layer. Dendritic growth and differentiation starts when the cells reach their final position in the cortical plate, and proceeds from the innermost cells outward. Due to the inadequacy of our methods for identifying radial glial fibers, the mechanism of migration of this type of cell (type II) remains unclear.Our results confirm that the inferior colliculus of the rat is organized as a central nuclear mass surrounded by a thin cortex. As previously observed in other regions of the nervous system, the modes of cell migration and differentiation in the cortical and non-cortical structures of the inferior colliculus appear different.     

Broca's area in the human brain is involved in the selection of grammatical gender for language production: evidence from event-related functional magnetic resonance imaging

We investigated cortical functions of two Unverricht-Lundborg disease (ULD) patients suffering from myoclonic jerks, but no generalized tonic-clonic seizures. Somatosensory cortical responses were recorded to median nerve stimuli and coherence was calculated between cortical and muscle signals during isometric contraction of hand muscle. In contrast to ULD patients with generalized tonic-clonic seizures, responses of the primary somatosensory (SI) cortex were only slightly enhanced in the left and normal in the right hemisphere, and no early responses were observed in the ipsilateral SI. Cortex-muscle coherence was remarkably enhanced. We conclude that in ULD patients without generalized tonic-clonic seizures, both the excitability of the SI and transcallosal conduction are relatively normal, probably decreasing susceptibility to generalized seizures. Disturbed cortical control of muscle contraction indicates selective alteration of the motor cortex activation.     

Expression of the transcription factor Zif268 in the visual cortex of monocularly deprived rats: effects of nerve growth factor.

Neurotrophins are known to be involved in experience-dependent plasticity of the visual cortex. Here, we have characterized in detail the effects of intraventricular nerve growth factor infusion in monocularly deprived rats by using immunostaining for the immediate-early gene product Zif268 as a marker of functional activity with cellular resolution. We have taken advantage of the rapid regulation of Zif268 by visual input to reveal the cortical units that are responsive to the deprived eye after a period of monocular deprivation. We found that responses to the deprived eye were significantly preserved in the cortex of monocularly deprived rats infused with nerve growth factor. The effects of nerve growth factor were greater for cortical cells located in deep layers and with more peripheral receptive fields. Results from Zif268 staining correlated very well with those obtained by single-cell recordings from the visual cortex. Our results demonstrate that exogenous nerve growth factor preserves the functional input from the deprived eye, enabling cortical neurons to activate immediate-early gene expression in response to stimulation of the deprived eye. Furthermore, we show that the intraventricular infusion of nerve growth factor differentially affects the ocular dominance of cells at various depths and eccentricities in the developing cortex.     

Mitotic spindle fiber orientation in relation to cell migration in the neo-cortex of normal and reeler mouse.

The angular distribution of mitotic spindle fibers has been studied in the telencephalic ventricular zone (VZ) of mouse embryos and has been found to be monophasic and invariant during the early cortical histogenesis both in the normal and in a reeler embryo. This observation suggests that the orientation of mitotic spindle fibers is independent of nerve cell migration in the embryonic cortex.     

Immunoelectron microscopic study of tyrosine hydroxylase immunoreactive nerve fibers and ganglion cells in the rat adrenal gland.

The present immunocytochemical study used an antiserum to tyrosine hydroxylase (TH), the first enzyme in the biosynthetic pathway of catecholamines, and revealed TH immunoreactivity in the ganglion cells and in the varicose nerve fibers of the cortex and medulla in the rat adrenal gland. TH immunoreactive nerve fibers in the cortex and medulla contained large and small granular vesicles, and also small clear vesicles. The immunoreactive nerve fibers were in close apposition to cortical cells in the cortex and in apposition to smooth muscle cells of blood vessels in both the cortex and medulla. Furthermore, TH immunoreactive nerve fibers were sometimes in close apposition to pericytes of blood vessels in the cortex and chromaffin cells in the medulla. The present results suggest that the catecholaminergic nerve fibers in the rat adrenal gland may be both intrinsic and extrinsic in origin.     

