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Aims: Previous studies have demonstrated that rapamycin prevents seizure-induced anxiety-like behaviors. However, rapamycin had been used at a higher dose of 3 mg/kg and resulted in side effects in immature animals. This work was designed to explore whether a lower dose of rapamycin has similar efficacy but has milder side effects.Methods: Acute seizures were induced by injection of pilocarpine at postnatal 10-day Sprague-Dawley rats. Western blot analysis was used to detect changes in mammalian target of rapamycin (mTOR) pathway after seizure. Immunofluorescent intensity of doublecortin (DCX) was conducted to evaluate the development of neurons in hippocampus. Morris water maze and Y-maze test were used to assess cognitive functions and open-field test and elevated plus maze were used to detect anxiety-like behaviors 4 weeks after seizure onset.Results: mTOR pathway was abnormally activated with two peaks after pilocarpine-induced seizures, and no difference of DCX-positive cells and body weight were noticed between control and pilocarpine-induced seizure rats. Pilocarpine-induced seizure in postnatal 10 days rats did not exert impairment on cognitive functions, but resulted in obvious anxiety-like behaviors. Low dose of rapamycin at 0.3 mg/kg significantly reversed seizure-induced increase of p-S6 levels as well as abnormal anxiety-like behaviors. In addition, rapamycin at the dose of 0.3mg/kg did not affect normal development and cognitive functions.Conclusion: lower doses of rapamycin should be used in infants compared with older children or adults.

Seizure activity stimulates adult neurogenesis, the birth of new neurons, in the hippocampus. Many new neurons that develop in the presence of repeatedly induced seizures acquire abnormal morphological and functional characteristics that can promote network hyperexcitability and hippocampal dysfunction. However, the impact of seizure induced neurogenesis on behaviour remains poorly understood. In this study, we investigated whether adult-born neurons generated immediately before and during chronic seizures were capable of integration into behaviorally relevant hippocampal networks. Adult rats underwent pentylenetetrazole (PTZ) kindling for either 1 or 2 weeks. Proliferating cells were labelled with BrdU immediately before kindling commenced. Twenty-four hours after receiving their last kindling treatment, rats were placed in a novel environment and allowed to freely explore for 30 min. The rats were euthanized 90 min later to examine for behaviourally-induced immediate early gene expression (c-fos, Zif268). Using this approach, we found that PTZ kindled rats did not differ from control rats in regards to exploratory behaviour, but there was a marked attenuation in behaviour-induced expression of Fos and Zif268 for rats that received 2 weeks of PTZ kindling. Further examination revealed that PTZ kindled rats showed reduced colocalization of Fos and Zif268 in 2.5 week old BrdU + cells. The proportion of immature granule cells (doublecortin-positive) expressing behaviorally induced Zif268 was also significantly lower for PTZ kindled rats than control rats. These results suggest that chronic seizures can potentially disrupt the ability of adult-born cells to functionally integrate into hippocampal circuits important for the processing of spatial information.

The brain has long been considered a site of "immune privilege"; however, recent evidence indicates the presence of brain-immune interactions in physiological and pathological conditions. Neurogenesis, a process of generating functionally integrated neurons, occurs in the adult brain of mammals. The adult neurogenesis predominantly takes place in the subgranular zone (SGZ) of the hippocampal dentate gyrus and the subventricular zone (SVZ). Several studies have shown that an immune reaction or alteration could affect adult neurogenesis activity, suggesting a link between the immune system and adult neurogenesis. Helminth infection is one of the activators of Th2 immune response. However, the influence of this type of immune reaction on adult neurogenesis is not well studied. In this study, we evaluated adult neurogenesis in mice infected with the helminth Nippostrongylus brasiliensis (Nb). Immunohistochemically, the number of both doublecortin-positive cells and doublecortin/5-bromodeoxyuridine (BrdU)-double-positive cells was decreased in the SGZ of Nb-infected mice by day 9 after infection. However, the total number of BrdU-positive newborn cells in the SGZ did not change. In no significant alterations were detected in the SVZ of infected mice. In addition, using reverse transcription-quantitative polymerase chain reaction, we observed no significant changes in the expression levels of neurotropic factors important for neurogenesis in the hippocampus. In conclusion, our results indicate that adult neurogenesis in SGZ, but not in SVZ, is inhibited by Nb infection. Th2 immune response might have a suppressive effect on hippocampal neurogenesis.

