The GTPase Rab21 is required for neuronal development and migration in the cerebral cortex.

Development of the mammalian neocortex requires proper inside-out migration of developing cortical neurons from the germinal ventricular zone toward the cortical plate. The mechanics of this migration requires precise coordination of different cellular phenomena including cytoskeleton dynamics, membrane trafficking, and cell adhesion. The small GTPases play a central role in all these events. The small GTPase Rab21 regulates migration and neurite growth in developing neurons. Moreover, regulators and effectors of Rab21 have been implicated in brain pathologies with cortical malformations, suggesting a key function for the Rab21 signaling pathway in cortical development. Mechanistically, it has been posited that Rab21 influences cell migration by controlling the trafficking of endocytic vesicles containing adhesion molecules. However, direct evidence of the participation of Rab21 or its mechanism of action in the regulation of cortical migration is still incomplete. In this study, we demonstrate that Rab21 plays a critical role in the differentiation and migration of pyramidal neurons by regulating the levels of the amyloid precursor protein on the neuronal cell surface. Rab21 loss of function increased the levels of membrane-exposed APP, resulting in impaired cortical neuronal differentiation and migration. These findings further our understanding of the processes governing the development of the cerebral cortex and shed light onto the molecular mechanisms behind cortical development disorders derived from the malfunctioning of Rab21 signaling effectors.

Ethanol Gestational Exposure Impairs Vascular Development and Endothelial Potential to Control BBB-Associated Astrocyte Function in the Developing Cerebral Cortex.

Ethanol consumption during pregnancy or lactation period can induce permanent damage to the development of the central nervous system (CNS), resulting in fetal alcohol spectrum disorders (FASD). CNS development depends on proper neural cells and blood vessel (BV) development and blood-brain barrier (BBB) establishment; however, little is known about how ethanol affects these events. Here, we investigated the impact of ethanol exposure to endothelial cells (ECs) function and to ECs interaction with astrocytes in the context of BBB establishment. Cerebral cortex of newborn mice exposed in utero to ethanol (FASD model) presented a hypervascularized phenotype, revealed by augmented vessel density, length, and branch points. Further, aberrant distribution of the tight junction ZO-1 protein along BVs and increased rates of perivascular astrocytic endfeet around BVs were observed. In vitro exposure of human brain microcapillary ECs (HBMEC) to ethanol significantly disrupted ZO-1 distribution, decreased Claudin-5 and GLUT-1 expression and impaired glucose uptake, and increased nitric oxide secretion. These events were accompanied by upregulation of angiogenesis-related secreted proteins by ECs in response to ethanol exposure. Treatment of cortical astrocytes with conditioned medium (CM) from ethanol exposed ECs, upregulated astrocyte's expression of GFAP, Cx43, and Lipocalin-2 genes, as well as the pro-inflammatory genes, IL-1beta, IL-6, and TNF-alpha, which was accompanied by NF-kappa B protein nuclear accumulation. Our findings suggest that ethanol triggers a dysfunctional phenotype in brain ECs, leading to impairment of cortical vascular network formation, and promotes ECs-induced astrocyte dysfunction, which could dramatically affect BBB establishment in the developing brain.

Long-Term Functional and Cytoarchitectonic Effects of the Systemic Administration of the Histamine H1 Receptor Antagonist/Inverse Agonist Chlorpheniramine During Gestation in the Rat Offspring Primary Motor Cortex.

The transient histaminergic system is among the first neurotransmitter systems to appear during brain development in the rat mesencephalon/rhombencephalon. Histamine increases FOXP2-positive deep-layer neuron differentiation of cortical neural stem cells through H1 receptor activation in vitro. The in utero or systemic administration of chlorpheniramine (H1 receptor antagonist/inverse agonist) during deep-layer cortical neurogenesis decreases FOXP2 neurons in the developing cortex, and H1R- or histidine decarboxylase-knockout mice show impairment in learning and memory, wakefulness and nociception, functions modulated by the cerebral cortex. Due to the role of H1R in cortical neural stem cell neurogenesis, the purpose of this study was to evaluate the postnatal impact of the systemic administration of chlorpheniramine during deep-layer cortical neuron differentiation (E12-14) in the primary motor cortex (M1) of neonates (P0) and 21-day-old pups (P21). Chlorpheniramine or vehicle were systemically administered (5 mg/kg, i.p.) to pregnant Wistar rats at gestational days 12-14, and the expression and distribution of deep- (FOXP2 and TBR1) and superficial-layer (SATB2) neuronal cortical markers were analyzed in neonates from both groups. The qRT-PCR analysis revealed a reduction in the expression of Satb2 and FoxP2. However, Western blot and immunofluorescence showed increased protein levels in the chlorpheniramine-treated group. In P21 pups, the three markers showed impaired distribution and increased immunofluorescence in the experimental group. The Sholl analysis evidenced altered dendritic arborization of deep-layer neurons, with lower excitability in response to histamine, as evaluated by whole-cell patch-clamp recording, as well as diminished depolarization-evoked [3H]-glutamate release from striatal slices. Overall, these results suggest long-lasting effects of blocking H1Rs during early neurogenesis that may impact the pathways involved in voluntary motor activity and cognition.