Immunoelectron microscopic study of tyrosine hydroxylase immunoreactive nerve fibers and ganglion cells in the rat adrenal gland

The present immunocytochemical study used an antiserum to tyrosine hydroxylase (TH), the first enzyme in the biosynthetic pathway of catecholamines, and revealed TH immunoreactivity in the ganglion cells and in the varicose nerve fibers of the cortex and medulla in the rat adrenal gland. TH immunoreactive nerve fibers in the cortex and medulla contained large and small granular vesicles, and also small clear vesicles. The immunoreactive nerve fibers were in close apposition to cortical cells in the cortex and in apposition to smooth muscle cells of blood vessels in both the cortex and medulla. Furthermore, TH immunoreactive nerve fibers were sometimes in close apposition to pericytes of blood vessels in the cortex and chromaffin cells in the medulla. The present results suggest that the catecholaminergic nerve fibers in the rat adrenal gland may be both intrinsic and extrinsic in origin.     

The release of γ-aminobutyric acid during inhibition in the cat visual cortex

1. The release of gamma-aminobutyric acid (GABA) from the surface of the posterior lateral gyrus of the cerebral cortex was measured by a sensitive enzymic fluorimetric assay procedure. Experiments were performed with anaesthetized cats during resting conditions and during cortical inhibition produced by electrical stimulation of the brain surface or of the lateral geniculate nucleus (l.g.n.).2. The average resting release of endogenous GABA was 0.20 n-mole/ 7 min.cm(2) cortex; this was increased during stimulation of both the cortical surface (2.9 times resting release during monopolar stimulation and 7.4 times resting release during bipolar stimulation) and the l.g.n. (5.7 times resting release).3. Removal of calcium ions from the collection fluid did not affect the resting release of endogenous GABA but prevented the increase in GABA release normally evoked by stimulation of the cortical surface.4. The stimulus parameters used to increase the release of GABA also inhibited the glutamate-induced firing of single cells in the visual cortex and this inhibition was abolished in the absence of calcium ions.5. In three experiments the total amino acid content of cortical samples was examined using an amino acid analyser. With the exception of GABA, there were no significant differences between the rates of release of any other detected amino acids during periods with and without electrical stimulation of the cortex.6. It is suggested that since the release of GABA observed during inhibitory stimulation of the cortex is calcium-dependent and specific, it may originate from inhibitory nerve terminals in the cortex. The present findings support the view that GABA is a central inhibitory neurotransmitter.     

Age changes in bovine lens endopeptidase activity

Lens endopeptidase activity and thermal stability have been determined as a function of cell development, cell age, and animal age. Lenses from animals aged 3 months to 15 years (lens weights 1.15-2.80 g) were divided into epithelial (outermost), cortical (peripheral), and nuclear (central) regions. Changes accompanying cell development were determined by measuring specific activity in epithelial (undifferentiated), outer cortical (differentiating), inner cortical (mature) and nuclear (aged) regions of individual lenses. Thermal stability of the enzyme activity obtained from the outer cortical and nuclear regions of the same lenses was also determined. Specific activity and thermal stability were found to decrease as a function of lens cell development. Changes with cell development represent the effects of both differentiation and increasing cell age. To determine the effects of cell age alone, activity was determined in the same population of aged, fully differentiated cells in lenses of different ages. Specific activity decreased as a function of cell age alone. Changes with animal age were determined by comparing cells of the same developmental stage from animals of different ages (e.g., differentiating cells of the cortex in animals 3 months to 15 years old). Specific activity for the cortical region increased with animal age while specific activity in the nuclear region appeared to remain constant or decrease slightly with increasing animal age. Thermal stability of the enzyme activity from the cortex was different in young and adult lenses. The change in stability occurred early in the lifespan and was therefore more closely related to animal development than to aging.     

