Vascular adventitia generates neuronal progenitors in the monkey hippocampus after ischemia

In the adult hippocampus, neurogenesis proceeds in the subgranular zone (SGZ) of the dentate gyrus (DG), but not in the cornu Ammonis (CA). Recently, we demonstrated in monkeys that transient brain ischemia induces an increase of the neuronal progenitor cells in the SGZ, but not in CA1, in the second week after the insult. To identify the origin of primary neuronal progenitors in vivo, we compared the postischemic monkey DG and CA1, using light and electron microscopy, focusing on specific phenotype markers, as well as the expression of neurotrophic factors. Laser confocal microscopy showed that 1-3% of 5-bromo-2'-deoxyuridine (BrdU)-positive cells in the SGZ after 2-96 h labeling were also positive for neuronal markers such as TUC4, betaIII tubulin, and NeuN on days 9 and 15. In contrast, despite the presence of numerous BrdU-positive cells, CA1 showed no neurogenesis at any time points, and all the progenitors were positive for glial markers: Iba1 or S-100beta on days 4, 9, and 15. Highly polysialylated neural cell adhesion molecule (PSA-NCAM)-positive cells were abundant in the SGZ, but were absent in CA1. On day 9, most of the immature neurons positive for betaIII-tubulin in SGZ showed an increase in PSA-NCAM immunoreactivity. The immunoreactivity of brain-derived neurotrophic factor (BDNF) was abundant at the vascular adventitia of the SGZ, but was absent at the adventitia of CA1. BrdU-positive progenitor cells were frequently seen in the vicinity of proliferating blood vessels. Ultrastructural analysis indicated that most of the neuronal progenitor cells and microglia originated from the pericytes of capillaries and/or adventitial cells of arterioles (called vascular adventitia). The detaching adventitial cells showed mitotic figures in the perivascular space, and the resultant neuronal progenitor cells made contact with dendritic spines associated with synaptic vesicles or boutons. These data implicate the vascular adventitia as a novel potential source of neuronal progenitor cells in the postischemic primate SGZ.     

Centrally administered interleukin-1 beta sensitizes to the central pressor action of angiotensin II

Proliferating astrocytes and proliferating neuroblasts have been observed in the subgranular zone (SGZ) of the dentate gyrus (DG) in the hippocampus of adult rats under normal conditions. However, whether these proliferating cells are stimulated by running has not been determined. Using immunohistochemical techniques, we examined the effects of chronic treadmill running on proliferating astrocytes (PCNA+/GFAP+ cells), proliferating neuroblasts (PCNA+/DCX+ cells) and newly generated postmitotic neurons (DCX+/NeuN+ cells) in the DG of the hippocampus of adult rats and also characterized the morphological features of PCNA+/GFAP+ cells and PCNA+/DCX+ cells. PCNA+/GFAP+ cells with few processes and PCNA+/DCX+ cells without long processes were detected in the SGZ, and we determined that these are morphological features of the astrocytes and neuroblasts with proliferative ability. Chronic treadmill running (at a speed of 22 m/min, 30 min/days for 7 days) significantly increased the numbers of PCNA+/GFAP+ cells and DCX+/NeuN+ cells, and the number of PCNA+/DCX+ cells tended to increase by chronic treadmill running. These results indicate that chronic treadmill running stimulates the proliferation of astrocytes in the SGZ. Furthermore, the present study indicates that chronic treadmill running increases DCX+/NeuN+ cells that are detected in a transient stage during the neuronal maturation process. These events may be the cellular basis mediating both running-induced increases of new neurons in the DG of the hippocampus and running-induced improvement of learning and memory functions of adult rats.     

