Mercedes F. Paredes

Intrinsically determined cell death of developing cortical interneurons

Cortical inhibitory circuits are formed by γ-aminobutyric acid (GABA)-secreting interneurons, a cell population that originates far from the cerebral cortex in the embryonic ventral forebrain. Given their distant developmental origins, it is intriguing how the number of cortical interneurons is ultimately determined. One possibility, suggested by the neurotrophic hypothesis, is that cortical interneurons are overproduced, and then after their migration into cortex the excess interneurons are eliminated through a competition for extrinsically derived trophic signals. Here we characterize the developmental cell death of mouse cortical interneurons in vivo, in vitro and after transplantation. We found that 40% of developing cortical interneurons were eliminated through Bax (Bcl-2-associated X)-dependent apoptosis during postnatal life. When cultured in vitro or transplanted into the cortex, interneuron precursors died at a cellular age similar to that at which endogenous interneurons died during normal development. Over transplant sizes that varied 200-fold, a constant fraction of the transplanted population underwent cell death. The death of transplanted neurons was not affected by the cell-autonomous disruption of TrkB (tropomyosin kinase receptor B), the main neurotrophin receptor expressed by neurons of the central nervous system. Transplantation expanded the cortical interneuron population by up to 35%, but the frequency of inhibitory synaptic events did not scale with the number of transplanted interneurons. Taken together, our findings indicate that interneuron cell death is determined intrinsically, either cell-autonomously or through a population-autonomous competition for survival signals derived from other interneurons.

Stromal-Derived Factor-1 (CXCL12) Regulates Laminar Position of Cajal-Retzius Cells in Normal and Dysplastic Brains

Normal brain development requires a series of highly complex and interrelated steps. This process presents many opportunities for errors to occur, which could result in developmental defects in the brain, clinically referred to as malformations of cortical development. The marginal zone and Cajal-Retzius cells are key players in cortical development and are established early, yet there is little understanding of the factors resulting in the disruption of the marginal zone in many types of cortical malformation syndromes. We showed previously that treatment with methylazoxymethanol in rats causes marginal zone dysplasia with displacement of Cajal-Retzius cells to deeper cortical layers. Here we establish that loss of activity of the chemokine stromal-derived factor-1 (SDF1) (CXCL12), which is expressed by the leptomeninges, is necessary and sufficient to cause marginal zone disorganization in this widely used teratogenic animal model. We also found that mice with mutations in the main receptor for SDF1 (CXCR4) have Cajal-Retzius cells displaced to deeper cortical layers. Furthermore, by inhibiting SDF1 signaling in utero by intraventricular injection of a receptor antagonist, we establish that SDF1 signaling is required for the maintenance of Cajal-Retzius cell position in the marginal zone during normal cortical development. Our data imply that cortical layering is not a static process, but rather requires input from locally produced molecular cues for maintenance, and that complex syndromes of cortical malformation as a result of environmental insults may still be amenable to explanation by interruption of specific molecular signaling pathways.

Human hippocampal neurogenesis drops sharply in children to undetectable levels in adults

New neurons continue to be generated in the subgranular zone of the dentate gyrus of the adult mammalian hippocampus. This process has been linked to learning and memory, stress and exercise, and is thought to be altered in neurological disease. In humans, some studies have suggested that hundreds of new neurons are added to the adult dentate gyrus every day, whereas other studies find many fewer putative new neurons. Despite these discrepancies, it is generally believed that the adult human hippocampus continues to generate new neurons. Here we show that a defined population of progenitor cells does not coalesce in the subgranular zone during human fetal or postnatal development. We also find that the number of proliferating progenitors and young neurons in the dentate gyrus declines sharply during the first year of life and only a few isolated young neurons are observed by 7 and 13 years of age. In adult patients with epilepsy and healthy adults (18–77 years; n = 17 post-mortem samples from controls; n = 12 surgical resection samples from patients with epilepsy), young neurons were not detected in the dentate gyrus. In the monkey (Macaca mulatta) hippocampus, proliferation of neurons in the subgranular zone was found in early postnatal life, but this diminished during juvenile development as neurogenesis decreased. We conclude that recruitment of young neurons to the primate hippocampus decreases rapidly during the first years of life, and that neurogenesis in the dentate gyrus does not continue, or is extremely rare, in adult humans. The early decline in hippocampal neurogenesis raises questions about how the function of the dentate gyrus differs between humans and other species in which adult hippocampal neurogenesis is preserved.