Prenatal development of neural excitation in rat thalamocortical projections studied by optical recording

To elucidate the formation of early thalamocortical synapses we recorded optical images with voltage-sensitive dyes from the cerebral cortex of prenatal rats by selective thalamic stimulation of thalamocortical slice preparations. At embryonic day (E) 17, thalamic stimulation elicited excitation that rapidly propagated through the internal capsule to the cortex. These responses lasted less than 15 ms, and were not affected by the application of glutamate receptor antagonists, suggesting that they might reflect presynaptic fiber responses. At E18, long-lasting (more than 300 ms) responses appeared in the internal capsule and in subplate. By E19, long-lasting responses increased in the cortical subplate. By E21, shortly before birth, the deep cortical layers were also activated in addition to the subplate. These long-lasting responses seen in the internal capsule and subplate were blocked by the antagonist perfusion, but the first spike-like responses still remained. The laminar location of the responses was confirmed in the same slices by Nissl staining and subplate cells were labeled by birthdating with bromodeoxyuridine at E13. Our results demonstrate that there is a few days delay between the arrival of thalamocortical axons at the subplate at E16 and the appearance of functional thalamocortical synaptic transmission at E19. Since thalamocortical connections are already functional within the subplate and in the deep cortical plate at embryonic ages, prenatal thalamocortical synaptic connections could influence cortical circuit formation before birth.     

Expression of highly polysialylated NCAM in the neocortex and piriform cortex of the developing and the adult rat

Summary The expression of a highly polysialylated form of the neural cell adhesion molecule (NCAM-H) has been investigated in the neocortex and piriform cortex of the developing and the adult rat by using a monoclonal antibody 12E3, which has been found to recognize the polysialic acid portion of NCAM-H. Immunoblot analysis of the cortical homogenates showed that NCAM-H was temporarily expressed during the late embryonic and early postnatal stages. Further, immunohistochemical observations revealed that NCAM-H appeared at embryonic day 13 (E13) in the plexiform primordium in horizontally-oriented cells, probably Cajal-Retzius cells, which are the first neurons to differentiate. During the late embryonic stage, the marginal zone, subplate, and intermediate zone strongly stained, whereas the ventricular zone stained weakly. After birth, the NCAM-H expression was progressively attenuated from a week onwards, and almost vanished in the adult neocortex. In the primordium of the piriform cortex, NCAM-H immunoreactivity also became positive at E13. The time sequences of the NCAM-H expression in these neurons were similar to those of the neurons in the neocortical area. In the piriform cortex, however, the expression remained in a number of neurons in the layer II, which receives a large number of olfactory fibers from the olfactory bulb, where prolonged neurogenesis and construction of neural circuits take place in adulthood. These results suggest that NCAM-H not only plays an important role in the developing rat cortex, but also may be involved in some functions related to reorganization in the adult piriform cortex.     

Comparative analysis of extra-ventricular mitoses at early stages of cortical development in rat and human

Embryonic germinal zones of the dorsal and ventral telencephalon generate cortical neurons during the final week of gestation in rodent and during several months in human. Whereas the vast majority of cortical interneurons originate from the ventral telencephalon, excitatory neurons are locally generated within the germinal zone of the dorsal telencephalon, the future cerebral cortex, itself. However, a number of studies have described proliferating cells external to the ventricular and subventricular germinal zones in the developing dorsal telencephalon. In this study, we performed a comprehensive cell density analysis of such ‘extra-ventricular proliferating cells’ (EVPCs) during corticogenesis in rat and human using a mitotic marker anti-phospho-histone H3. Subsequently, we performed double-labelling studies with other mitotic and cell type specific markers to undertake phenotypic characterisation of EVPCs. Our findings show: (1) the densities of extra-ventricular H3-positive (H3⁺) cells were surprisingly similar in preplate stage rat and human; (2) extra-ventricular proliferation continues during mid-and late corticogenesis in rat and in early fetal human cortex; and (3) extra-ventricular cells appear to be mitotic precursors as they are not immunoreactive for a panel of early post-mitotic and cell type-specific markers, although (4) a subset of EVPCs are proliferating microglia. These data suggest that some aspects of early corticogenesis are conserved between rodent and human despite marked differences in the duration of neurogenesis and the anatomical organisation of the developing cerebral cortex. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

