The temporal origin of dentate granule neurons dictates their role in spatial memory.

The dentate gyrus is one of the only brain regions that continues its development after birth in rodents. Adolescence is a very sensitive period during which cognitive competences are programmed. We investigated the role of dentate granule neurons (DGNs) born during adolescence in spatial memory and compared them with those generated earlier in life (in embryos or neonates) or during adulthood by combining functional imaging, retroviral and optogenetic tools to tag and silence DGNs. By imaging DGNs expressing Zif268, a proxy for neuronal activity, we found that neurons generated in adolescent rats (and not embryos or neonates) are transiently involved in spatial memory processing. In contrast, adult-generated DGNs are recruited at a later time point when animals are older. A causal relationship between the temporal origin of DGNs and spatial memory was confirmed by silencing DGNs in behaving animals. Our results demonstrate that the emergence of spatial memory depends on neurons born during adolescence, a function later assumed by neurons generated during adulthood.

Reelin Regulates the Maturation of Dendritic Spines, Synaptogenesis and Glial Ensheathment of Newborn Granule Cells.

Significance Statement: The extracellular protein Reelin has an important role in neurological diseases, including epilepsy, Alzheimer's disease and psychiatric diseases, targeting hippocampal circuits. Here we address the role of Reelin in the development of synaptic contacts in adult-generated granule cells (GCs), a neuronal population that is crucial for learning and memory and implicated in neurological and psychiatric diseases. We found that the Reelin pathway controls the shapes, sizes, and types of dendritic spines, the complexity of multisynaptic innervations and the degree of the perisynaptic astroglial ensheathment that controls synaptic homeostasis. These findings show a pivotal role of Reelin in GC synaptogenesis and provide a foundation for structural circuit alterations caused by Reelin deregulation that may occur in neurological and psychiatric disorders.

Cell-autonomous inactivation of the reelin pathway impairs adult neurogenesis in the hippocampus.

Adult hippocampal neurogenesis is thought to be essential for learning and memory, and has been implicated in the pathogenesis of several disorders. Although recent studies have identified key factors regulating neuroprogenitor proliferation in the adult hippocampus, the mechanisms that control the migration and integration of adult-born neurons into circuits are largely unknown. Reelin is an extracellular matrix protein that is vital for neuronal development. Activation of the Reelin cascade leads to phosphorylation of Disabled-1, an adaptor protein required for Reelin signaling. Here we used transgenic mouse and retroviral reporters along with Reelin signaling gain-of-function and loss-of-function studies to show that the Reelin pathway regulates migration and dendritic development of adult-generated hippocampal neurons. Whereas overexpression of Reelin accelerated dendritic maturation, inactivation of the Reelin signaling pathway specifically in adult neuroprogenitor cells resulted in aberrant migration, decreased dendrite development, formation of ectopic dendrites in the hilus, and the establishment of aberrant circuits. Our findings support a cell-autonomous and critical role for the Reelin pathway in regulating dendritic development and the integration of adult-generated granule cells and point to this pathway as a key regulator of adult neurogenesis. Moreover, our data reveal a novel role of the Reelin cascade in adult brain function with potential implications for the pathogenesis of several neurological and psychiatric disorders.

Adult-specific Reelin expression alters striatal neuronal organization: implications for neuropsychiatric disorders.

In addition to neuronal migration, brain development, and adult plasticity, the extracellular matrix protein Reelin has been extensively implicated in human psychiatric disorders such as schizophrenia, bipolar disorder, and autism spectrum disorder. Moreover, heterozygous reeler mice exhibit features reminiscent of these disorders, while overexpression of Reelin protects against its manifestation. However, how Reelin influences the structure and circuits of the striatal complex, a key region for the above-mentioned disorders, is far from being understood, especially when altered Reelin expression levels are found at adult stages. In the present study, we took advantage of complementary conditional gain- and loss-of-function mouse models to investigate how Reelin levels may modify adult brain striatal structure and neuronal composition. Using immunohistochemical techniques, we determined that Reelin does not seem to influence the striatal patch and matrix organization (studied by µ-opioid receptor immunohistochemistry) nor the density of medium spiny neurons (MSNs, studied with DARPP-32). We show that overexpression of Reelin leads to increased numbers of striatal parvalbumin- and cholinergic-interneurons, and to a slight increase in tyrosine hydroxylase-positive projections. We conclude that increased Reelin levels might modulate the numbers of striatal interneurons and the density of the nigrostriatal dopaminergic projections, suggesting that these changes may be involved in the protection of Reelin against neuropsychiatric disorders.

Chemogenetic stimulation of adult neurogenesis, and not neonatal neurogenesis, is sufficient to improve long-term memory accuracy.

Hippocampal adult neurogenesis is involved in many memory processes from learning, to remembering and forgetting. However, whether or not the stimulation of adult neurogenesis is a sufficient condition to improve memory performance remains unclear. Here, we developed and validated, using ex-vivo electrophysiology, a chemogenetic approach that combines selective tagging and activation of discrete adult-born neuron populations. Then we demonstrated that, in rats, this activation can improve accuracy and strength of remote memory. These results show that stimulation of adult-born neuron activity can counteract the natural fading of memory traces that occurs with the passage of time. This opens up new avenues for treating memory problems that may arise over time.

