Differences in Doublecortin Immunoreactivity and Protein Levels in the Hippocampal Dentate Gyrus Between Adult and Aged Dogs

Doublecortin (DCX), a microtubule-associated protein, specifically expresses in neuronal precursors. This protein has been used as a marker for neuronal precursors and neurogenesis. In the present study, we observed differences in DCX immunoreactivity and its protein levels in the hippocampal dentate gyrus between adult and aged dogs. In the adult dog, DCX immunoreactive cells with well-stained processes were detected in the subgranular zone of the dentate gyrus. Numbers of DCX immunoreactive cells in the dentate gyrus of the aged dog were significantly decreased compared to those in the adult dog. DCX immunoreactive cells in both adult and aged dog did not show NeuN (a marker for mature neurons) immunoreactivity. NeuN immunoreactivity in the aged dog was poor compared to that in the adult dog. DCX protein level in the aged dentate gyrus was decreased by 80% compared to that in the adult dog. These results suggest that the reduction of DCX in the aged hippocampal dentate gyrus may be involved in some neural deficits related to the hippocampus.     

Adult neurogenesis in the mammalian brain: significant answers and significant questions

Adult neurogenesis, a process of generating functional neurons from adult neural precursors, occurs throughout life in restricted brain regions in mammals. The past decade has witnessed tremendous progress in addressing questions related to almost every aspect of adult neurogenesis in the mammalian brain. Here we review major advances in our understanding of adult mammalian neurogenesis in the dentate gyrus of the hippocampus and from the subventricular zone of the lateral ventricle, the rostral migratory stream to the olfactory bulb. We highlight emerging principles that have significant implications for stem cell biology, developmental neurobiology, neural plasticity, and disease mechanisms. We also discuss remaining questions related to adult neural stem cells and their niches, underlying regulatory mechanisms, and potential functions of newborn neurons in the adult brain. Building upon the recent progress and aided by new technologies, the adult neurogenesis field is poised to leap forward in the next decade.     

Temporal Changes in Prosaposin Expression in the Rat Dentate Gyrus after Birth

Neurogenesis in the hippocampal dentate gyrus occurs constitutively throughout postnatal life. Adult neurogenesis includes a multistep process that ends with the formation of a postmitotic and functionally integrated new neuron. During adult neurogenesis, various markers are expressed, including GFAP, nestin, Pax6, polysialic acid-neural cell adhesion molecule (PSA-NCAM), neuronal nuclei (NeuN), doublecortin, TUC-4, Tuj-1, and calretinin. Prosaposin is the precursor of saposins A–D; it is found in various organs and can be excreted. Strong prosaposin expression has been demonstrated in the developing brain including the hippocampus, and its neurotrophic activity has been proposed. This study investigated changes in prosaposin in the dentate gyrus of young and adult rats using double immunohistochemistry with antibodies to prosaposin, PSA-NCAM, and NeuN. Prosaposin immunoreactivity was intense in the dentate gyrus at postnatal day 3 (P3) and P7, but decreased gradually after P14. In the dentate gyrus at P28, immature PSA-NCAM-positive neurons localized exclusively in the subgranular zone were prosaposin-negative, whereas mature Neu-N-positive neurons were positive for prosaposin. Furthermore, these prosaposin-negative immature neurons were saposin B-positive, suggesting that the neurons take up and degrade prosaposin. In situ hybridization assays showed that prosaposin in the adult dentate gyrus is dominantly the Pro+9 type, a secreted type of prosaposin. These results imply that prosaposin secreted from mature neurons stimulates proliferation and maturation of immature neurons in the dentate gyrus.     

The microtubule destabilizing protein stathmin controls the transition from dividing neuronal precursors to postmitotic neurons during adult hippocampal neurogenesis

The hippocampus is one of the two areas in the mammalian brain where adult neurogenesis occurs. Adult neurogenesis is well known to be involved in hippocampal physiological functions as well as pathophysiological conditions. Microtubules (MTs), providing intracellular transport, stability, and transmitting force, are indispensable for neurogenesis by facilitating cell division, migration, growth, and differentiation. Although there are several examples of MT‐stabilizing proteins regulating different aspects of adult neurogenesis, relatively little is known about the function of MT‐destabilizing proteins. Stathmin is such a MT‐destabilizing protein largely restricted to the CNS, and in contrast to its developmental family members, stathmin is also expressed at significant levels in the adult brain, notably in areas involved in adult neurogenesis. Here, we show an important role for stathmin during adult neurogenesis in the subgranular zone of the mouse hippocampus. After carefully mapping stathmin expression in the adult dentate gyrus (DG), we investigated its role in hippocampal neurogenesis making use of stathmin knockout mice. Although hippocampus development appears normal in these animals, different aspects of adult neurogenesis are affected. First, the number of proliferating Ki‐67+ cells is decreased in stathmin knockout mice, as well as the expression of the immature markers Nestin and PSA‐NCAM. However, newborn cells that do survive express more frequently the adult marker NeuN and have a more mature morphology. Furthermore, our data suggest that migration in the DG might be affected. We propose a model in which stathmin controls the transition from neuronal precursors to early postmitotic neurons. © 2014 Wiley Periodicals, Inc. Develop Neurobiol 74: 1226–1242, 2014     