Developmental exposure to valproic acid (VPA), a model compound for experimental autism, has shown to primarily target GABAergic interneuron subpopulations in hippocampal neurogenesis of rat offspring. The VPA-exposed animals had revealed late effects on granule cell lineages, involving progenitor cell proliferation and synaptic plasticity. To investigate the possibility whether hippocampal neurogenesis in postpubertal rats in a protocol of 28-day repeated exposure is affected in relation with the property of a developmental neurotoxicant by developmental exposure, VPA was orally administered to 5-week-old male rats at 0, 200, 800 and 900 mg/kg body weight/day for 28 days. At 900 mg/kg, GFAP+ cells increased in number, but DCX+ cells decreased in number in the granule cell lineages. Moreover, CHRNB2+ cells and NeuN+ postmitotic neurons decreased in number in the hilus of the dentate gyrus. Transcript level examined at 900 mg/kg in the dentate gyrus was increased with Kit, but decreased with Dpsyl3, Btg2, Pvalb and Chrnb2. These results suggest that VPA increased type-1 stem cells in relation to the activation of SCF-KIT signaling and suppression of BTG2-mediated antiproliferative effect on stem cells. VPA also decreased type-3 progenitor cells and immature granule cells probably in relation with PVALB+ interneuron hypofunction and reduced CHRNB2+ interneuron subpopulation in the hilus, as well as with suppression of BTG2-mediated terminal differentiation of progenitor cells. Thus, the disruption pattern of VPA by postpubertal exposure was different from developmental exposure. However, disruption itself can be detected, suggesting availability of hippocampal neurogenesis in detecting developmental neurotoxicants in a 28-day toxicity study.

In the adult hippocampal dentate gyrus (DG), the majority of newly generated cells are eliminated by apoptotic mechanisms. The apoptosis repressor with caspase recruitment domain (ARC), encoded by the Nol3 gene, is a potent and multifunctional death repressor that inhibits both death receptor and mitochondrial apoptotic signaling. The aim of the present study was to parse the role of ARC in the development of new granule cell neurons. Nol3 gene expression as revealed by in situ hybridization is present in the entire dentate granule cell layer. Moreover, a comparison of Nol3 expression between FACS-sorted Sox2-positive neural stem cells and Doublecortin (DCX)-positive immature neurons demonstrates upregulation of Nol3 during neurogenesis. Using ARC-deficient mice, we show that proliferation and survival of BrdU birth-dated cells are strongly reduced in the absence of ARC while neuronal-glial fate choice is not affected. Both the number of DCX-positive cells and the number of calretinin (CR)-positive immature postmitotic neurons are reduced in the hippocampus of ARC-/- mice. ARC knockout is not associated with increased numbers of microglia or with microglia activation. However, hippocampal brain-derived neurotrophic factor (BDNF) protein content is significantly increased in ARC-/- mice, possibly representing a compensatory response. Collectively, our results suggest that ARC plays a critical cell-autonomous role in preventing cell death during adult granule cell neogenesis.

To develop biological approaches to restore vision, we developed a method of transplanting stem cell-derived retinal tissue into the subretinal space of a large-eye animal model (cat). Human embryonic stem cells (hESC) were differentiated to retinal organoids in a dish. hESC-derived retinal tissue was introduced into the subretinal space of wild-type cats following a pars plana vitrectomy. The cats were systemically immunosuppressed with either prednisolone or prednisolone plus cyclosporine A. The eyes were examined by fundoscopy and spectral-domain optical coherence tomography imaging for adverse effects due to the presence of the subretinal grafts. Immunohistochemistry was done with antibodies to retinal and human markers to delineate graft survival, differentiation, and integration into cat retina. We successfully delivered hESC-derived retinal tissue into the subretinal space of the cat eye. We observed strong infiltration of immune cells in the graft and surrounding tissue in the cats treated with prednisolone. In contrast, we showed better survival and low immune response to the graft in cats treated with prednisolone plus cyclosporine A. Immunohistochemistry with antibodies (STEM121, CALB2, DCX, and SMI-312) revealed large number of graft-derived fibers connecting the graft and the host. We also show presence of human-specific synaptophysin puncta in the cat retina. This work demonstrates feasibility of engrafting hESC-derived retinal tissue into the subretinal space of large-eye animal models. Transplanting retinal tissue in degenerating cat retina will enable rapid development of preclinical in vivo work focused on vision restoration.