The early development of thalamocortical and corticothalamic projections in the mouse

The initial ingrowth of corticothalamic and thalamocortical projections was examined in mice at embryonic and perinatal stages. Fibers, in fixed brains, were labeled with the carbocyanine dye 1,1’-dioctadecyl-3,3,3’,3’-tetramethylindocarbocianine perchlorate (DiI). By E13, the corticofugal fibers had entered the lowest intermediate zone through which they ran, turned over the corpus striatum, and left the cortex. The fibers were arranged in scattered bundles throughout the corpus striatum. At E14 corticofugal axons reached the internal capsule and at E14.5–E15 they established contact within the thalamus. Meanwhile, the thalamocortical afferents reached the neocortex at E13. At this time fibers ran tangentially within the intermediate zone, immediately underneath the cortical plate. By E14, the fibers had started to invade the subplate and, by E15, thalamocortical fibers had begun their radial growth into the cortex. Such radial growth proceeded steadily, invading each cortical layer as it differentiated cytoarchitectonically from the dense cortical plate. The first retrogradely labeled cells were detected at the cortical plate at E15. By the day of birth (E20), thalamocortical fibers had formed a dense branching system within layers VI and V. Our observations indicate that, in mice, the thalamic axons reach the cortex before corticothalamic projections enter the thalamic nuclei. Moreover, the results suggest that the pathway followed by each fiber system is different. By DiI injections into the internal capsule we have also determined that subplate cells are the first to send axons to the thalamus. Accepted: 11 August 1999     

An early development defect in the cerebral cortex of the reeler mouse

Summary Brains of reeler and normal mouse embryos have been studied on semi-thin sections and with Golgi impregnations. No change can be seen in the neuroepithelium or in the primary cortical organization. The first evidence of a morphological abnormality appears at E 14, in the cortical plate. Instead of being closely packed and radially oriented, nerve cells are loosely arranged and show quite variable orientations of their long axis and apical dendrite. The axons run obliquely through the cortical plate and do not display the characteristic angular course seen in the normal animal.It is suggested that the primary defect in reeler mice may be in the plasma membrane of cortical plate cells, resulting in a loss of their capacity for mutual recognition and binding. This could account for the cytoarchitectonic disorganization in this mutant, especially the absence of a molecular layer and the inversion of the histogenetic gradient in the developing cerebral cortex.Aspirant au Fonds National de la Recherche Scientifique de Belgique     

Development of astrocytes in the mouse embryonic cerebrum tracked by tenascin-C gene expression

The development of astrocytes in the rodent cerebrum has been considered to occur mainly in the perinatal stage after the end of neurogenesis, and the mode of generation and migration of the astroglial precursors in the embryonic cerebrum remains poorly understood. Tenascin-C (TN-C) is an astroglia-derived extracellular matrix molecule, and its expression can be traced to the ventricular germinative zone of the embryonic cerebrum. TN-C expression may therefore be used as a marker of astroglial precursors prior to the expression of the glial fibrillary acidic protein (GFAP), a marker for the late stage of astroglial differentiation. The development of astrocytes in the mouse embryonic cerebrum was examined by in situ hybridization histochemistry for TN-C mRNA as an early marker. Observations showed that TN-C-positive cells were aligned in the ventricular germinative zone and began to migrate radially at E (embryonic day) 15. They were dispersed into the cortex as early as E17, and some were still in the DNA synthetic phase (S phase) even after lodgement in the cortex. TN-C-positive cells differentiated into GFAP-immunopositive astrocytes in the cortical and subcortical areas at the neonatal stage. The distribution of their active generation sites in the ventricular germinative zone was not homogeneous but displayed a restricted pattern. The mode of their migration towards the cortical and subcortical areas also showed a characteristic pattern that correlated to the radial glial arrangement. These findings indicate that TN-C-positive cells represent a new subset of astroglial precursors which arise directly from the ventricular germinative zone, and that astrogliogenesis from this precursor occurs as early as the period of neurogenesis at the embryonic stage.     