Adult-born neurons immature during learning are necessary for remote memory reconsolidation in rats.

Memory reconsolidation, the process by which memories are again stabilized after being reactivated, has strengthened the idea that memory stabilization is a highly plastic process. To date, the molecular and cellular bases of reconsolidation have been extensively investigated particularly within the hippocampus. However, the role of adult neurogenesis in memory reconsolidation is unclear. Here, we combined functional imaging, retroviral and chemogenetic approaches in rats to tag and manipulate different populations of rat adult-born neurons. We find that both mature and immature adult-born neurons are activated by remote memory retrieval. However, only specific silencing of the adult-born neurons immature during learning impairs remote memory retrieval-induced reconsolidation. Hence, our findings show that adult-born neurons immature during learning are required for the maintenance and update of remote memory reconsolidation.

Corrigendum: FIB/SEM technology and high-throughput 3D reconstruction of dendritic spines and synapses in GFP-labeled adult-generated neurons.

[This corrects the article on p. 60 in vol. 9, PMID: 26052271.].

FIB/SEM technology and high-throughput 3D reconstruction of dendritic spines and synapses in GFP-labeled adult-generated neurons.

The fine analysis of synaptic contacts is usually performed using transmission electron microscopy (TEM) and its combination with neuronal labeling techniques. However, the complex 3D architecture of neuronal samples calls for their reconstruction from serial sections. Here we show that focused ion beam/scanning electron microscopy (FIB/SEM) allows efficient, complete, and automatic 3D reconstruction of identified dendrites, including their spines and synapses, from GFP/DAB-labeled neurons, with a resolution comparable to that of TEM. We applied this technology to analyze the synaptogenesis of labeled adult-generated granule cells (GCs) in mice. 3D reconstruction of dendritic spines in GCs aged 3-4 and 8-9 weeks revealed two different stages of dendritic spine development and unexpected features of synapse formation, including vacant and branched dendritic spines and presynaptic terminals establishing synapses with up to 10 dendritic spines. Given the reliability, efficiency, and high resolution of FIB/SEM technology and the wide use of DAB in conventional EM, we consider FIB/SEM fundamental for the detailed characterization of identified synaptic contacts in neurons in a high-throughput manner.

Transient downregulation of Dab1 protein levels during development leads to behavioral and structural deficits: relevance for psychiatric disorders.

Psychiatric disorders have been hypothesized to originate during development, with genetic and environmental factors interacting in the etiology of disease. Therefore, developmentally regulated genes have received attention as risk modulators in psychiatric diseases. Reelin is an extracellular protein essential for neuronal migration and maturation during development, and its expression levels are reduced in psychiatric disorders. Interestingly, several perinatal insults that increase the risk of behavioral deficits alter Reelin signaling. However, it is not known whether a dysfunction in Reelin signaling during perinatal stages increases the risk of psychiatric disorders. Here we used a floxed dab1 allele to study whether a transient decrease in Dab1, a key component of the Reelin pathway, is sufficient to induce behavioral deficits related to psychiatric disorders. We found that transient Dab1 downregulation during perinatal stages leads to permanent abnormalities of structural layering in the neocortex and hippocampus. In contrast, conditional inactivation of the dab1 gene in the adult brain does not result in additional layering abnormalities. Furthermore, perinatal Dab1 downregulation causes behavior impairments in adult mice, such as deficits in memory, maternal care, pre-pulse inhibition, and response to cocaine. Some of these deficits were also found to be present in adolescence. We also show that D-cycloserine rescues the cognitive deficits observed in floxed dab1 mice with layering alterations in the hippocampus and neocortex. Our results indicate a causal relation between the downregulation of Dab1 protein levels during development and the structural and behavioral deficits associated with psychiatric diseases in the adult.

Overexpression of Reelin prevents the manifestation of behavioral phenotypes related to schizophrenia and bipolar disorder.

Despite the impact of schizophrenia and mood disorders, which in extreme cases can lead to death, recent decades have brought little progress in the development of new treatments. Recent studies have shown that Reelin, an extracellular protein that is critical for neuronal development, is reduced in schizophrenia and bipolar disorder patients. However, data on a causal or protective role of Reelin in psychiatric diseases is scarce. In order to study the direct influence of Reelin's levels on behavior, we subjected two mouse lines, in which Reelin levels are either reduced (Reelin heterozygous mice) or increased (Reelin overexpressing mice), to a battery of behavioral tests: open-field, black-white box, novelty-suppressed-feeding, forced-swim-test, chronic corticosterone treatment followed by forced-swim-test, cocaine sensitization and pre-pulse inhibition (PPI) deficits induced by N-methyl-D-aspartate (NMDA) antagonists. These tests were designed to model some aspects of psychiatric disorders such as schizophrenia, mood, and anxiety disorders. We found no differences between Reeler heterozygous mice and their wild-type littermates. However, Reelin overexpression in the mouse forebrain reduced the time spent floating in the forced-swim-test in mice subjected to chronic corticosterone treatment, reduced behavioral sensitization to cocaine, and reduced PPI deficits induced by a NMDA antagonist. In addition, we demonstrate that while stress increased NMDA NR2B-mediated synaptic transmission, known to be implicated in depression, Reelin overexpression significantly reduced it. Together, these results point to the Reelin signaling pathway as a relevant drug target for the treatment of a range of psychiatric disorders.