Time-of-Day-Dependent Enhancement of Adult Neurogenesis in the Hippocampus

Background

Adult neurogenesis occurs in specific regions of the mammalian brain such as the dentate gyrus of the hippocampus. In the neurogenic region, neural progenitor cells continuously divide and give birth to new neurons. Although biological properties of neurons and glia in the hippocampus have been demonstrated to fluctuate depending on specific times of the day, it is unclear if neural progenitors and neurogenesis in the adult brain are temporally controlled within the day.

Methodology/Principal Findings

Here we demonstrate that in the dentate gyrus of the adult mouse hippocampus, the number of M-phase cells shows a day/night variation throughout the day, with a significant increase during the nighttime. The M-phase cell number is constant throughout the day in the subventricular zone of the forebrain, another site of adult neurogenesis, indicating the daily rhythm of progenitor mitosis is region-specific. Importantly, the nighttime enhancement of hippocampal progenitor mitosis is accompanied by a nighttime increase of newborn neurons.

Conclusions/Significance

These results indicate that neurogenesis in the adult hippocampus occurs in a time-of-day-dependent fashion, which may dictate daily modifications of dentate gyrus physiology.     

Targeting the Hippocampal Mossy Fiber Synapse for the Treatment of Psychiatric Disorders

It is widely known that new neurons are continuously generated in the dentate gyrus of the hippocampus in the adult mammalian brain. This neurogenesis has been implicated in depression and antidepressant treatments. Recent evidence also suggests that the dentate gyrus is involved in the neuropathology and pathophysiology of schizophrenia and other related psychiatric disorders. Especially, abnormal neuronal development in the dentate gyrus may be a plausible risk factor for the diseases. The synapse made by the mossy fiber, the output fiber of the dentate gyrus, plays a critical role in regulating neuronal activity in its target CA3 area. The mossy fiber synapse is characterized by remarkable activity-dependent short-term synaptic plasticity that is established during the postnatal development and is supposed to be central to the functional role of the mossy fiber. Any defects, including developmental abnormalities, in the dentate gyrus and drugs acting on the dentate gyrus can modulate the mossy fiber-CA3 synaptic transmission, which may eventually affect hippocampal functions. In this paper, I review recent evidence for involvement of the dentate gyrus and mossy fiber synapse in psychiatric disorders and discuss potential importance of drugs targeting the mossy fiber synapse either directly or indirectly in the therapeutic treatments of psychiatric disorders.     

Blockade of L-type voltage-gated Ca channel inhibits ischemia-induced neurogenesis by down-regulating iNOS expression in adult mouse

Neurogenesis in the adult mammalian hippocampus may contribute to repairing the brain after injury. The signals that regulate neurogenesis in the dentate gyrus following ischemic stroke insult are not well known. We have previously reported that inducible nitric oxide synthase (iNOS) expression is necessary for ischemia-stimulated neurogenesis in the adult dentate gyrus. Here, we show that mice subjected to 90 min of middle cerebral artery occlusion (MCAO) significantly increased the number of new neurons and up-regulated iNOS expression in the dentate gyrus. Blockade of the L-type voltage-gated Ca(2+) channel (L-VGCC) prevented neurogenesis in the dentate gyrus and subventricular zone (SVZ), and down-regulated iNOS expression in the dentate gyrus after cerebral ischemia. This study suggests that Ca(2+) influx through L-VGCC is involved in ischemia-induced neurogenesis by up-regulating iNOS expression.     

Development of neural stem cell in the adult brain

New neurons are continuously generated in the dentate gyrus of the mammalian hippocampus and in the subventricular zone of the lateral ventricles throughout life. The origin of these new neurons is believed to be from multipotent adult neural stem cells. Aided by new methodologies, significant progress has been made in the characterization of neural stem cells and their development in the adult brain. Recent studies have also begun to reveal essential extrinsic and intrinsic molecular mechanisms that govern sequential steps of adult neurogenesis in the hippocampus and subventricular zone/olfactory bulb, from proliferation and fate specification of neural progenitors to maturation, navigation, and synaptic integration of the neuronal progeny. Future identification of molecular mechanisms and physiological functions of adult neurogenesis will provide further insight into the plasticity and regenerative capacity of the mature central nervous system.     

