Enhanced production and dendritic growth of new dentate granule cells in the middle-aged hippocampus following intracerebroventricular FGF-2 infusions

Declined production and diminished dendritic growth of new dentate granule cells in the middle-aged and aged hippocampus are correlated with diminished concentration of fibroblast growth factor-2 (FGF-2). This study examined whether increased FGF-2 concentration in the milieu boosts both production and dendritic growth of new dentate granule cells in the middle-aged hippocampus. The FGF-2 or vehicle was infused into the posterior lateral ventricle of middle-aged Fischer (F)344 rats for 2 weeks using osmotic minipumps. New cells born during the first 12 days of infusions were labeled via daily intraperitoneal injections of 5'-bromodeoxyuridine (BrdU) and analysed at 10 days after the last BrdU injection. Measurement of BrdU(+) cells revealed a considerably enhanced number of new cells in the subgranular zone (SGZ) and granule cell layer (GCL) of the dentate gyrus (DG) ipsilateral to FGF-2 infusions. Characterization of beta-III tubulin(+) neurons among newly born cells suggested an increased addition of new neurons to the SGZ/GCL ipsilateral to FGF-2 infusions. Quantification of DG neurogenesis at 8 days post-infusions via doublecortin (DCX) immunostaining also revealed the presence of an enhanced DG neurogenesis ipsilateral to FGF-2 infusions. Furthermore, DCX(+) neurons in FGF-2-infused rats exhibited enhanced dendritic growth compared with their counterparts in vehicle-infused rats. Thus, subchronic infusion of FGF-2 is efficacious for stimulating an enhanced DG neurogenesis from neural stem/progenitor cells in the middle-aged hippocampus. As dentate neurogenesis is important for hippocampal-dependent learning and memory and DG long-term potentiation, strategies that maintain increased FGF-2 concentration during ageing may be beneficial for thwarting some of the age-related cognitive impairments.     

Deafferentation enhances neurogenesis in the young and middle aged hippocampus but not in the aged hippocampus

Increased neurogenesis in the dentate gyrus (DG) after brain insults such as excitotoxic lesions, seizures, or stroke is a well known phenomenon in the young hippocampus. This plasticity reflects an innate compensatory response of neural stem cells (NSCs) in the young hippocampus to preserve function or minimize damage after injury. However, injuries to the middle‐aged and aged hippocampi elicit either no or dampened neurogenesis response, which could be due to an altered plasticity of NSCs and/or the hippocampus with age. We examined whether the plasticity of NSCs to increase neurogenesis in response to a milder injury such as partial deafferentation is preserved during aging. We quantified DG neurogenesis in the hippocampus of young, middle‐aged, and aged F344 rats after partial deafferentation. A partial deafferentation of the left hippocampus without any apparent cell loss was induced via administration of Kainic acid (0.5 μg in 1.0 μl) into the right lateral ventricle of the brain. In this model, degeneration of CA3 pyramidal neurons and dentate hilar neurons in the right hippocampus results in loss of commissural axons which leads to partial deafferentation of the dendrites of dentate granule cells and CA1‐CA3 pyramidal neurons in the left hippocampus. Quantification of newly born cells that are added to the dentate granule cell layer at postdeafferentation days 4–15 using 5′‐bromodeoxyuridine (BrdU) labeling revealed greatly increased addition of newly born cells (～three fold increase) in the deafferented young and middle‐aged hippocampi but not in the deafferented aged hippocampus. Measurement of newly born neurons using doublecortin (DCX) immunostaining also revealed similar findings. Analyses using BrdU‐DCX dual immunofluorescence demonstrated no changes in neuronal fate‐choice decision of newly born cells after deafferentation, in comparison to the age‐matched naive hippocampus in all age groups. Thus, the plasticity of hippocampal NSCs to increase DG neurogenesis in response to a milder injury such as partial hippocampal deafferentation is preserved until middle age but lost at old age. © 2010 Wiley‐Liss, Inc.     