Extensive migration of young neurons into the infant human frontal lobe

Building the human brain As the brain develops, neurons migrate from zones of proliferation to their final locations, where they begin to build circuits. Paredes et al. have discovered that shortly after birth, a group of neurons that proliferates near the ventricles migrates in chains alongside circulatory vessels into the frontal lobes (see the Perspective by McKenzie and Fishell). Young neurons that migrate postnatally into the anterior cingulate cortex then develop features of inhibitory interneurons. The number of migratory cells decreases over the first 7 months of life, and by 2 years of age, migratory cells are not evident. Any damage during migration, such as hypoxia, may affect the childs subsequent physical and behavioral development. Science, this issue p. 81; see also p. 38 Neurons are still finding their places as inhibitory circuits are established in the developing postnatal brain. [Also see Perspective by McKenzie] INTRODUCTION Inhibitory interneurons balance the excitation and inhibition of neural networks and therefore are key to normal brain function. In the developing brain, young interneurons migrate from their sites of birth into distant locations, where they functionally integrate. Although this neuronal migration is largely complete before birth, some young inhibitory interneurons continue to travel and add to circuits in restricted regions of the juvenile and adult mammalian brain. For example, postnatally migrating inhibitory neurons travel from the walls of the lateral ventricle, along the rostral migratory stream (RMS) into the olfactory bulb. In humans, an additional ventral route branching off the RMS, the medial migratory stream (MMS), takes young neurons into the medial prefrontal cortex. It has been suggested that recruitment of neurons during postnatal life could help shape neural circuits according to experience. Specifically, inhibitory interneuron maturation during postnatal development is associated with critical periods of brain plasticity. We asked whether neuronal recruitment continues into early childhood in the frontal lobe, a region of the human brain that has greatly increased in size and complexity during evolution. RATIONALE Migrating young neurons persist for several months after birth in an extensive region of the subventricular zone (SVZ) around the anterior lateral ventricles in the human brain. Are all these young neurons migrating into the RMS and MMS, or do they have other destinations? Using high-resolution magnetic resonance imaging (MRI), histology, and time-lapse confocal microscopy, we observed the migration of many young inhibitory interneurons around the dorsal anterior walls of the lateral ventricle and into multiple cortical regions of the human frontal cortex. We determined the location and orientation of these young neurons, demonstrated their active translocation, and inferred their fates in the postnatal anterior forebrain. RESULTS A large collection of cells expressing doublecortin (DCX), a marker of young migrating neurons, traveled and integrated within the infant frontal lobe. This migratory stream, which was most prominent during the first 2 months after birth and persisted until at least 5 months, formed a caplike structure surrounding the anterior body of the lateral ventricle. We refer to this population of young neurons as the Arc. This structure could also be visualized by brain MRI. Young neurons in the Arc appeared to move long distances in distinct regions around the ventricular wall and the developing white matter. The orientation of elongated DCX+ cells suggested that migratory neurons closer to the ventricular wall dispersed tangentially. In contrast, migratory neurons within the developing white matter tended to be orientated toward the overlying cortex. These cells expressed markers of interneurons, and their entry into the anterior cingulate cortex (a major target of the Arc used for quantification) was correlated with the emergence of specific subtypes of γ-aminobutyric acid (GABA)–expressing interneurons (neuropeptide Y, somatostatin, calretinin, and calbindin). Expression of transcription factors associated with specific sites of origin suggested that these neurons arise from ventral telencephalon progenitor domains. CONCLUSION Widespread neuronal migration into the human frontal lobe continues for several months after birth. Young neurons express markers of cortical inhibitory interneurons and originate outside the cortex, likely in the ventral forebrain. The postnatal recruitment of large populations of inhibitory neurons may contribute to maturation and plasticity in the human frontal cortex. Defects in the migration of these neurons could result in circuit dysfunction associated with neurodevelopmental disorders. Widespread neuronal migration into the human frontal lobe continues during postnatal life. (A) Sagittal schematic of the newborn forebrain shows prominent collections of young migratory neurons (illustrated in green) adjacent to the lateral ventricle (LV) and in the overlying white matter. Directional axes: D, dorsal; A, anterior. (B and C) DCX+ cells coexpress GABA and GAD67, markers of inhibitory interneurons (marked by arrows). The first few months after birth, when a child begins to interact with the environment, are critical to human brain development. The human frontal lobe is important for social behavior and executive function; it has increased in size and complexity relative to other species, but the processes that have contributed to this expansion are unknown. Our studies of postmortem infant human brains revealed a collection of neurons that migrate and integrate widely into the frontal lobe during infancy. Chains of young neurons move tangentially close to the walls of the lateral ventricles and along blood vessels. These cells then individually disperse long distances to reach cortical tissue, where they differentiate and contribute to inhibitory circuits. Late-arriving interneurons could contribute to developmental plasticity, and the disruption of their postnatal migration or differentiation may underlie neurodevelopmental disorders.