Wnt7a Regulates Multiple Steps of Neurogenesis

Although Wnt7a has been implicated in axon guidance and synapse formation, investigations of its role in the early steps of neurogenesis have just begun. We show here that Wnt7a is essential for neural stem cell self-renewal and neural progenitor cell cycle progression in adult mouse brains. Loss of Wnt7a expression dramatically reduced the neural stem cell population and increased the rate of cell cycle exit in neural progenitors in the hippocampal dentate gyrus of adult mice. Furthermore, Wnt7a is important for neuronal differentiation and maturation. Loss of Wnt7a expression led to a substantial decrease in the number of newborn neurons in the hippocampal dentate gyrus. Wnt7a^−/− dentate granule neurons exhibited dramatically impaired dendritic development. Moreover, Wnt7a activated β-catenin and its downstream target genes to regulate neural stem cell proliferation and differentiation. Wnt7a stimulated neural stem cell proliferation by activating the β-catenin–cyclin D1 pathway and promoted neuronal differentiation and maturation by inducing the β-catenin–neurogenin 2 pathway. Thus, Wnt7a exercised critical control over multiple steps of neurogenesis by regulating genes involved in both cell cycle control and neuronal differentiation.     

The common properties of neurogenesis in the adult brain: from invertebrates to vertebrates

Until recently, it was believed that adult brains were unable to generate any new neurons. However, it is now commonly known that stem cells remain in the adult central nervous system and that adult vertebrates as well as adult invertebrates are currently adding new neurons in some specialized structures of their central nervous system. In vertebrates, the subventricular zone and the dentate gyrus of the hippocampus are the sites of neuronal precursor proliferation. In some insects, persistent neurogenesis occurs in the mushroom bodies, which are brain structures involved in learning and memory and considered as functional analogues of the hippocampus. In both vertebrates and invertebrates, secondary neurogenesis (including neuroblast proliferation and neuron differentiation) appears to be regulated by hormones, transmitters, growth factors and environmental cues. The functional implications of adult neurogenesis have not yet been clearly demonstrated and comparative study of the various model systems could contribute to better understand this phenomenon. Here, we review and discuss the common characteristics of adult neurogenesis in the various animal models studied so far.     

成年哺乳动物皮质区神经发生的研究进展

神经发生是神经干细胞在适当的条件下分化成功能性整合神经元的过程,主要包括细胞的增殖、迁移、分化和存活。成年神经发生区以前脑室管膜下区(Subventricular zones,SVZ)和海马齿状回颗粒层下区(Subgranular zones,SGZ)为主,但皮质作为神经元和神经胶质细胞数量最多、分布最广,同时也是哺乳动物高度发展的脑区,是否有成年神经元新生,已成为近年来神经科学领域的研究热点[1,2]。现本文就未成熟神经元在皮质区的研究方法、分布、来源与转归、病理生理功能影响等方面探讨成年哺乳动物皮质神经发生现象。     

Deficient neurogenesis in forebrain-specific presenilin-1 knockout mice is associated with reduced clearance of hippocampal memory traces.

To examine the in vivo function of presenilin-1 (PS1), we selectively deleted the PS1 gene in excitatory neurons of the adult mouse forebrain. These conditional knockout mice were viable and grew normally, but they exhibited a pronounced deficiency in enrichment-induced neurogenesis in the dentate gyrus. This reduction in neurogenesis did not result in appreciable learning deficits, indicating that addition of new neurons is not required for memory formation. However, our postlearning enrichment experiments lead us to postulate that adult dentate neurogenesis may play a role in the periodic clearance of outdated hippocampal memory traces after cortical memory consolidation, thereby ensuring that the hippocampus is continuously available to process new memories. A chronic, abnormal clearance process in the hippocampus may conceivably lead to memory disorders in the mammalian brain.     

Decreased Hippocampal Neurogenesis Following Olfactory Bulbectomy is Reversed by Repeated Citalopram Administration

1. Whereas much progress has been made in the treatment of depression, the exact pathogenetic mechanisms of the disorder are still poorly understood. It has been proposed that one possible mechanism could be a decrease in adult hippocampal neurogenesis.2. The olfactory bulbectomy (OB) in rats is widely accepted as an animal model of depression. In the present study, we investigated whether hippocampal neurogenesis is affected by an OB, and whether chronic citalopram, a serotonin selective reuptake inhibitor, counteracts OB-induced impairment of neurogenesis.3. Our study shows that OB decreases proliferation of the neuronal precursors in the dentate gyrus and retards their differentiation into mature granule neurons. In OB rats, repeated administration of citalopram restores reduced proliferative activity and enhances the differentiation of precursors into mature calbindin-positive neurons.4. The obtained data demonstrate that a citalopram-induced increase in neurogenesis in OB rats could be one possible mechanism by which antidepressants alleviate OB-induced depressive-like behavior.     

