John G. Parnavelas

The Medial Ganglionic Eminence Gives Rise to a Population of Early Neurons in the Developing Cerebral Cortex

During development of the neocortex, the marginal zone (layer I) and the subplate (layer VII) are the first layers to form from a primordial plexiform neoropil. The cortical plate (layers II–VI) is subsequently established between these superficial and deep components of the primordial plexiform neuropil. Neurons in the early zones are thought to play important roles in the formation of the cortex: the Cajal-Retzius cells of the marginal zone are instrumental in neuronal migration and laminar formation, and cells of the subplate are involved in the formation of cortical connections. Using the fluorescent tracer 1,1′-dioctodecyl-3,3,3′,3′-tetramethylindocarbocyanine (DiI), we have shown here that a substantial proportion of neurons of the marginal zone, including cells with features of Cajal-Retzius cells, and of the subplate and lower intermediate zone are not born in the ventricular neuroepithelium but instead originate in the medial ganglionic eminence (MGE), the pallidal primordium. These neurons follow a tangential migratory route to their positions in the developing cortex. They express the neurotransmitter GABA but seem to lack the calcium binding protein calretinin; some migrating cells found in the marginal zone express reelin. In addition, migrating cells express the LIM-homeobox gene Lhx6, a characteristic marker of the MGE. It is suggested that this gene uniquely or in combination with other transcription factors may be involved in the decision of MGE cells to differentiate in situ or migrate to the neocortex.

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.

The formation and maturation of synapses in the visual cortex of the rat. II. Quantitative analysis

SummarySynapse formation and maturation were examined in the visual cortex of albino rats from birth to maturity. During the first few days of postnatal life, synapses were sparsely scattered in the subplate zone and in layer I. They appeared immature as judged by the irregular shapes of the presynaptic and postsynaptic profiles, the relatively poorly defined membrane specializations and the presence of only a few synaptic vesicles in the presynaptic structures. As the neuropil matured, synapses were observed throughout the cortex, showing increased thickening of the membrane specializations and more vesicles. However, it was not until the end of the fourth postnatal week that they appeared qualitatively indistinguishable from synapses identified in the adult material.A feature characteristic of the developing visual cortex was the presence of vacant membrane specializations that resembled type I postsynaptic densities. These specializations, which were located either opposite extracellular space or opposite another neuronal process, were only evident during the initial stages of synaptogenesis and their frequency decreased as the number of synapses increased. In addition, transitional forms between these densities and true type I synapses were identified during the first two postnatal weeks. Structures that resembled vacant postsynaptic densities typical of type II synapses were not observed. The earliest identified forms of type II synaptic contacts identified consisted of two profiles that exhibited symmetrical membrane specializations and cleft material. Based on these observations, a scheme has been proposed for the formation of type I and type II synapses in the visual cortex of the rat.

Apoptosis and Its Relation to the Cell Cycle in the Developing Cerebral Cortex

Large numbers of dying cells are found in proliferating tissues, suggesting a link between cell death and cell division. We detected and quantified dying cells during pre- and early postnatal development of the rat cerebral cortex using in situ end labeling of DNA fragmentation [terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL)] and electron microscopy. The proliferative zones that give rise to the neuronal and glial cell types of the cortex, the ventricular and, to a larger extent, the subventricular zones showed higher incidence of cell death than other regions of the developing cortex during the period of neurogenesis. Gel electrophoresis of DNA isolated from the subventricular zone of newborn animals showed a ladder pattern that is characteristic of apoptosis. The number of apoptotic cells remained high in this zone for at least 2 weeks, during which period cells continued to divide. The correlation between cell division and cell death was studied in the subventricular zone of newborn rats; cumulative labeling with bromodeoxyuridine showed that 71% of TUNEL-labeled cells had taken up this S-phase marker before undergoing cell death. Using bromodeoxyuridine and [3H]-thymidine in succession to identify a cohort of proliferating cells, we found that the clearance time of TUNEL-positive nuclei was 2 hr and 20 min. A comparison between the number of mitotic figures and that of TUNEL-positive nuclei showed that cell death affects one in every 14 cells produced by dividing ventricular zone cells at embryonic day 16 and one in every 1.5 cells produced in the subventricular zone of newborn rats. In addition, we found that most of TUNEL-positive cells were in the G1 phase of their cell cycle. We conclude that apoptosis is prominent in the proliferating neuroepithelium of the developing rat cerebral cortex and that it is related to the progression of the cell cycle.

Area identity shifts in the early cerebral cortex of Emx2 −/− mutant mice

The specification of area identities in the cerebral cortex is a complex process, primed by intrinsic cortical cues and refined after the arrival of afferent fibers from the thalamus. Little is known about the genetic control of the early steps of this process, but the distinctive expression pattern of the homeogene Emx2 in the developing cortex has prompted suggestions that it is critical in this context. We tested this hypothesis using Emx2−/− mice. We found that the normal spectrum of cortical areal identities was encoded in these mutants, but areas with caudal–medial identities were reduced and those with anterior–lateral identities were relatively expanded in the cortex.