Impaired Cortical Cytoarchitecture and Reduced Excitability of Deep-Layer Neurons in the Offspring of Diabetic Rats.

Maternal diabetes has been related to low verbal task scores, impaired fine and gross motor skills, and poor performance in graphic and visuospatial tasks during childhood. The primary motor cortex is important for controlling motor functions, and embryos exposed to high glucose show changes in cell proliferation, migration, and differentiation during corticogenesis. However, the existing studies do not discriminate between embryos with or without neural tube defects, making it difficult to conclude whether the reported changes are related to neural tube defects or other anomalies. Furthermore, postnatal effects on central nervous system cytoarchitecture and function have been scarcely addressed. Through molecular, biochemical, morphological, and electrophysiological approaches, we provide evidence of impaired primary motor cerebral cortex lamination and neuronal function in pups from diabetic rats, showing an altered distribution of SATB2, FOXP2, and TBR1, impaired cell migration and polarity, and decreased excitability of deep-layer cortical neurons, suggesting abnormalities in cortico-cortical and extra-cortical innervation. Furthermore, phase-plot analysis of action potentials suggests changes in the activity of potassium channels. These results indicate that high-glucose insult during development promotes complex changes in migration, neurogenesis, cell polarity establishment, and dendritic arborization, which in turn lead to reduced excitability of deep-layer cortical neurons.

Radial Glia Cells Control Angiogenesis in the Developing Cerebral Cortex Through TGF-ß1 Signaling.

Neuroangiogenesis in the developing central nervous system is controlled by interactions between endothelial cells (ECs) and radial glia (RG) neural stem cells, although RG-derived molecules implicated in these events are not fully known. Here, we investigated the role of RG-secreted TGF-ß1, in angiogenesis in the developing cerebral cortex. By isolation of murine microcapillary brain endothelial cells (MBECs), we demonstrate that conditioned medium from RG cultures (RG-CM) promoted MBEC migration and formation of vessel-like structures in vitro, in a TGF-ß1-dependent manner. These events were followed by endothelial regulation of GPR124 and BAI-1 gene expression by RG-CM. Proteome profile of RG-CM identified angiogenesis-related molecules IGFBP2/3, osteopontin, endostatin, SDF1, fractalkine, TIMP1/4, Ang-1, pentraxin3, and Cyr61, some of them modulated by TGF-ß1 induction. In vivo gain and loss of function assays targeting RG cells demonstrates a specific TGF-ß1-dependent control of blood vessels branching in the cerebral cortex. Together, our results point to TGF-ß1 signaling pathway as a potential mediator of the RG-EC interactions and shed light to the key role of RG in paving the brain vascular network.

Sonic hedgehog signaling regulates mode of cell division of early cerebral cortex progenitors and increases astrogliogenesis.

The morphogen Sonic Hedgehog (SHH) plays a critical role in the development of different tissues. In the central nervous system, SHH is well known to contribute to the patterning of the spinal cord and separation of the brain hemispheres. In addition, it has recently been shown that SHH signaling also contributes to the patterning of the telencephalon and establishment of adult neurogenic niches. In this work, we investigated whether SHH signaling influences the behavior of neural progenitors isolated from the dorsal telencephalon, which generate excitatory neurons and macroglial cells in vitro. We observed that SHH increases proliferation of cortical progenitors and generation of astrocytes, whereas blocking SHH signaling with cyclopamine has opposite effects. In both cases, generation of neurons did not seem to be affected. However, cell survival was broadly affected by blockade of SHH signaling. SHH effects were related to three different cell phenomena: mode of cell division, cell cycle length and cell growth. Together, our data in vitro demonstrate that SHH signaling controls cell behaviors that are important for proliferation of cerebral cortex progenitors, as well as differentiation and survival of neurons and astroglial cells.