Cux2 activity defines a subpopulation of perinatal neurogenic progenitors in the hippocampus

The hippocampus arises from the medial region of the subventricular (SVZ) within the telencephalon. It is one of two regions in the postnatal brain that harbors neural progenitors (NPs) capable of giving rise to new neurons. Neurogenesis in the hippocampus is restricted to the subgranular zone (SGZ) of the dentate gyrus (DG) where it contributes to the generation of granule cell layer (gcl) neurons. It is thought that SGZ progenitors are heterogeneous, differing in their morphology, expression profiles, and developmental potential, however it is currently unknown whether they display differences in their developmental origins and cell fate‐restriction in the DG. Here we demonstrate that Cux2 is a marker for SGZ progenitors and nascent granule cell neurons in the perinatal brain. Cux2 was expressed in the presumptive hippocampal forming region of the embryonic forebrain from E14.5 onwards. At fetal stages, Cux2 was expressed in early‐forming Prox1⁺ granule cell neurons as well as the SVZ of the DG germinal matrix. In the postnatal brain, Cux2 was expressed in several types of progenitors in the SGZ of the DG, including Nestin/Sox2 double‐positive radial glia, Sox2⁺ cells that lacked a radial glial process, DCX⁺ neuroblasts, and Calretinin‐expressing nascent neurons. Another domain characterized by a low level of Cux2 expression emerged in Calbindin⁺ neurons of the developing DG blades. We used Cux2‐Cre mice in genetic fate‐mapping studies and showed almost exclusive labeling of Calbindin‐positive gcl neurons, but not in any progenitor cell types or astroglia. This suggests that Cux2⁺ progenitors directly differentiate into gcl neurons and do not self‐renew. Interestingly, developmental profiling of cell fate revealed an outside‐in formation of gcl neurons in the DG, likely reflecting the activity of Cux2 in the germinative matrices during DG formation and maturation. However, DG morphogenesis proceeded largely normally in hypomorphic Cux2 mutants lacking Cux2 expression. Taken together we conclude that Cux2 expression reflects hippocampal neurogenesis and identifies non‐self‐renewing NPs in the SGZ. © 2014 The Authors Hippocampus Published by Wiley Periodicals, Inc.     

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.     

Transient elevation of adult hippocampal neurogenesis after dopamine depletion

Degeneration of the midbrain dopaminergic neurons during Parkinson's disease (PD) may affect remote regions of the brain that are innervated by the projections of these neurons. The dentate gyrus (DG), a site of continuous production of new neurons in the adult hippocampus, receives dopaminergic inputs from the neurons of the substantia nigra (SN). Thus, depletion of the SN neurons during disease or in experimental settings may directly affect adult hippocampal neurogenesis. We show that experimental ablation of dopaminergic neurons in the 1-methyl-4-phenyl-1,2,3,6-tetrahydopyridine (MPTP) mouse model of PD results in a transient increase in cell division in the subgranular zone (SGZ) of the DG. This increase is evident for the amplifying neural progenitors and for their postmitotic progeny; our results also indicate that MPTP treatment affects division of the normally quiescent stem cells in the SGZ. We also show that l-DOPA, used in the clinical treatment of PD, while attenuating the MPTP-induced death of dopaminergic neurons, does not alter the effect of MPTP on cell division in the DG. Our results suggest that a decrease in dopaminergic signaling in the hippocampus leads to a transient activation of stem and progenitor cells in the DG.     

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.     

有氧训练对阿尔茨海默病模型大鼠海马神经细胞凋亡及再生的影响

目的:观察有氧训练对Aβ_25-35诱导的阿尔茨海默病（AD）大鼠海马神经细胞凋亡及再生的影响。方法:SD大鼠48只,随机分为3组（均n=16）:①假手术组（于大鼠侧脑室注射等量生理盐水+4周有氧训练）;②AD+有氧训练组(于大鼠侧脑室注射Aβ_25-35制作AD大鼠模型+4周有氧训练);③AD组(仅于大鼠侧脑室注射Aβ_25-35制作AD模型)。AD造模后第3天开始进行连续4周的无负重游泳训练,训练结束后各组大鼠分别行Morris水迷宫行为学检测认知行为能力、Hoechst染色观察海马神经细胞凋亡、BrdU/NeuN荧光双染观察齿状回（DG）区新生神经细胞分化及成熟。结果:①Morris水迷宫实验显示,与对照组相比,AD组逃避潜伏期显著延长（P<0.05）;AD+有氧训练组逃避潜伏期较AD组显著缩短（P<0.05）。②与对照组比,AD组齿状回区Hoechst阳性细胞显著增多（P<0.05）;AD+有氧训练组Hoechst阳性细胞较AD组显著减少（P<0.05）。③与对照组比,AD组齿状回Brdu、Brdu/NeuN双染阳性细胞明显减少（P<0.05）;AD+有氧训练组较AD组相比齿状回Brdu、Brdu/NeuN双染阳性细胞明显增多（P<0.05）。结论:有氧训练后,AD大鼠海马亚颗粒细胞区神经细胞的凋亡减少,齿状回神经再生增加,上述改变可能为有氧训练改善AD大鼠认知行为能力的机制之一。     