The morphology and distribution of peptide-containing neurons in the adult and developing visual cortex of the rat. I. Somatostatin

SummaryUsing conventional immunocytochemical techniques, we have examined the morphology and distribution of somatostatin-like immunoreactive neurons in the visual cortex of albino rats between the first postnatal day and maturity. In the adult, somatostatin-immunoreactive neurons were observed in layers II to VI but were concentrated in layers II and III. These cells displayed morphological features characteristic of the multipolar and bitufted varieties of cortical non-pyramidal neurons as described in Golgi preparations of rat visual cortex.On the first postnatal day and in the subsequent few days, immunoreactivity was confined to immature bipolar and multipolar neurons concentrated in layers V and VI. Labelled cells first appeared in the more superficial layers at the beginning of the second postnatal week and attained a distribution similar to that observed in adult animals at the end of this week. At this time they closely resembled their adult counterparts from which they appeared indistinguishable by the end of the third postnatal week. The late appearance of labelled cells in the superficial layers, where they are predominantly located in adult animals, suggests that the somatostatin immunoreactivity exhibited by most of these neurons develops several days after they have completed their migration and assumed their positions in the visual cortex.

Robo1 regulates the development of major axon tracts and interneuron migration in the forebrain

The Slit genes encode secreted ligands that regulate axon branching, commissural axon pathfinding and neuronal migration. The principal identified receptor for Slit is Robo (Roundabout in Drosophila). To investigate Slit signalling in forebrain development, we generated Robo1 knockout mice by targeted deletion of exon 5 of the Robo1 gene. Homozygote knockout mice died at birth, but prenatally displayed major defects in axon pathfinding and cortical interneuron migration. Axon pathfinding defects included dysgenesis of the corpus callosum and hippocampal commissure, and abnormalities in corticothalamic and thalamocortical targeting. Slit2 and Slit1/2 double mutants display malformations in callosal development, and in corticothalamic and thalamocortical targeting, as well as optic tract defects. In these animals, corticothalamic axons form large fasciculated bundles that aberrantly cross the midline at the level of the hippocampal and anterior commissures, and more caudally at the medial preoptic area. Such phenotypes of corticothalamic targeting were not observed in Robo1 knockout mice but, instead, both corticothalamic and thalamocortical axons aberrantly arrived at their respective targets at least 1 day earlier than controls. By contrast, in Slit mutants, fewer thalamic axons actually arrive in the cortex during development. Finally, significantly more interneurons (up to twice as many at E12.5 and E15.5) migrated into the cortex of Robo1 knockout mice, particularly in both rostral and parietal regions, but not caudal cortex. These results indicate that Robo1 mutants have distinct phenotypes, some of which are different from those described in Slit mutants, suggesting that additional ligands, receptors or receptor partners are likely to be involved in Slit/Robo signalling.

Ventricle-directed migration in the developing cerebral cortex

It is believed that postmitotic neurons migrate away from their sites of origin in the germinal zones to populate distant targets. Contrary to this notion, we found, using time-lapse imaging of brain slices, populations of neurons positioned at various levels of the developing neocortex that migrate towards the cortical ventricular zone. After a pause in this proliferative zone, they migrate radially in the direction of the pial surface to take up positions in the cortical plate. Immunohistochemical analysis together with tracer labeling in brain slices showed that cells showing ventricle-directed migration in the developing cortex are GABAergic interneurons originating in the ganglionic eminence in the ventral telencephalon. We speculate that combinations of chemoattractant and chemorepellent molecules are involved in this ventricle-directed migration and that interneurons may seek the cortical ventricular zone to receive layer information.

Cell and molecular mechanisms involved in the migration of cortical interneurons

Since the discovery that the vast majority of the GABA‐containing interneurons of the cerebral cortex arise in the subpallium, considerable effort has been put into the description of the precise origin of these neurons in subdivisions of the ganglionic eminence and in the migratory routes they follow on their way to the developing cortex. More recently, studies have focused on the molecular and cellular mechanisms that guide their migration. Investigations of the molecular mechanisms involved have demonstrated important roles for numerous transcription factors, motogenic factors and guidance molecules. Here, we review results of very recent analyses of the underlying cellular mechanisms and specifically of the movement of the nucleus, cytoplasmic components and neuritic processes during interneuron migration.

Gap junctions in the adult cerebral cortex: Regional differences in their distribution and cellular expression of connexins

Gap junctions are membrane channels that mediate electrical and metabolic coupling between adjacent cells. Immunocytochemical analysis by using a panel of anti‐connexin antibodies, as well as electron microscopy of thin sections and freeze‐fracture replicas, has shown that gap junctions and their constituent proteins are abundant in the cerebral cortex of the adult rat. Their frequency and distribution vary in different cortical regions, which may reflect differences in the cellular and functional organization of these areas of the cortex. Gap junctions were identified between glial cells and, less frequently, between neuronal elements. Heterologous junctions were also identified between astrocytes and oligodendrocytes and between neurons and glia; the latter category included abundant junctions between astrocytic processes and neurons. Double‐antibody labelling experiments in tissue sections and in acutely dissociated cells showed that connexin 32 was expressed in neurons and oligodendrocytes, whereas connexin 43, widely believed to be expressed only in astrocytes, was also localized in a population of cortical neurons. These results show that gap junctions can provide a major nonsynaptic means of communication between cortical cell types.