Background Over 50% of acute stroke patients have hyperglycemia, which is associated with a poorer prognosis and outcome. Our aim was to investigate the impact of hyperglycemia on behavioral recovery and brain repair of delivered human adipose tissue-derived mesenchymal stem cells (hAD-MSCs) in a rat model of permanent middle cerebral artery occlusion (pMCAO). Methods Hyperglycemia was induced in rats by the administration of nicotinamide and streptozotocin. The rats were then subjected to stroke by a pMCAO model. At 48 h post-stroke, 1 × 106 hAD-MSCs or saline were intravenously administered. We evaluated behavioral outcome, infarct size by MRI, and brain plasticity markers by immunohistochemistry (glial fibrillary acidic protein [GFAP], Iba-1, synaptophysin, doublecortin, CD-31, collagen-IV, and α-smooth muscle actin [α-SMA]). Results The hyperglycemic group exhibited more severe neurological deficits; lesion size and diffusion coefficient were larger compared with the non-hyperglycemic rats. GFAP, Iba-1, and α-SMA were increased in the hyperglycemic group. The hyperglycemic rats administered hAD-MSCs at 48 h after pMCAO had improved neurological impairment. Although T2-MRI did not show differences in lesion size between groups, the rADC values were lower in the treated group. Finally, the levels of GFAP, Iba-1, and arterial wall thickness were lower in the treated hyperglycemic group than in the nontreated hyperglycemic group at 6 weeks post-stroke. Conclusions Our data suggest that rats with hyperglycemic ischemic stroke exhibit increased lesion size and impaired brain repair processes, which lead to impairments in behavioral recovery after pMCAO. More importantly, hAD-MSC administration induced better anatomical tissue preservation, associated with a good behavioral outcome. Electronic supplementary material The online version of this article (10.1186/s13287-019-1322-x) contains supplementary material, which is available to authorized users.

Macroautophagy/autophagy is generally regarded as a cytoprotective mechanism, and it remains a matter of controversy whether autophagy can cause cell death in mammals. Here, we show that chronic restraint stress suppresses adult hippocampal neurogenesis in mice by inducing autophagic cell death (ACD) of hippocampal neural stem cells (NSCs). We generated NSC-specific, inducible Atg7 conditional knockout mice and found that they had an intact number of NSCs and neurogenesis level under chronic restraint stress and were resilient to stress- or corticosterone-induced cognitive and mood deficits. Corticosterone treatment of adult hippocampal NSC cultures induced ACD via SGK3 (serum/glucocorticoid regulated kinase 3) without signs of apoptosis. Our results demonstrate that ACD is biologically important in a mammalian system in vivo and would be an attractive target for therapeutic intervention for psychological stress-induced disorders. Abbreviations: AAV: adeno-associated virus; ACD: autophagic cell death; ACTB: actin, beta; Atg: autophagy-related; ASCL1/MASH1: achaete-scute family bHLH transcription factor 1; BafA1: bafilomycin A1; BrdU: Bromodeoxyuridine/5-bromo-2ʹ-deoxyuridine; CASP3: caspase 3; cKO: conditional knockout; CLEM: correlative light and electron microscopy; CORT: corticosterone; CRS: chronic restraint stress; DAB: 3,3ʹ–diaminobenzidine; DCX: doublecortin; DG: dentate gyrus; GC: glucocorticoid; GFAP: glial fibrillary acidic protein; HCN: hippocampal neural stem; i.p.: intraperitoneal; MAP1LC3B: microtubule-associated protein 1 light chain 3 beta; MKI67/Ki67: antigen identified by monoclonal antibody Ki 67; MWM: Morris water maze; Nec-1: necrostatin-1; NES: nestin; NR3C1/GR: nuclear receptor subfamily 3, group C, member 1; NSC: neural stem cell; PCD: programmed cell death; PFA: paraformaldehyde; PX: Phox homology; PtdIns3P: phosphatidylinositol-3-phosphate; RBFOX3/NeuN: RNA binding protein, fox-1 homolog (C. elegans) 3; SGK: serum/glucocorticoid-regulated kinases; SGZ: subgranular zone; SOX2: SRY (sex determining region Y)-box 2; SQSTM1: sequestosome 1; STS: staurosporine; TAM: tamoxifen; Ulk1: unc-51 like kinase 1; TUNEL: terminal deoxynucleotidyl transferase dUTP nick end labeling; VIM: vimentin; WT: wild type; ZFYVE1: zinc finger, FYVE domain containing 1; Z-VAD/Z-VAD-FMK: pan-caspase inhibitor