Expression of SDF-1 and CXCR4 during Reorganization of the Postnatal Dentate Gyrus

Previous studies have demonstrated that stromal cell-derived factor 1 (SDF-1) is crucial for early dentate development; however, the mouse mutants for this chemokine and its only receptor, CXCR4, are neonatally lethal, making conclusions about the role of these molecules in postnatal development difficult to sustain. Previous expression analyses have used single labeling, but the distribution of CXCR4 is complex and to determine the cell types expressing CXCR4 requires multiple marker labeling. In this study, we examined the distribution of SDF-1 and CXCR4 mRNAs during the first postnatal weeks, combining these markers with several other cell-type-specific markers. We found that SDF-1 has three sites of expression: (1) continuation of prenatal expression in the meninges; (2) expression in Cajal-Retzius cells occupying the molecular layer of the upper and lower blades of the dentate, and (3) the maturing dentate granule neurons themselves. The timing of expression in these three sites corresponds to alterations in the distribution of the primary cell types expressing CXCR4 during the same periods, notably the expression of CXCR4 in radial-glial-like GFAP-expressing dentate precursors and immature dentate granule neurons. Taken together, our data suggest potential ongoing roles for SDF-1/CXCR4 signaling in the dentate gyrus during the early postnatal period that will be tested in the future with more precise genetic approaches.

Brain size and limits to adult neurogenesis.

The walls of the cerebral ventricles in the developing embryo harbor the primary neural stem cells from which most neurons and glia derive. In many vertebrates, neurogenesis continues postnatally and into adulthood in this region. Adult neurogenesis at the ventricle has been most extensively studied in organisms with small brains, such as reptiles, birds, and rodents. In reptiles and birds, these progenitor cells give rise to young neurons that migrate into many regions of the forebrain. Neurogenesis in adult rodents is also relatively widespread along the lateral ventricles, but migration is largely restricted to the rostral migratory stream into the olfactory bulb. Recent work indicates that the wall of the lateral ventricle is highly regionalized, with progenitor cells giving rise to different types of neurons depending on their location. In species with larger brains, young neurons born in these spatially specified domains become dramatically separated from potential final destinations. Here we hypothesize that the increase in size and topographical complexity (e.g., intervening white matter tracts) in larger brains may severely limit the long‐term contribution of new neurons born close to, or in, the ventricular wall. We compare the process of adult neuronal birth, migration, and integration across species with different brain sizes, and discuss how early regional specification of progenitor cells may interact with brain size and affect where and when new neurons are added. J. Comp. Neurol. 524:646–664, 2016.

Neuropeptide Y modulates a G protein-coupled inwardly rectifying potassium current in the mouse hippocampus

Neuropeptide Y (NPY) is an abundant brain peptide with endogenous antiepileptic activity. Here we examined the role played by Y1 receptors (Y1R) in the mouse hippocampus. Using whole-cell patch-clamp recordings, we show that hilar neurons in acute mouse hippocampal slices exhibit a G-protein coupled inwardly rectifying potassium (GIRK) current that is significantly enhanced during exogenous NPY application. NPY-mediated enhancement of GIRK current was observed on 47% of putative interneurons and was mimicked by application of Y1R specific agonist (Leu(31)Pro(34) NPY). Immunostaining revealed the presence of Y1R on cell somas of hilar NPY-containing interneurons. Thus, our results suggest that Y1R on hilar interneurons may act as a peptide autoreceptor.