Decreased neuronal differentiation of newly generated cells underlies reduced hippocampal neurogenesis in chronic temporal lobe epilepsy

Hippocampal neurogenesis declines substantially in chronic temporal lobe epilepsy (TLE). However, it is unclear whether this decline is linked to altered production of new cells and/or diminished survival and neuronal fate‐choice decision of newly born cells. We quantified different components of hippocampal neurogenesis in rats exhibiting chronic TLE. Through intraperitoneal administration of 5′‐bromodeoxyuridine (BrdU) for 12 days, we measured numbers of newly born cells in the subgranular zone‐granule cell layer (SGZ‐GCL) at 24 h and 2.5 months post‐BrdU administration. Furthermore, the differentiation of newly added cells into neurons and glia was quantified via dual immunofluorescence for BrdU and various markers of neurons and glia. Addition of new cells to the SGZ‐GCL over 12 days was comparable between the chronically epileptic hippocampus and the age‐matched intact hippocampus. Furthermore, comparison of BrdU+ cells measured at 24 h and 2.5 months post‐BrdU administration revealed similar survival of newly born cells between the two groups. However, only 4–5% of newly born cells (i.e., BrdU+ cells) differentiated into neurons in the chronically epileptic hippocampus, in comparison to 73–80% of such cells exhibiting neuronal differentiation in the intact hippocampus. Moreover, differentiation of newly born cells into S‐100β+ astrocytes or NG2+ oligodendrocyte progenitors increased to ∼79% in the chronically epileptic hippocampus from ∼25% observed in the intact hippocampus. Interestingly, the extent of proliferation of astrocytes and microglia (identified through Ki‐67 and S‐100β and Ki‐67 and OX‐42 dual immunofluorescence) in the SGZ‐GCL was similar between the chronically epileptic hippocampus and the age‐matched intact hippocampus, implying that the proliferation of neural stem/progenitor cells in the SGZ‐GCL of the chronically epileptic hippocampus was not obscured by an increased division of glia. Thus, severely diminished DG neurogenesis in chronic TLE is not associated with either decreased production of new cells or reduced survival of newly born cells in the SGZ‐GCL. Rather, it is linked to a dramatic decline in the neuronal fate‐choice decision of newly generated cells. Overall, the differentiation of newly born cells turns mainly into glia with chronic TLE from predominantly neuronal differentiation seen in control conditions. © 2009 Wiley‐Liss, Inc.     

CXCR4 prevents dispersion of granule neuron precursors in the adult dentate gyrus

Neurogenesis in the adult dentate gyrus (DG) generates new granule neurons that differentiate in the inner one‐third of the granule cell layer (GCL). The migrating precursors of these neurons arise from neural stem cells (NSCs) in the subgranular zone (SGZ). Although it is established that pathological conditions, including epilepsy and stroke, cause dispersion of granule neuron precursors, little is known about the factors that regulate their normal placement. Based on the high expression of the chemokine CXCL12 in the adult GCL and its role in guiding neuronal migration in development, we addressed the function of the CXCL12 receptor CXCR4 in adult neurogenesis. Using transgenic reporter mice, we detected Cxcr4‐GFP expression in NSCs, neuronal‐committed progenitors, and immature neurons of adult and aged mice. Analyses of hippocampal NSC cultures and hippocampal tissue by immunoblot and immunohistochemistry provided evidence for CXCL12‐promoted phosphorylation/activation of CXCR4 receptors in NSCs in vivo and in vitro. Cxcr4 deletion in NSCs of the postnatal or mature DG using Cre technology reduced neurogenesis. Fifty days after Cxcr4 ablation in the mature DG, the SGZ showed a severe reduction of Sox2‐positive neural stem/early progenitor cells, NeuroD‐positive neuronal‐committed progenitors, and DCX‐positive immature neurons. Many immature neurons were ectopically placed in the hilus and inner molecular layer, and some developed an aberrant dendritic morphology. Only few misplaced cells survived permanently as ectopic neurons. Thus, CXCR4 signaling maintains the NSC pool in the DG and specifies the inner one‐third of the GCL as differentiation area for immature granule neurons. © 2013 Wiley Periodicals, Inc.     