The orphan nuclear receptor TLX (Nr2e1) is a key regulator of hippocampal neurogenesis. Impaired adult hippocampal neurogenesis has been reported in neurodegenerative and psychiatric conditions including dementia and stress-related depression. Neuroinflammation is also implicated in the neuropathology of these disorders, and has been shown to negatively affect hippocampal neurogenesis. To investigate a role for TLX in hippocampal neuroinflammation, we assessed microglial activation in the hippocampus of mice with a spontaneous deletion of TLX. Results from our study suggest that a lack of TLX is implicated in deregulation of microglial phenotype and that consequently, the survival and function of newborn cells in the hippocampus is impaired. TLX may be an important target in understanding inflammatory-associated impairments in neurogenesis.

Neurogenesis in the substantia nigra (SN) has been a controversial issue. Here we report that neurogenesis can be induced in the adult rodent SN by transplantation of embryoid body cells (EBCs) derived from mouse embryonic stem cells. The detection of Sox2+ dividing (BrdU+) putative host neural precursor cells (NPCs) between 1 and 6 days post-transplantation (dpt) supported the neurogenic capacity of the adult SN. In agreement with the awakening of NPCs by EBCs, only host cells from implant-bearing SN were able to generate neurosphere-like aggregates in the presence of Egf and Fgf2. Later, at 15 dpt, a significant number of SN Dcx+ neuroblasts were detected. However, a continuous BrdU administration after transplantation showed that only a fraction (about 20–30%) of those host Dcx+ progeny derived from dividing cells and few BrdU+ cells, some of them NeuN+, survived up to 30 dpt. Unexpectedly, 25–30% of Dcx+ or Psa-Ncam+ cells at 15 dpt displayed astrocytic markers such as Gfap and S100b. Using a genetic lineage tracing strategy, we demonstrated that a large proportion of host Dcx+ and/or Tubb3+ neuroblasts originated from Gfap+ cells. Remarkably, new blood vessels formed in association with the neurogenic process that, when precluded, caused a reduction in neuroblast production. Accordingly, two proteins secreted by EBCs, Fgf2 and Vegf, were able to promote the emergence of Dcx+/Psa-Ncam+, Tubb3+ and NeuN+/BrdU+ cells in vivo in the absence of EBCs. We propose that the adult SN is a mostly silent neurogenic niche with the ability to generate new neurons by typical and atypical mechanisms.

We combined translating ribosome affinity purification (TRAP) with in utero electroporation (IUE), called iTRAP to identify the molecular profile of specific neuronal populations during neonatal development without the need for viral approaches and FACS sorting. We electroporated a plasmid encoding EGFP-tagged ribosomal protein L10a at embryonic day (E) 14–15 to target layer 2–4 cortical neurons of the somatosensory cortex. At three postnatal (P) ages—P0, P7, and P14—when morphogenesis occurs and synapses are forming, TRAP and molecular profiling was performed from electroporated regions. We found that ribosome bound (Ribo)-mRNAs from ~7,300 genes were significantly altered over time and included classical neuronal genes known to decrease (e.g., Tbrl, Dcx) or increase (e.g., Eno2, Camk2a, Synl) as neurons mature. This approach led to the identification of specific developmental patterns for Ribo-mRNAs not previously reported to be developmentally regulated in neurons, providing rationale for future examination of their role in selective biological processes. These include upregulation of Lynxl, Nrnl, Cntnapl over time; downregulation of St8sia2 and Draxin; and bidirectional changes to Fkbplb. iTRAP is a versatile approach that allows researchers to easily assess the molecular profile of specific neuronal populations in selective brain regions under various conditions, including overexpression and knockdown of target genes, and in disease settings.