Cellular localization of epidermal‐type and brain‐type fatty acid‐binding proteins in adult hippocampus and their response to cerebral ischemia

This study aimed at an analysis of expression of epidermal‐type and brain‐type fatty acid‐binding proteins (E‐FABP and B‐FABP, also called FABP5 and FABP7, respectively) in adult hippocampus and their potential value as neuroprotective factors after ischemic brain damage in monkey model. The immunostaining and Western blotting results show that FABP5 was mainly expressed in neurons, whereas FABP7 was primarily expressed in astrocytes and progenitors of the subgranular zone (SGZ). Interestingly, FABP5 expression in neurons increased in cornu Ammonis 1 (CA1) and remains stable within dentate gyrus (DG) after ischemia; FABP7 expression increased within both CA1 and SGZ. This indicates a potential role for FABP5 and FABP7 in intracellular fatty acid transport within different neural cells. The change in FABP5–7 expression within CA1 and DG of the adult postischemic hippocampus was compatible with previous findings of downregulation in CA1 neurons and upregulation in SGZ progenitor cells after ischemia. Altogether, the present data suggest that polyunsaturated fatty acids, such as docosahexaenoic acid, may act via FABP5 or 7 to regulate adult postischemic hippocampal neuronal antiapoptosis or neurogenesis in primates. © 2009 Wiley‐Liss, Inc.     

蛋白偶联受体GPR40在缺血性成年猴海马的表达

目的 利用猴短暂性全脑缺血模型,研究蛋白偶联受体40(GPR40)在海马的表达,从而探讨GPR40受体在缺血性卒中的作用.方法 参照Yamashima建立成年猴短暂性全脑缺血模型,24只猴随机数字表法分成4组,即对照组、缺血后4 d组(d4)、缺血后9 d组(d9)和缺血后15 d(d15),采用Western印迹和免疫荧光在蛋白质水平观察GPR40的表达.结果 Western blotting结果显示CA1区域GPR40在短暂性全脑缺血后表达逐渐下降(15 d内);而在DG区域表达升高,与对照组比较有统计学意义(P<0.05).免疫荧光分析显示CA1区域与GPR40共表达的神经元细胞数逐渐下降.到d15约下降50%;而DG区域则无明显变化,邻近SGZ区域与GPR40共表达的星形细胞数量迅速升高,d4升高约2.5倍.结论 脑缺血后海马CA1和DG两区域GPR40表达差异.特别SGZ星形细胞GPR40表达升高,提示可通过增加GPR40配体中长链不饱和脂肪酸如DHA来改善脑缺血后海马神经元的损伤.     

Impairment of dentate gyrus neuronal progenitor cell differentiation in a mouse model of temporal lobe epilepsy

Unilateral intrahippocampal injection of kainic acid (KA) in adult mice induces an epileptic focus replicating major histopathological features of temporal lobe epilepsy (TLE). In this model, neurogenesis is impaired in the lesioned dentate gyrus, although cell proliferation transiently is increased bilaterally in the subgranular zone (SGZ). To investigate further the relationship between epileptogenesis and neurogenesis, we compared the differentiation of cells born shortly before and after KA injection. Immunohistochemical staining for doublecortin and PSA-NCAM, two markers of young neurons, revealed a rapid downregulation of both markers ipsilaterally, whereas they were increased transiently on the contralateral side. To determine whether KA treatment directly affects neural progenitors in the SGZ, dividing cells were prelabeled with 5'-bromo-2'deoxyuridine (BrdU) treatment before unilateral injection of KA. Double staining with the proliferation marker PCNA showed that prelabeled BrdU cells survived KA exposure and proliferated bilaterally. Unexpectedly, the neuronal differentiation of these cells, as assessed after 2 weeks with doublecortin and NeuN triple-staining, occurred to the same extent as on the contralateral side. Only 5% of pre-labeled BrdU cells were GFAP-positive within the lesion. Therefore, SGZ progenitor cells committed to a neuronal phenotype before KA treatment complete their differentiation despite the rapid down-regulation of doublecortin and PSA-NCAM. These findings suggest impaired fate commitment and/or early differentiation of proliferating cells in the lesioned dentate gyrus. Loss of neurogenesis in this TLE model likely reflects an irreversible alteration of the SGZ germinal niche during development of the epileptic focus and may therefore be relevant for human TLE.     