Newly born cells in the ageing dentate gyrus display normal migration, survival and neuronal fate choice but endure retarded early maturation

Addition of new granule cells to the dentate gyrus (DG) from stem or progenitor cells declines considerably during ageing. However, potential age-related alterations in migration, enduring survival and neuronal fate choice of newly born cells, and rate of maturation and dendritic growth of newly differentiated neurons are mostly unknown. We addressed these issues by analysing cells that are positive for 5'-bromodeoxyuridine (BrdU), doublecortin (DCX), BrdU and DCX, and BrdU and neuron-specific nuclear antigen (NeuN) in the DG of young adult, middle-aged and aged F344 rats treated with daily injections of BrdU for 12 consecutive days. Analyses performed at 24 h, 10 days and 5 months after BrdU injections reveal that the extent of new cell production decreases dramatically by middle age but exhibits no change thereafter. Interestingly, fractions of newly formed cells that exhibit appropriate migration and prolonged survival, and fractions of newly born cells that differentiate into neurons, remain stable during ageing. However, in newly formed neurons of the middle-aged and aged DG, the expression of mature neuronal marker NeuN is delayed and early dendritic growth is retarded. Thus, the presence of far fewer new granule cells in the aged DG is not due to alterations in the long term survival and phenotypic differentiation of newly generated cells but solely owing to diminished production of new cells. The results also underscore that the capability of the DG milieu to support neuronal fate choice, migration and enduring survival of newly born cells remains stable even during senescence but its ability to promote rapid neuronal maturation and dendritic growth is diminished as early as middle age.     

Efficacy of doublecortin as a marker to analyse the absolute number and dendritic growth of newly generated neurons in the adult dentate gyrus

Doublecortin (DCX), a microtubule-associated phosphoprotein, has been recently utilized as a marker of newly born neurons in the adult dentate gyrus (DG). Nonetheless, it is unknown whether DCX exclusively labels newly formed neurons, as certain granule cells with the phenotype of differentiated neurons express DCX. We addressed the authenticity of DCX as a marker of new neurons in the adult DG by quantifying cells that are positive for 5'-bromodeoxyuridine (BrdU), DCX and both BrdU and DCX in hippocampal tissues of adult rats treated with daily injections of BrdU for 12 consecutive days. We provide new evidence that neurons visualized with DCX immunostaining in the adult rat DG are new neurons that are predominantly born during the 12 days before euthanasia. This is confirmed by the robust expression of BrdU in 90% of DCX-positive neurons in the DG of animals injected with BrdU for 12 days. Furthermore, DCX expression is specific to newly generated healthy neurons, as virtually all DCX-positive cells express early neuronal antigens but lack antigens specific to glia, undifferentiated cells or apoptotic cells. As DCX expression is also robust in the dendrites, DCX immunocytochemistry of thicker sections facilitates quantification of the dendritic growth in newly born neurons. Thus, both absolute number and dendritic growth of new neurons that are generated in the adult DG over a 12-day period can be quantified reliably with DCX immunostaining. This could be particularly useful for analysing changes in dentate neurogenesis in human hippocampal tissues as a function of ageing or neurodegenerative diseases.     