Effective delivery of trophic factors to cure neurological disorders and traumatic injuries is a major challenge. With promising therapeutic effects of epidermal neural crest stem cells (EPI-NCSCs) in preclinical spinal cord injury, there is an implication that these stem cells might provide supportive role through releasing various trophic agents. Hence, the present study was designed to assess the influence of valproic acid (VPA), a well-known histone deacetylases inhibitor, on mRNA expression of selected trophic factors. In this study, following stem cell migration from explanted hair bulges, immunostaining against Nestin, SOX-10, DCX, β-III tubulin and GFAP was carried out. Then, cells were treated with various clinically relevant concentrations of VPA and the survival rate was defined by MTT assay. Finally, stem cells were treated with 0.1 and 1 mM VPA and the drug impact on the transcription level of BDNF, GDNF, VEGF, NGF and NT3 at 6, 24, 72, 168 h was assessed by quantitative real-time PCR. The examined proteins expressions in the population of migrated cells confirmed the identity of stem cells as EPI-NCSCs. In addition, MTT assay showed that all three tested concentrations of VPA were suitable to treat these cells. Trophic factors assessment, following treatment revealed the mRNA expression level of BDNF, GDNF and VEGF could be significantly up- regulated at various time points, mainly by 1 mM VPA. However, NGF and NT3 transcripts were enhanced at few limited time points. Our findings showed that EPI-NCSCs due to secretion of various trophic factors are potential candidate to deliver the required trophic agents and their potential can be enhanced by 1 mM VPA, predominantly following 168 h treatment. Hence, these cells can be utilized to modulate destructive context of neurological disorders and injuries.

Subcortical band heterotopia (SBH), also known as doublecortex syndrome, is a malformation of cortical development resulting from mutations in the doublecortin gene (DCX). It is characterized by a lack of migration of cortical neurons that accumulate in the white matter forming a heterotopic band. Patients with SBH may present mild to moderate intellectual disability as well as epilepsy. The SBH condition can be modeled in rats by in utero knockdown (KD) of Dcx. The affected cells form an SBH reminiscent of that observed in human patients and the animals develop a chronic epileptic condition in adulthood. Here, we investigated if the presence of a SBH is sufficient to induce cognitive impairment in juvenile Dcx-KD rats, before the onset of epilepsy. Using a wide range of behavioral tests, we found that the presence of SBH did not appear to affect motor control or somatosensory processing. In addition, cognitive abilities such as learning, short-term and long-term memory, were normal in pre-epileptic Dcx-KD rats. We suggest that the SBH presence is not sufficient to impair these behavioral functions.