Maternal-fetal unit interactions and eutherian neocortical development and evolution.

The conserved brain design that primates inherited from early mammals differs from the variable adult brain size and species-specific brain dominances observed across mammals. This variability relies on the emergence of specialized cerebral cortical regions and sub-compartments, triggering an increase in brain size, areal interconnectivity and histological complexity that ultimately lies on the activation of developmental programs. Structural placental features are not well correlated with brain enlargement; however, several endocrine pathways could be tuned with the activation of neuronal progenitors in the proliferative neocortical compartments. In this article, we reviewed some mechanisms of eutherians maternal-fetal unit interactions associated with brain development and evolution. We propose a hypothesis of brain evolution where proliferative compartments in primates become activated by "non-classical" endocrine placental signals participating in different steps of corticogenesis. Changes in the inner placental structure, along with placenta endocrine stimuli over the cortical proliferative activity would allow mammalian brain enlargement with a concomitant shorter gestation span, as an evolutionary strategy to escape from parent-offspring conflict.

Efectos de la exposición prenatal a paraquat sobre el desarrollo de la transmisión sináptica aminoacídica en la corteza cerebral parietal del ratón / Effects of prenatal expossure to paraquat on the development of amino acid synaptic transmission in mouse cerebral parietal cortex

Se estudiaron los efectos de la exposición prenatal al paraquat (PQ), sobre el desarrollo postnatal de la transmisión sináptica aminoacídica de la corteza cerebral parietal del ratón. Las ratonas NMRI preñadas del grupo experimental recibieron 5 dosis de 10mg/kg de peso corporal de PQ, entre el día de gestación (G)12 y G20, y el grupo control recibió solución salina. Mediante HPLC, se determinaron los niveles de aspartato, glutamato, glicina, GABA y taurina de las crías, entre la edad postnatal (P)1 y P30. Entre P3 y P15, se observó un incremento significativo de los neurotransmisores excitatorios, aspartato y glutamato, en los ratones expuestos a PQ. Con respecto a la neurotransmisión inhibidora, los cambios más importantes se observaron en glicina: sus niveles se mantuvieron significativamente por debajo del grupo control entre P1 y P7, y significativamente por encima en P11 y P15. Para taurina, entre P1 y P7 sus niveles se mantuvieron significativamente altos con respecto al grupo control. En P30, los niveles de todos los neurotransmisores se encontraron significativamente por debajo del grupo control. En conclusión, podríamos decir que la exposición prenatal a PQ tuvo efectos tóxicos que se reflejaron en una alteración de los niveles basales de los neurotransmisores aminoacídicos durante el desarrollo postnatal de la corteza parietal del ratón, predominando la excitación sobre la inhibición durante todo el período estudiado. Estas alteraciones podrían indicar la ocurrencia de importantes daños corticales, tales como la disminución de algunas poblaciones neuronales, la inadecuada formación de los circuitos corticales y alteraciones en el proceso de sinaptogénesis.

The effects of prenatal expossure to paraquat (PQ) were studied on postnatal development of mouse parietal cerebral cortex, in particular, the ontogenesis of amino acid synaptic transmission. Pregnant NMRI mice were separated into two groups: the experimental group received 5 doses of 10mg PQ/kg body weight, between days of gestation (G)12 and G20, whereas the control group received physiological saline solution. Levels of neurotransmitter amino acids: Asp, Glu, Gly, GABA and Tau were determined by HPLC between postnatal (P) days P1 and P30. Between P3 and P15, a significant increment in the levels of excitatory amino acids, Asp and Glu, were observed in mice exposed to PQ, as compared with the control group. With respect to the inhibitory neurotransmitter levels, in the group exposed to PQ, the more important changes were observed in Gly between P1 and P15. In relation to taurine, its levels remained significantly higher between P1 and P7 with respect to the control group. It is important to emphasize that at P30, the levels of all neurotransmitters in the experimental group were significantly lower than those of control. In conclusion, prenatal exposure to PQ caused neurotoxicity in the developing mouse parietal cortex, as shown by the alterations in the basal levels of amino acid neurotransmitters, with the excitatory predominating over inhibitory neurotransmission, throughout the studied developmental period. These alterations could indicate the occurrence of important cortical injuries, such as decrement in some neuronal populations, inadequate formation of intrinsic cortical circuits and alterations in synaptogenic processes.