Long-term neuronal replacement in adult rat hippocampus after status epilepticus despite chronic inflammation

Dentate gyrus (DG) neurogenesis is transiently increased during the first weeks after status epilepticus (SE). Survival of the new neurons is initially compromised by an acute inflammatory response, but the long-term fate of the remaining ones in the post-SE environment is unknown. Here adult rats were subjected to 2 h electrically evoked self-sustained SE and perfused after 5 weeks or 6 months. Rats exhibited partial or generalized SE followed by spontaneous behavioural seizures and abnormal electroencephalographic activity during 6 months. Numbers of activated microglia in the dentate subgranular zone (SGZ)-granule cell layer (GCL) and in the hilus declined after 5 weeks, but were still elevated at 6 months after SE, with no differences between the milder partial and the more severe generalized SE. At 6 months, partial and generalized SE rats showed a seven-fold increase in the number of mature SGZ-GCL neurons formed during the first 2 weeks along with aberrant neurons in the hilus. Total numbers of mature neurons in SGZ-GCL were unaltered, indicating that SE-generated neurons replaced dead granule cells. Neuroblast formation had returned to normal levels in SGZ-GCL but generation of aberrant neurons in the hilus was still ongoing at 6 months. Our data indicate that long-term impairment of neurogenesis, as reported previously after kainic acid-induced SE, is not a general feature of chronic epilepsy. We have found that a substantial proportion of the mature granule cells at 6 months are generated during the first 2 weeks after SE and survive despite chronic inflammation, and that SE triggers continuous production of aberrant hilar neurons.     

Injury-Induced Neurogenesis: Consideration of Resident Microglia as Supportive of Neural Progenitor Cells

The induction of neurogenesis in the adult subgranular zone (SGZ) by injury is often accompanied by changes in the extracellular environment that can have significant impacts on neural progenitor cells (NPCs). We examined the induction of neurogenesis in the SGZ at 72 h following an injection of the hippocampal toxicant, trimethyltin (TMT; 2 mg/kg, ip) inducing apoptosis in dentate granule neurons. BrdU+ incorporation during the active period of neuronal death indicated NPC proliferation and migration of newly generated cells into the granule cell layer (GCL). BrdU+ cells were transiently in contact with process bearing microglia within the inner SGZ layer. Contact with GFAP+ astrocyte processes occurred once cells were within the GCL. A small percentage of the BrdU+ cells within the SGZ region showed immunoreactivity for tumor necrosis factor (TNF) p75 receptor (TNFp75R). In mice deficient for TNFp75R, TMT injection produced an equivalent level of dentate granule cell death however; BrdU+ cells were localized at the SGZ as compared to the presence of cells within the GCL in the WT mice dosed with TMT. These data suggest that cells generated by NPCs in the SGZ induced with a focal lesion to the dentate granule neurons of adolescent mice maintain the capacity to utilize the neuroinflammation and microglia responses within their environment for migration into the GCL.     

Targeted transgene expression in neuronal precursors: watching young neurons in the old brain

Progress in the field of neurogenesis is limited by the lack of animal models allowing direct detection and analysis of living cells participating in neurogenesis. We engineered a transgenic mouse model that expresses the fluorescent reporter proteins enhanced green fluorescent protein or Discoma sp. reef coral red fluorescent protein under the control of the doublecortin (DCX) promoter, a gene specifically and transiently active in neuronal precursors and young neurons. The expression of the reporter proteins correlated with expression of the endogenous DCX protein, and with developmental and adult neurogenesis. Neurogenesis was unaffected by the presence of the fluorescent proteins. The transgenic mice allowed direct identification of the very few newly generated neurons present in the aged brain. We performed electrophysiological analysis and established that newly generated hippocampal granule cells in aged and young mice shared identical physiological properties. Hence, although the rate of neurogenesis tapers with ageing, a population of highly excitable young neurons indistinguishable to those found in younger animals is continuously generated. Therefore, maintenance of the fundamental properties of neuronal precursors even at advanced age suggests that stimulation of neurogenesis may constitute a valid strategy to counteract age-related neuronal loss and cognitive declines.     