Depression is a major health issue that causes severe societal economic and health burden. Aromatherapy, a practice that uses essential oils for preventive and therapeutic purposes, represents a promising therapeutic alternative for the alleviation of depressive symptoms. Lavender essential oil (LEO) has been the focus of clinical studies due to its positive effect on mood. An animal model of chronic administration of high dose corticosterone to induce depression- and anxiety-like behavior and reduced neurogenesis was used to explore the biological changes brought by aromatherapy. Twenty-four adult male Sprague Dawley rats were randomly assigned into four groups: Control, corticosterone (Cort) group with high dose of corticosterone, LEO group with daily exposure to LEO by inhalation, and LEO + Cort. At the end of the 14-day treatment period, behavioral tests were carried out. Serum samples were collected 2–3 days after the 14-day period treatment and before perfusion to carry out biochemical analyses to measure BDNF, corticosterone and oxytocin. After perfusion, brains were collected for immunohistochemical analysis to detect BrdU and DCX positive cells in the hippocampus and subventricular zone. Results showed that treatment with LEO ameliorated the depression-like behavior induced by the chronic administration of corticosterone as observed in the LEO + Cort group. Cort treatment reduced the number of BrdU positive cells in the hippocampus and the subventricular zone. Treatment with LEO prevented the corticosterone-induced reduction in the number of BrdU positive cells (LEO + Cort group) demonstrating the neurogenic effect of LEO under high corticosterone conditions. Chronic administration of high dose of corticosterone significantly reduced the dendritic complexity of immature neurons. On the contrary, treatment with LEO increased dendritic complexity of immature neurons under high corticosterone conditions (LEO + Cort group). The improved neurogenesis and dendritic complexity observed in the LEO + Cort group demonstrated a clear restorative effect of LEO under high corticosterone conditions. However, 2–3 days after the treatment, the levels of BDNF were upregulated in the LEO and LEO + Cort groups. Furthermore, the concentration of oxytocin in serum, 2–3 days after the treatment, showed to be upregulated in the LEO group alone. The present study has provided evidence of the biological effect of LEO on neuroplasticity and neurogenesis. Also, this study contributes to the understanding of the mechanism of action of LEO in an animal model where depression- and anxiety-like behavior and reduced neurogenesis were induced by high corticosterone administration.

Doublecortin X (DCX) plays essential roles in neuronal development via its regulation of cytoskeleton dynamics. This is mediated through direct interactions between its doublecortin (DC) domains (DC1 and DC2) with microtubules (MTs) and indirect association with actin filaments (F-ACT). While the regulatory role of the DCX C-terminus following DC2 (i.e. DCX residues 275-366) has been established, less is known of the possible contributions made by the DCX N-terminus preceding DC1 (i.e. DCX residues 1-44). Here, we assessed the influence of DCX Ser28 within the DCX N-terminus, on the association of DCX with MTs and F-ACT. We compared the cytoskeletal interactions of the DCX S28E phosphomimetic and DCX S28A phospho-resistant mutants and wild-type DCX. Immunoprecipitation and colocalisation analyses indicated increased association of DCX S28E with F-ACT but decreased interaction with MTs, and conversely enhanced DCX S28A association with MTs but decreased association with F-ACT. To evaluate the impact of DCX mutants on cytoskeletal filaments we performed fluorescence recovery after photobleaching (FRAP) studies on SiR-tubulin and β-actin-mCherry and observed comparable tubulin and actin exchange rates in the presence of DCX WT and DCX S28A. However, we observed faster tubulin exchange rates but slower actin exchange rates in the presence of DCX S28E. Moreover, DCX S28E enhanced the association with the actin-binding protein spinophilin (Spn) suggesting the shift to favour association with both F-ACT and Spn in the presence of DCX S28E. Taken together, our results highlight a new role for DCX S28 as a regulatory switch for cytoskeletal organisation.

BDNF and proBDNF play an opposite role in hippocampal neurogenesis. What remains to be known is the effect of balance between BDNF and proBDNF in the ischemic hippocampus on pathogenesis of post-stroke depression (PSD) and the potential mechanisms of aerobic exercise (AE) on PSD. Wistar rats were randomly divided into control, Sham, Sedentary and AE groups. After PSD model was successful made, the blood lactate threshold corresponding speed (SLT) were measured. The behavioral tests (open field, forced swimming and sucrose preference tests) were performed before and after 4 weeks of aerobic treadmill training. HE staining and immunostaining for doublecortin (DCX)+ neurons were used to observe the changes of neuronal cell morphology and proliferation, migration of the neural progenitor cells (NPCs). The expression of mature brain-derived neurotrophic factor (BDNF), tropomyosin receptor kinase B (TrkB), precursor BDNF (proBDNF), pan-neurotrophin receptor 75 (p75NTR) proteins, BDNF mRNA in the ischemic hippocampus and serum adrenocorticotropic hormone (ACTH) and corticosterone (CORT) were detected by Western blotting, immunohistochemistry, RT-PCR and ELISA. Higher immobility time and levels of proBDNF, p75NTR, ACTH, CORT proteins and lower sucrose preference, total distance, climbing frequency and levels of BDNF, TrkB proteins, BDNF mRNA were observed in the Sedentary group. Neuronal cells in the ischemic hippocampus were loosely arranged and expression of DCX reduced in the Sedentary group. There were significant differences in above results between Sedentary and AE groups after 4 weeks of aerobic exercise. The balance between BDNF and proBDNF in the ischemic hippocampus are likely to play an important role in the pathogenesis of PSD. And AE could improve depression, hippocampal neurogenesis, and increase BDNF/proBDNF ratio in the ischemic hippocampus of the PSD rats.