Region- and neuronal phenotype-specific expression of NELL2 in the adult rat brain

NELL2, a neural tissue-enriched protein, is produced in the embryo, and postembryonically in the mammalian brain, with a broad distribution. Although its synthesis is required for neuronal differentiation in chicks, not much is known about its function in the adult mammalian brain. We investigated the distribution of NELL2 in various regions of the adult rat brain to study its potential functions in brain physiology. Consistent with previous reports, NELL2-immunoreactivity (ir) was found in the cytoplasm of neurons, but not in glial fibrillary acidic protein (GFAP)-positive glial cells. The highest levels of NELL2 were detected in the hippocampus and the cerebellum. Interestingly, in the cerebellar cortex NELL2 was observed only in the GABAergic Purkinje cells not in the excitatory granular cells. In contrast, it was found mainly in the hippocampal dentate gyrus and pyramidal cell layer that contains mainly glutamatergic neurons. In the dentate gyrus, NELL2 was not detected in the GFAP-positive neural precursor cells, but was generally present in mature neurons of the subgranular zone, suggesting a role in this region restricted to mature neurons.     

Neurogenin 2 has an essential role in development of the dentate gyrus

The dentate gyrus (DG) of the hippocampus has a central role in learning and memory in adult rodents. The DG is generated soon after birth, although new neurons continue to be generated in the DG throughout life. The proneural factors Mash1 (Ascl1) and neurogenin 2 (Ngn2) are expressed during formation of the DG but their role in the development of this structure has not yet been addressed. Here, we show that Ngn2 is essential for the development of the DG. Ngn2 mutant mice have fewer DG progenitors and these cells present defects in neuronal differentiation. By contrast, the DG is normal in Mash1 mutant mice at birth, and loss of both Mash1 and Ngn2 does not aggravate the defect observed in Ngn2 single mutants. These data establish a unique role of Ngn2 in DG neurogenesis during development and raise the possibility that Ngn2 has a similar function in adult neurogenesis.     

Three-dimensional ultrastructural and immunohistochemical study of immature neurons in the subgranular zone of the rat dentate gyrus

The present study is devoted to three-dimensional ultrastructural organization of mitotically dividing immature neurons in dentate gyrus using biophysical approaches. In adult vertebrate brain, cell proliferation persists throughout life mainly in dentate gyrus of the hippocampus (DG) and olfactory bulb. Neurogenesis has been demonstrated using tagged thymidine analogues incorporated into the S phase of the cell cycle, but these may also detect repaired DNA in postmitotic neurons. Recent retroviral labelling has shown that neuronal progenitors/neuroblasts divide and produce functional neurons. Providing ultrastructural evidence of mitotically active cells has proven problematical, not only because of technical issues of identifying dividing cells at electron microscope level, but also because it is difficult to demonstrate unequivocally that neurons identified in the electron microscope are really post mitotic. However by characterising post mitotic cells labelled with BrdU and doublecortin and comparing these with post mitotic cells reconstructed in 3-dimensions from ultrathin serial sections, we have been able to illustrate individual mitotic elements and phases of cells within the GC layer of adult rat dentate gyrus. Here we show dividing cells in metaphase within clusters of immature GCs in subgranular zone (SGZ). These reconstructions provide ultrastructural confirmation that cells expressing doublecortin (DCX), a microtubule-associated protein expressed in migrating neurons, localize as clusters in the subgranular zone (SGZ) of dentate gyrus (DG) in the hippocampus during all animal life. Such DG cells with clear synaptic specializations, somatic spines and basal dendrites are exclusive to immature GC that appear to re-enter the cell cycle, suggesting the possibility that newly generated neurons within the DG might arise not only from precursors, but also from clusters of immature GC.     

Spatial relational memory requires hippocampal adult neurogenesis

The dentate gyrus of the hippocampus is one of the few regions of the mammalian brain where new neurons are generated throughout adulthood. This adult neurogenesis has been proposed as a novel mechanism that mediates spatial memory. However, data showing a causal relationship between neurogenesis and spatial memory are controversial. Here, we developed an inducible transgenic strategy allowing specific ablation of adult-born hippocampal neurons. This resulted in an impairment of spatial relational memory, which supports a capacity for flexible, inferential memory expression. In contrast, less complex forms of spatial knowledge were unaltered. These findings demonstrate that adult-born neurons are necessary for complex forms of hippocampus-mediated learning.