Long-term survival and cell death of newly generated neurons in the adult rat olfactory bulb

In the adult rat olfactory bulb, neurons are continually generated from progenitors that reside in the lateral ventricle wall. This study investigates long-term survival and cell death of newly generated cells within the adult olfactory bulb. After injecting rats at 2 months of age with 5-bromodeoxyuridine (BrdU), the newly generated cells were quantified over a period of 19 months. A peak of BrdU-positive cells was reached in the olfactory bulb 1 month after BrdU injection, when all new cells have finished migrating from the ventricle wall. Thereafter, a reduction of BrdU-positive cells to about 50% was observed and it was confirmed by dUTP-nick end-labelling (TUNEL) that progenitors and young neurons undergo programmed cell death. However, cells that survived the first 3 months after BrdU injection persisted for up to 19 months. The majority of the BrdU-positive cells that reach the olfactory bulb differentiate into granule cells, but a small fraction migrate further into the glomerular layer. These newborn cells differentiate more slowly into periglomerular interneurons, with a delay of more than 1 month when compared to the granule cells. The newly generated periglomerular neurons, among them a significant fraction of dopaminergic cells, showed a similar decline in number compared to the granule cell layer and long-term survival for the remaining new neurons of up to 19 months. Rather than replacing old neurons, this data suggests that adult olfactory bulb neurogenesis utilizes the overproduction and turnover of young neurons, which is reminiscent of the cellular dynamics observed during brain development.     

Chronic temporal lobe epilepsy is associated with severely declined dentate neurogenesis in the adult hippocampus

While it is clear that acute hippocampal injury or status epilepticus increases the production of new neurons in the adult dentate gyrus (DG), the effects of chronic epilepsy on dentate neurogenesis are unknown. We hypothesize that epileptogenic changes and spontaneous recurrent motor seizures (SRMS) that ensue after hippocampal injury or status epilepticus considerably decrease dentate neurogenesis. We addressed this issue by quantifying the number of cells that are positive for doublecortin (DCX, a marker of new neurons) in the DG of adult F344 rats at 16 days and 5 months after an intracerebroventricular kainic acid (ICV KA) administration or after graded intraperitoneal KA (IP KA) injections, models of temporal lobe epilepsy (TLE). At early post-KA administration, the injured hippocampus exhibited increased dentate neurogenesis in both models. Conversely, at 5 months post-KA administration, the chronically epileptic hippocampus demonstrated severely declined neurogenesis, which was associated with considerable SRMS in both KA models. Additionally, stem/progenitor cell proliferation factors, FGF-2 and IGF-1, were decreased in the chronically epileptic hippocampus. Interestingly, the overall decrease in neurogenesis and the extent of SRMS were greater in rats receiving IP KA than rats receiving ICV KA, suggesting that the extent of neurogenesis during chronic TLE exhibits an inverse relationship with SRMS. These results provide novel evidence that chronic TLE is associated with extremely declined dentate neurogenesis. As fraction of newly born neurons become GABA-ergic interneurons, declined neurogenesis may contribute to the increased seizure-susceptibility of the DG in chronic TLE. Likewise, the hippocampal-dependent learning and memory deficits observed in chronic TLE could be linked at least partially to the declined neurogenesis.     

Caspase inhibitors increase short-term survival of progenitor-cell progeny in the adult rat dentate gyrus following status epilepticus

The dentate gyrus (DG) is one of the few regions in the brain that continues to produce new neurons throughout adulthood. Seizures not only increase neurogenesis, but also lead to death of DG neurons. We investigated the relationship between cell death and neurogenesis following seizures in the DG of adult rats by blocking caspases, which are key components of apoptotic cell death. Multiple intracerebroventricular infusions of caspase inhibitors (pancaspase inhibitor zVADfmk, and caspase 3 and 9 inhibitor) prior to, just after, 1 day after, and 1 week following 2 h of lithium-pilocarpine-induced status epilepticus reduced the number of terminal deoxynucleotidyl transferase-mediated fluorescein-dUTP nick-end labelled (TUNEL) cells and increased the number of bromodeoxyuridine (BrdU) -stained proliferated cells in the subgranular zone at 1 week. The caspase inhibitor-treated group did not differ from control at 2 days or 5 weeks following the epileptic insult. Our findings suggest that caspases modulate seizure-induced neurogenesis in the DG, probably by regulating apoptosis of newly born neurons, and that this action can be suppressed transiently by caspase inhibitors. Furthermore, although previous studies have indicated that increased neuronal death can trigger neurogenesis, we show here that reduction in apoptotic death may be associated with increased neurogenesis.     