Most avian neurogenesis studies focused on the song control system and little attention has been given to non-song birds such as the Japanese quail. However, the only few neurogenesis studies in quails mainly focused on the sex steroid sensitive areas of the brain such as the medial preoptic and lateral septal nuclei. Despite the important role the quail telencephalon plays in filial imprinting and passive avoidance learning, neurogenesis in this structure has been completely overlooked. The aim of this study was therefore to quantitatively determine how DCX expression in the Japanese quail telencephalon changes with post hatching age (3-12 weeks) and life history stage. In this study, DCX was used as a proxy for neuronal incorporation. Bipolar and multipolar DCX immunoreactive cells were observed in the entire telencephalon except for the entopallium and arcopallium. In addition, DCX expression in all the eight telencephalic areas quantified was strongly negatively correlated with post-hatching age. Furthermore, numbers of bipolar and multipolar DCX immunoreactive cells were higher in the juvenile compared to subadult and adult quails. In conclusion, neuronal incorporation in the quail telencephalon is widespread but it declines with post hatching age. In addition, the most dramatic decline in neuronal incorporation in the telencephalic areas quantified takes place just after the birds have attained sexual maturity.

Repetitive transcranial magnetic stimulation (rTMS) is a physical treatment applied during recovery after intracerebral hemorrhage (ICH). With in vivo and in vitro assays, the present study sought to investigate how rTMS influences neural stem cells (NSCs) after ICH and the possible mechanism. Following a collagenase-induced ICH, adult male C57BL/6 J mice were subjected to rTMS treatment every 24 h for 5 days using the following parameters: frequency, 10 Hz; duration, 2 s; wait time, 5.5 s; 960 trains (500 µV/div, 5 ms/div, default setting). Brain water content and neurobehavioral score were assessed at days 1, 3, and 5 after ICH. The proliferation and differentiation of NSCs were observed using immunofluorescence staining for Nestin, Ki-67, DCX, and GFAP on day 3 after ICH, and rTMS treatment with the same parameters was applied to NSCs in vitro. We found that rTMS significantly reduced brain edema and alleviated neural functional deficits. The mice that underwent ICH recovered faster after rTMS treatment, with apparent proliferation and neuronal differentiation of NSCs and attenuation of glial differentiation and GFAP aggregation. Accordingly, proliferation and neuronal differentiation of isolated NSCs were promoted, while glial differentiation was reduced. In addition, microarray analysis, western blotting assays, and calcium imaging were applied to initially investigate the potential mechanism. Bioinformatics showed that the positive effect of rTMS on NSCs after ICH was largely related to the MAPK signaling pathway, which might be a potential hub signaling pathway under the complex effect exerted by rTMS. The results of the microarray data analysis also revealed that Ca2+ might be the connection between physical treatment and the MAPK signaling pathway. These predictions were further identified by western blotting analysis and calcium imaging. Taken together, our findings showed that rTMS after ICH exhibited a restorative effect by enhancing the proliferation and neuronal differentiation of NSCs, potentially through the MAPK signaling pathway.