Loss of true blue labelling from the medial septum following transection of the fimbria-fornix: evidence for the death of cholinergic and non-cholinergic neurons

Many neurons in the medial septal nucleus lose their transmitter-associated enzyme staining following axotomy in the proximal fimbria-fornix (FF), but it is not clear if these neurons have died or persist in a shrunken and subfunctional state. To investigate this further, septal neurons projecting through the FF were labelled with the fluorescent dye, True blue, by retrograde transport from multiple bilateral injection sites in the hippocampus. True blue-labelled neurons and cholinergic neurons immunohistochemically stained for choline acetyltransferase (ChAT) were then quantitatively compared in neighbouring sections through the medial septum 28 days after complete unilateral transections of the proximal FF. The number of True blue and ChAT positive cells ipsilateral to the FF lesion showed significant (P less than 0.001) declines of 51.4% and 71.1%, respectively, relative to the unlesioned side. Cell loss was considerably more severe among large neurons, such that 78.0% and 92.7% of True blue and ChAT labelled cells larger than the normal mean, but only 40.1% and 68.0% of True blue and ChAT labelled cells smaller than the normal mean size were lost. This indicates either that larger neurons were more prone to cell loss, or that some (but not all) large neurons persisted in a shrunken form. Histograms showed no increase in cell number in any of the smaller size categories and a substantial decrease in most cases, indicating that shrinkage alone could not account for the loss of all large neurons. Since True blue can remain present in brainstem cholinergic neurons surviving for over 365 days after axotomy, loss of True blue suggests breakdown of membrane integrity and cell death.(ABSTRACT TRUNCATED AT 250 WORDS)     

Immunohistochemical changes of neuronal calcium-binding proteins parvalbumin and calbindin-D-28k following unilateral deafferentation in the rat visual system.

The neuron-specific calcium-binding proteins, parvalbumin and calbindin-D-28k, were studied in the subcortical visual system of normal and unilaterally deafferented albino rats. Immunohistochemistry with monoclonal antibodies was used on vibratome sections through optic tract (OT), dorsal lateral geniculate nucleus (dLGN), olivary pretectal nucleus (OPN), and superior colliculus (SC). In controls, OT stained strongly for parvalbumin and weakly for calbindin-D-28k. The dLGN contained a plexus of parvalbumin-positive fibers. In dLGN, calbindin-D-28k-antibodies showed strong labeling of some neurons with long dendrites and weak staining of the cytoplasm in other neurons. In OPN, parvalbumin stained a ring of neurons and terminals in the shell region, whereas calbindin-D-28k was contained in medial cell populations. In SC, parvalbumin was contained in fibers, terminals, and neurons throughout the visual layer. Calbindin-D-28k showed a laminar distribution of neurons with a predominance in deep portions of superficial grey matter and in ventral portions of stratum opticum. Following unilateral deafferentation induced by optic nerve section, retinal axons showed immunohistochemical changes related to Wallerian degeneration and target neurons reacted by changes of calcium-binding proteins. Parvalbumin and calbindin-D-28k immunostaining decreased during Wallerian degeneration of OT. In the deafferented dLGN, immunohistochemical labeling for calbindin-D-28k declined in strongly stained neurons from 4 to 21 days after lesion. Measurement of dendritic length per number of cells or per area of dLGN showed a significant decline for the contralateral side at 4, 8, and 21 days (ANOVA, P less than 0.05). In deafferented OPN, terminal-like staining for parvalbumin decreased and neuronal labeling was enhanced. In deafferented SC, the neuronal and dendritic staining for parvalbumin increased beginning from Day 1 on and persisting at Day 21, whereas fibers and terminal-like elements decreased in staining. Measurement of parvalbumin-positive neurons per area of SC showed a significant increase of labeling in the contralateral side from Day 1 to Day 21 (ANOVA, P less than 0.05). These studies show that cellular responses to deafferentation of visual neurons involve a regulation of calcium-binding proteins. The decline in staining for calbindin-D-28k in dLGN may relate to reduced retinal afferent activity. The progressive cellular changes in parvalbumin staining may be related to unmasking of intrinsic neurons after removal of parvalbumin-containing, afferent fibers and terminals. Additionally, the changes of parvalbumin labeling in SC neurons may reflect a plastic reorganization of local circuits known to occur in rat SC in response to deafferentation.     