Newly born dentate granule neurons after pilocarpine-induced epilepsy have hilar basal dendrites with immature synapses

Neurogenesis in the subgranular zone of the dentate gyrus persists throughout the lifespan of mammals, and the resulting newly born neurons are incorporated into existing hippocampal circuitry. Seizures increase the rate of neurogenesis in the adult rodent brain and result in granule cells in the dentate gyrus with basal dendrites. Using doublecortin (DCX) immunocytochemistry to label newly generated neurons the current study focuses on the electron microscopic features of DCX-labeled cell bodies and dendritic processes in the dentate gyrus of rats with pilocarpine-induced epilepsy. At the base of the granule cell layer clusters of cells that include up to six DCX-labeled cell bodies were observed. The cell bodies in these clusters lacked a one-to-one association with an astrocyte cell body and its processes, a relationship that is typical for newly born granule cells in control rats. Also, DCX-labeled basal dendrites in the hilus had immature synapses while those in control rats lacked synapses. These results indicate that increased neurogenesis after seizures alters the one-to-one relationship between astrocytes and DCX-labeled newly generated neurons at the base of the granule cell layer. The data also suggest that the synapses on DCX-labeled hilar basal dendrites contribute to the persistence of hilar basal dendrites on neurons born after pilocarpine-induced seizures.     

Early survival and delayed death of developmentally‐born dentate gyrus neurons

The storage and persistence of memories depends on plasticity in the hippocampus. Adult neurogenesis produces new neurons that mature through critical periods for plasticity and cellular survival, which determine their contributions to learning and memory. However, most granule neurons are generated prior to adulthood; the maturational timecourse of these neurons is poorly understood compared to adult‐born neurons but is essential to identify how the dentate gyrus (DG), as a whole, contributes to behavior. To characterize neurons born in the early postnatal period, we labeled DG neurons born on postnatal day 6 (P6) with BrdU and quantified maturation and survival across early (1 hr to 8 weeks old) and late (2–6 months old) cell ages. We find that the dynamics of developmentally‐born neuron survival is essentially the opposite of neurons born in adulthood: P6‐born neurons did not go through a period of cell death during their immature stages (from 1 to 8 weeks). In contrast, 17% of P6‐born neurons died after reaching maturity, between 2 and 6 months of age. Delayed death was evident from the loss of BrdU⁺ cells as well as pyknotic BrdU⁺caspase3⁺ neurons within the superficial granule cell layer. Patterns of DCX, NeuN, and activity‐dependent Fos expression indicate that developmentally‐born neurons mature over several weeks and a sharp peak in zif268 expression at 2 weeks suggests that developmentally‐born neurons mature faster than adult‐born neurons (which peak at 3 weeks). Collectively, our findings are relevant for understanding how developmentally‐born DG neurons contribute to memory and disorders throughout the lifespan. High levels of early survival and zif268 expression may promote learning, while also rendering neurons sensitive to insults at defined stages. Late neuronal death in young adulthood may result in the loss of hundreds of thousands of DG neurons, which could impact memory persistence and contribute to hippocampal/DG atrophy in disorders such as depression.     