Distinct and separable roles for EZH2 in neurogenic astroglia

The epigenetic mechanisms that enable specialized astrocytes to retain neurogenic competence throughout adult life are still poorly understood. Here we show that astrocytes that serve as neural stem cells (NSCs) in the adult mouse subventricular zone (SVZ) express the histone methyltransferase EZH2. This Polycomb repressive factor is required for neurogenesis independent of its role in SVZ NSC proliferation, as Ink4a/Arf-deficiency in Ezh2-deleted SVZ NSCs rescues cell proliferation, but neurogenesis remains defective. Olig2 is a direct target of EZH2, and repression of this bHLH transcription factor is critical for neuronal differentiation. Furthermore, Ezh2 prevents the inappropriate activation of genes associated with non-SVZ neuronal subtypes. In the human brain, SVZ cells including local astroglia also express EZH2, correlating with postnatal neurogenesis. Thus, EZH2 is an epigenetic regulator that distinguishes neurogenic SVZ astrocytes, orchestrating distinct and separable aspects of adult stem cell biology, which has important implications for regenerative medicine and oncogenesis. DOI: http://dx.doi.org/10.7554/eLife.02439.001

Embryonic and Early Postnatal Abnormalities Contributing to the Development of Hippocampal Malformations in a Rodent Model of Dysplasia

While there are many recent examples of single gene deletions that lead to defects in cortical development, most human cases of cortical disorganization can be attributed to a combination of environmental and genetic factors. Elucidating the cellular or developmental basis of teratogenic exposures in experimental animals is an important approach to understanding how environmental insults at particular developmental junctures can lead to complex brain malformations. Rats with prenatal exposure to methylazoxymethanol (MAM) reproduce many anatomical features seen in epilepsy patients. Previous studies have shown that heterotopic clusters of neocortically derived neurons exhibit hyperexcitable firing activity and may be a source of heightened seizure susceptibility; however, the events that lead to the formation of these abnormal cell clusters is unclear. Here we used a panel of molecular markers and birthdating studies to show that in MAM‐exposed rats the abnormal cell clusters (heterotopia) first appear postnatally in the hippocampus (P1–2) and that their appearance is preceded by a distinct sequence of perturbations in neocortical development: 1) disruption of the radial glial scaffolding with premature astroglial differentiation, and 2) thickening of the marginal zone with redistribution of Cajal‐Retzius neurons to deeper layers. These initial events are followed by disruption of the cortical plate and appearance of subventricular zone nodules. Finally, we observed the erosion of neocortical subventricular zone nodules into the hippocampus around parturition followed by migration of nodules to hippocampus. We conclude that prenatal MAM exposure disrupts critical developmental processes and prenatal neocortical structures, ultimately resulting in neocortical disorganization and hippocampal malformations. J. Comp. Neurol. 495:133–148, 2006.

Hilar Mossy Cells Share Developmental Influences with Dentate Granule Neurons

Mossy cells are the major class of excitatory neurons in the dentate hilus. Although mossy cells are involved in a range of physiological and pathological conditions, very little is known about their ontogeny. To gain insight into this issue, we first determined the developmental stage at which mossy cells can be reliably identified with the molecular markers calretinin and GluR2/3 and found that hilar mossy cells were first identifiable around the end of the 1st postnatal week. Birthdating studies combined with staining for these markers revealed that the appearance of mossy cells coincided with the first wave of dentate granule cell production during mid-gestation. Since mossy cells are born as the first granule cells are produced and it is believed that mossy cells originate from the neuroepithelium adjacent to the dentate progenitor zone, we examined to what extent the development of mossy cells is controlled by the same molecular pathways as that of granule cells. To do this, we analyzed the production of mossy cells in Lef1 and NeuroD mutant animals, in which granule cell production is disrupted during precursor proliferation or neuronal differentiation, respectively. The production of mossy cells was almost entirely lost in both mutants. Collectively, these data suggests that hilar mossy cells, unlike CA subfield pyramidal cells, are influenced by many of the same developmental cues as dentate granule cells.