Expression of free fatty acid receptor GPR40 in the neurogenic niche of adult monkey hippocampus

Polyunsaturated free fatty acids (PUFAs) are known to play critical roles for the development, maintenance, and function of the brain. Recently, we reported that G-protein coupled receptor 40 (GPR40), one type of PUFA receptors, is expressed throughout the adult primate central nervous system including the hippocampus. This opens a possibility that PUFA might act as extracellular signaling molecules at the GPR40 receptor to regulate neuronal function. Here we studied protein expression of GPR40 in the neurogenic niche of the adult monkey hippocampus under normal and postischemic conditions. Confocal laser microscope analysis of immunostained sections revealed GPR40 immunoreactivity in neural progenitors, immature neurons, astrocytes and endothelial cells of the subgranular zone (SGZ) of the dentate gyrus (DG); a well-known neurogenic niche within the adult brain. Immunoblotting analysis showed that the GPR40 protein increased significantly in the second week after global cerebral ischemia as compared with the control. This was compatible with the postischemic increment of GPR40-positive cells in the SGZ as detected by immunofluorescence imaging. Taken together with our previous findings of the SGZ progenitor cell upregulation after ischemia, the present data suggest that PUFA such as docosahexaenoic acid may act via GPR40 to regulate adult hippocampal neurogenesis in primates.     

Status epilepticus during old age is not associated with enhanced hippocampal neurogenesis

Increased production of new neurons in the adult dentate gyrus (DG) by neural stem/progenitor cells (NSCs) following acute seizures or status epilepticus (SE) is a well known phenomenon. However, it is unknown whether NSCs in the aged DG have similar ability to upregulate neurogenesis in response to SE. We examined DG neurogenesis after the induction of continuous stages III-V seizures (SE) for over 4 h in both young adult (5-months old) and aged (24-months old) F344 rats. The seizures were induced through 2-4 graded intraperitoneal injections of the excitotoxin kainic acid (KA). Newly born cells in the DG were labeled via daily intraperitoneal injections of the 5'-bromodeoxyuridine (BrdU) for 12 days, which commenced shortly after the induction of SE in KA-treated rats. New cells and neurons in the subgranular zone (SGZ) and the granule cell layer (GCL) were analyzed at 24 h after the last BrdU injection using BrdU and doublecortin (DCX) immunostaining, BrdU-DCX and BrdU-NeuN dual immunofluorescence and confocal microscopy, and stereological cell counting. Status epilepticus enhanced the numbers of newly born cells (BrdU(+) cells) and neurons (DCX(+) neurons) in young adult rats. In contrast, similar seizures in aged rats, though greatly increased the number of newly born cells in the SGZ/GCL, failed to increase neurogenesis due to a greatly declined neuronal fate-choice decision of newly born cells. Only 9% of newly born cells in the SGZ/GCL differentiated into neurons in aged rats that underwent SE, in comparison to the 76% neuronal differentiation observed in age-matched control rats. Moreover, the number of newly born cells that migrate abnormally into the dentate hilus (i.e., ectopic granule cells) after SE in the aged hippocampus is 92% less than that observed in the young adult hippocampus after similar SE. Thus, SE fails to increase the addition of new granule cells to the GCL in the aged DG, despite a considerable upregulation in the production of new cells, and SE during old age leads to much fewer ectopic granule cells. These results have clinical relevance because earlier studies have implied that both increased and abnormal neurogenesis occurring after SE in young animals contributes to chronic epilepsy development.     