Positive Controls in Adults and Children Support That Very Few,If Any,New Neurons Are Born in the Adult Human Hippocampus

Adult hippocampal neurogenesis was originally discovered in rodents. Subsequent studies identified the adult neural stem cells and found important links between adult neurogenesis and plasticity, behavior, and disease. However, whether new neurons are produced in the human dentate gyrus (DG) during healthy aging is still debated. We and others readily observe proliferating neural progenitors in the infant hippocampus near immature cells expressing doublecortin (DCX), but the number of such cells decreases in children and few, if any, are present in adults. Recent investigations using dual antigen retrieval find many cells stained by DCX antibodies in adult human DG. This has been interpreted as evidence for high rates of adult neurogenesis, even at older ages. However, most of these DCX-labeled cells have mature morphology. Furthermore, studies in the adult human DG have not found a germinal region containing dividing progenitor cells. In this Dual Perspectives article, we show that dual antigen retrieval is not required for the detection of DCX in multiple human brain regions of infants or adults. We review prior studies and present new data showing that DCX is not uniquely expressed by newly born neurons: DCX is present in adult amygdala, entorhinal and parahippocampal cortex neurons despite being absent in the neighboring DG. Analysis of available RNA-sequencing datasets supports the view that DG neurogenesis is rare or absent in the adult human brain. To resolve the conflicting interpretations in humans, it is necessary to identify and visualize dividing neuronal precursors or develop new methods to evaluate the age of a neuron at the single-cell level.     

Temporally specific proliferation events are induced in the hippocampus following acute focal injury

In models of global brain injury, such as stroke or epilepsy, a large increase in neurogenesis occurs in the dentate gyrus (DG) days after the damage is induced. In contrast, more focal damage in the DG produces an increase in neurogenesis within 24 hr. To determine how cell proliferation and differentiation differs in the DG after acute injury in the DG, focal electrolytic lesions were made and mitotic activity was assessed at early (1 day) and late (5 day) time points. At the early time point, bromodeoxyuridine (BrdU)-positive cells were diffusely spread throughout the extent of the hippocampus that was ipsilateral to the lesion. No significant increase in the subgranular zone (SGZ) of the DG was observed. When BrdU was administered at the later time point, the number of BrdU+ cells in the SGZ of the DG was significantly increased. This fourfold increase in BrdU+ cells also resulted in a significant increase in neurogenesis, as measured 6 weeks following BrdU administration. This increase in neurogenesis was not observed when BrdU was administered at the early time point. These results indicate that focal injury in the DG activates two temporally specific proliferation events and that enhanced neurogenesis is observed only following a latent period after the lesion is made.     

Long-term gonadal hormone treatment and endogenous neurogenesis in the dentate gyrus of the adult female monkey

Neurogenesis occurs continually throughout life in all mammals and the extent of neurogenesis is influenced by many factors including gonadal hormones. Most research regarding hormones and neurogenesis has been performed on non-primate species. To determine whether gonadal hormones can modulate endogenous neurogenesis in the dentate gyrus (DG) of the hippocampus in non-human primates, ovariectomized (OVX) female rhesus monkeys received continuous, unopposed β-estradiol (OVX-E-Con), cyclic unopposed β-estradiol (OVX-E-Cyc), continuous β-estradiol + cyclic progesterone (OVX-E-Con + P-Cyc), or control (OVX-Veh) treatments. At week 29, all monkeys received BrdU injections for 4 consecutive days, in addition to the ongoing treatment. Twenty days after the last BrdU injection, all animals were sacrificed for tissue collection. In DG of hippocampus, scattered BrdU-ir cells were observed mainly in the subgranular zone (SGZ) and in the granule cell layer and occasionally these BrdU-ir cells in the SGZ formed clusters containing between 2 and 5 cells. In the granule cell layer and SGZ, virtually none of the BrdU-ir cells were either Dcx, a marker of immature neurons, or GFAP positive. However, an occasional BrdU-ir cell was positive for both neuronal marker NeuN or β III-tubulin. Unbiased stereological analysis of BrdU-ir cells within the SGZ and the granule cell layer of DG revealed that among the experimental groups, there was no significant difference in number of BrdU-ir cells within the SGZ and the granule cell layer of the DG: OVX-E-Con (1801 ± 218.7), OVX-E-Cyc (1783 ± 415.6), OVX-E-Con ± P-Cyc (1721 ± 229.6), and OVX-Veh (1263 ± 106.3), but a trend towards increased BrdU-ir cells was observed in all the experimental groups.     