Effects of changes in cortical arousal and of auditory cortex cooling on neuronal activity in the medial geniculate body

Summary The activity of cells in the medial geniculate body (MGB) of adult cats was recorded during different states of cortical arousal with and without cooling of the auditory cortex. In the absence of auditory cortex cooling, the overall mean unit spontaneous discharge rate was 49% higher during desynchronized Electrocorticogram (ECoG) periods (high cortical arousal) than during synchronized periods (low cortical arousal). Responses to sound were somewhat more prominent vis-à-vis the spontaneous activity during periods of high arousal. Changes in spontaneous discharge rate associated with arousal shifts were significantly reduced during auditory cortex cooling. When the ECoG changed from desynchronized to synchronized activity, MGB cells showed a change in discharge pattern, typically characterized by an increase in both high-rate bursts and long-interval pauses. These changes were duplicated for most cells by cooling of the auditory cortex. Corticofugal fiber discharge thus has an effect on MGB neuronal activity which is dependent on the level of cortical arousal. This effect is most likely a result of direct corticogeniculate activity, though indirect auditory cortex — brainstem — MGB routes may also be involved.Supported by a grant from the University of Illinois Campus Research Board     

TWO SIBLINGS, INCLUDING A FETUS, WITH TAY-SACHS DISEASE

This report consists of a morphological study on two siblings of Tay-Sachs disease. One is a girl of 3 years of age and the other is a 23-week fetus. In the first case, the nerve cells in the cerebral cortex showed histochemically abnormal accumulation of glycolipids and electron microscopically numerous membranous cytoplasmic bodies characteristic to gangliosidosis. In the fetal case, the nerve cells in the brain stem were well differentiated and here abnormal inclusions were found by histochemical study. The cortical neurons were immature in structure and inclusions were not demonstrated by histochemical study. In electron microscopic study, the cortical neurons also showed a small number of intracytoplasmic inclusions which contained electron dense granules and/or membranous structures. Causes of the differences in amount of abnormal lipid accumulation in the brain stem and the cerebral cortex were discussed.     

The early development of thalamocortical and corticothalamic projections in the mouse

The initial ingrowth of corticothalamic and thalamocortical projections was examined in mice at embryonic and perinatal stages. Fibers, in fixed brains, were labeled with the carbocyanine dye 1, 1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocianine perchlorate (DiI). By E13, the corticofugal fibers had entered the lowest intermediate zone through which they ran, turned over the corpus striatum, and left the cortex. The fibers were arranged in scattered bundles throughout the corpus striatum. At E14 corticofugal axons reached the internal capsule and at E14.5-E15 they established contact within the thalamus. Meanwhile, the thalamocortical afferents reached the neocortex at E13. At this time fibers ran tangentially within the intermediate zone, immediately underneath the cortical plate. By E14, the fibers had started to invade the subplate and, by E15, thalamocortical fibers had begun their radial growth into the cortex. Such radial growth proceeded steadily, invading each cortical layer as it differentiated cytoarchitectonically from the dense cortical plate. The first retrogradely labeled cells were detected at the cortical plate at E15. By the day of birth (E20), thalamocortical fibers had formed a dense branching system within layers VI and V. Our observations indicate that, in mice, the thalamic axons reach the cortex before corticothalamic projections enter the thalamic nuclei. Moreover, the results suggest that the pathway followed by each fiber system is different. By DiI injections into the internal capsule we have also determined that subplate cells are the first to send axons to the thalamus.