Enhanced neurogenesis during trimethyltin-induced neurodegeneration in the hippocampus of the adult rat

The occurrence of neurogenesis in the hippocampus of the adult rat during trimethyltin (TMT)-induced neurodegeneration was investigated using bromodeoxyuridine (BrdU). Fifteen days after TMT intoxication, BrdU-labeled cells were significantly more numerous in the hippocampus of treated animals, gradually decreasing towards the control value 21 days after intoxication in the dentate gyrus (DG), while in the CA3/hilus region BrdU-labeled cells were still more numerous in TMT-treated rats. In order to investigate the fate of newly-generated cells double labeling experiments using neuronal or glial markers were performed. Colocalization of the neuronal marker NeuN was detected in many BrdU-positive cells in the DG, while in the CA3/hilus region no colocalization of NeuN and BrdU could be observed. No colocalization of BrdU and the astroglial marker GFAP or the microglial marker OX-42 was detected either in the DG and or in the CA3/hilus region. The results indicate an enhancement of endogenous neurogenesis in the hippocampus during TMT-induced neurodegeneration, with the development of a subpopulation of regenerated cells into neurons in the DG, while in the CA3/hilus region the population of newly-generated cells should be regarded as undifferentiated.     

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.     

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.     

Temporal profile of neurogenesis in the subventricular zone,dentate gyrus and cerebral cortex following transient focal cerebral ischemia

Abstract

Background: In the adult mammalian brain, it is considered that neurogenesis persists in limited regions such as the hippocampal dentate gyrus (DG) and the subventricular zone (SVZ) of the lateral ventricle. On the other hand, neurogenesis in the cortex after cerebral ischemia and its role in post-stroke recovery have not been clarified yet. In this study, we investigated neurogenesis in the cortex and the spatiotemporal profile of neural progenitors in SVZ and DG of rats subjected to transient focal cerebral ischemia.

Materials and methods: Male Sprague–Dawley rats (270–300 g) were subjected to 60 minute middle cerebral artery occlusion. Proliferating cells were labeled by the cumulative administration of BrdU 1, 2, 3, 4, 6 and 8 weeks after ischemia induction (at weeks 1–4, 6 and 8). Double labeling was also performed with antibodies against BrdU and NeuN.

Results: BrdU-positive cells proliferated in DG and SVZ of the bilateral hemispheres, and their proliferation peaked at week 3 in SVZ and at week 4 in DG. In the peri-infarct zone of cerebral cortex, BrdU-positive cells co-expressed NeuN from weeks 3 to 8.

Conclusion: Neurogenesis was observed in the cerebral cortex and proliferation of neural progenitors occurred in SVZ and DG of rats subjected to transient focal cerebral ischemia. Our data might indicate that endogenous dormant neural stem cells residing in the cortex were activated by ischemic insult to induce the proliferation of neural progenitors and differentiation into mature neurons.     

Three steps of neural stem cells development in gerbil dentate gyrus after transient ischemia.

The stage of neurogenesis can be divided into three steps: proliferation, migration, and differentiation. To elucidate detailed relations between these three steps after ischemia, the authors evaluated the three steps in the adult gerbil dentate gyrus (DG) after 5 minutes of transient global ischemia using bromodeoxyuridine (BrdU), highly polysialylated neural cell adhesion molecule (PSA-NCAM), and neuronal nuclear antigen (NeuN) and glial fibrillary acidic protein (GFAP) as markers for proliferation, migration, and differentiation, respectively. Bromodeoxyuridine-labeled cells increased approximately sevenfold, and PSA-NCAM-positive cells increased approximately threefold in the subgranular zone (SGZ) with a peak 10 days after ischemia. Bromodeoxyuridine-labeled cells with PSA-NCAM expression were first detected both in the SGZ and the granule cell layer (GCL) 20 days after ischemia and gradually decreased after that, whereas BrdU-labeled cells with NeuN gradually increased in the GCL until 60 days after ischemia. A few BrdU-labeled cells with GFAP expression were detected in DG after ischemia; no PSA-NCAM-positive cells with GFAP expression were detected, but the radial processes of glial cells were partly in contact with PSA-NCAM-positive cell bodies and dendrites. These results suggest that neural stem cell proliferation begins at the SGZ, and that the cells then migrate into the GCL and differentiate mainly into neuronal cells. The majority of these three steps finished in 2 months after transient global ischemia.     

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.