Microglia engulf viable newborn cells in the epileptic dentate gyrus

Microglia, which are the brain's resident immune cells, engulf dead neural progenitor cells during adult neurogenesis in the subgranular zone (SGZ) of the dentate gyrus (DG). The number of newborn cells in the SGZ increases significantly after status epilepticus (SE), but whether and how microglia regulate the number of newborn cells after SE remain unclear. Here, we show that microglia rapidly eliminate newborn cells after SE by primary phagocytosis, a process by which viable cells are engulfed, thereby regulating the number of newborn cells that are incorporated into the DG. The number of newborn cells in the DG was increased at 5 days after SE in the adult mouse brain but rapidly decreased to the control levels within a week. During this period, microglia in the DG were highly active and engulfed newborn cells. We found that the majority of engulfed newborn cells were caspase‐negative viable cells. Finally, inactivation of microglia with minocycline maintained the increase in the number of newborn cells after SE. Furthermore, minocycline treatment after SE induced the emergence of hilar ectopic granule cells. Thus, our findings suggest that microglia may contribute to homeostasis of the dentate neurogenic niche by eliminating excess newborn cells after SE via primary phagocytosis. GLIA 2016;64:1508–1517     

Functional dentate gyrus neurogenesis in a rapid kindling seizure model

Neurogenesis in the adult mammalian hippocampus resulting in long-term persistence of new neurons with features of capacity for functional activation is recognized. Many stimuli are capable of increasing the rate of neurogenesis, including seizure activity. Whether these insults result in an increased number of new functionally active neurons over and above the baseline rate of neurogenesis is not known. The rapid electrical amygdala kindling (REAK) model of seizures isolates the effects of seizures alone in the absence of neuronal death and the resulting seizures induce expression of c-Fos in the vast majority of dentate gyrus (DG) granule cells. C57BL/6 mice were exposed to REAK then injected with bromodeoxyuridine (BrDU) to label dividing cells, then re-exposed to REAK after a delay period to allow detection of functional activation in new neurons by measurement c-Fos expression in response to seizures. Adult subgranular zone cells migrated into the DG granule cell layer (GCL), assumed a neuronal phenotype and demonstrated seizure-dependent responsiveness. Larger absolute numbers of new neurons demonstrating seizure-dependent activation were found in the GCL of previously kindled mice. Seizures are capable of increasing the number of new neurons with the capacity for functional activation laid down in the postseizure period and incorporated into seizure-activated circuitry.     

Seizure severity‐dependent selective vulnerability of the granule cell layer and aberrant neurogenesis in the rat hippocampus

The pilocarpine‐induced status epilepticus rodent model has been commonly used to analyze the mechanisms of human temporal lobe epilepsy. Recent studies using this model have demonstrated that epileptic seizures lead to increased adult neurogenesis of the dentate granule cells, and cause abnormal cellular organization in dentate neuronal circuits. In this study, we examined these structural changes in rats with seizures of varying severity. In rats with frequent severe seizures, we found a clear loss of Prox1 and NeuN expression in the dentate granule cell layer (GCL), which was confined mainly to the suprapyramidal blade of the GCL at the septal and middle regions of the septotemporal axis of the hippocampus. In the damaged suprapyramidal region, the number of immature neurons in the subgranular zone was markedly reduced. In contrast, in rats with less frequent severe seizures, there was almost no loss of Prox1 and NeuN expression, and the number of immature neurons was increased. In rats with no or slight loss of Prox1 expression in the GCL, ectopic immature neurons were detected in the molecular layer of the suprapyramidal blade in addition to the hilus, and formed chainlike aggregated structures along the blood vessels up to the hippocampal fissure, suggesting that newly generated neurons migrate at least partially along blood vessels to the hippocampal fissure. These results suggest that seizures of different severity cause different effects on GCL damage, neurogenesis, and the migration of new neurons, and that these structural changes are selective to subdivisions of the GCL and the septotemporal axis of the hippocampus.