The viral mimetic polyinosinic:polycytidylic acid (poly I:C) is an important tool to study the consequences of viral infection to the development of neuropsychiatric disorders. Here, based on the premise of omega-3 polyunsaturated fatty acids (n3 PUFAs) as supplemental treatment to antipsychotics in schizophrenia, we investigated the involvement of NFkB pathway in the effects of n3 PUFAs or of the atypical antipsychotic clozapine in hippocampal poly I:C-challenged neurons. Primary hippocampal neuronal cultures were exposed to n3 PUFAs (DHA4.35 μM/EPA7.10 μM, DHA 8.7 μM/EPA14.21 μM or DHA17.4 μM/EPA28.42 μM) or clozapine (1.5 or 3 μM) in the presence or absence of poly I:C. MTT assay revealed that poly I:C-induced reduction in cell viability was prevented by n3 PUFAs or clozapine. N3 PUFAs (DHA 8.7 μM/EPA14.21 μM) or clozapine (3 μM) significantly reduced poly I:C-induced increase in iNOS, NFkB (p50/p65), IL-6 and nitrite when compared to non-treated cells. Only n3 PUFAs prevented poly I:C-induced deficits in BDNF. On the other hand, poly I:C caused a marked reduction in DCX immunoexpression, which was prevented only by clozapine. Thus, n3 PUFAs and clozapine exert in vitro neuroprotective effects against poly I:C immune challenge in hippocampal neurons, by mechanisms possibly involving the inhibition of canonical NFkB pathway. The present study adds further evidences to the mechanisms underlying n3 PUFAs and clozapine neuroprotective effects against viral immune challenges. Since n3 PUFAs is a safe strategy for use during pregnancy, our results also add further evidence for the use of this supplement in order to prevent alterations induced by viral hits during this developmental period.

Hippocampal plasticity is hypothesized to play a role in the etiopathogenesis of depression and the antidepressant effect of medications. One form of plasticity that is unique to the hippocampus and is involved in depression-related behaviors in animal models is adult neurogenesis. While chronic electroconvulsive shock (ECS) strongly promotes neurogenesis, less is known about its acute effects and little is known about the neurogenic effects of other forms of stimulation therapy, such as repetitive transcranial magnetic stimulation (rTMS). Here, we investigated the time course of acute ECS and rTMS effects on markers of cell proliferation and neurogenesis in the adult hippocampus. Mice were subjected to a single session of ECS, 10 Hz rTMS (10–rTMS), or intermittent theta burst stimulation (iTBS). Mice in both TMS groups were injected with BrdU 2 days before stimulation to label immature cells. One, 3, or 7 days later, hippocampi were collected and immunostained for BrdU + cells, actively proliferating PCNA + cells, and immature DCX + neurons. Following ECS, mice displayed a transient increase in cell proliferation at 3 days post-stimulation. At 7 days post–stimulation there was an elevation in the number of proliferating neuronal precursor cells (PCNA + DCX +), specifically in the ventral hippocampus. iTBS and rTMS did not alter the number of BrdU + cells, proliferating cells, or immature neurons at any of the post-stimulation time points. Our results suggest that neurostimulation treatments exert different effects on hippocampal neurogenesis, where ECS may have greater neurogenic potential than iTBS and 10–rTMS.

Tamoxifen is frequently used in murine knockout systems with CreER/LoxP. Besides possible neuroprotective effects, tamoxifen is described as having a negative impact on adult neurogenesis. The present study investigated the effect of a high-dose tamoxifen application on Theiler’s murine encephalomyelitis virus (TMEV)-induced hippocampal damage. Two weeks after TMEV infection, 42% of the untreated TMEV-infected mice were affected by marked inflammation with neuronal loss, whereas 58% exhibited minor inflammation without neuronal loss. Irrespective of the presence of neuronal loss, untreated mice lacked TMEV antigen expression within the hippocampus at 14 days post-infection (dpi). Interestingly, tamoxifen application 0, 2 and 4, or 5, 7 and 9 dpi decelerated virus elimination and markedly increased neuronal loss to 94%, associated with increased reactive astrogliosis at 14 dpi. T cell infiltration, microgliosis and expression of water channels were similar within the inflammatory lesions, regardless of tamoxifen application. Applied at 0, 2 and 4 dpi, tamoxifen had a negative impact on the number of doublecortin (DCX)-positive cells within the dentate gyrus (DG) at 14 dpi, without a long-lasting effect on neuronal loss at 147 dpi. Thus, tamoxifen application during a TMEV infection is associated with transiently increased neuronal loss in the hippocampus, increased reactive astrogliosis and decreased neurogenesis in the DG.

The present in vivo study was conducted to evaluate whether hydrophilic (HL) or hydrophobic (HP) carbon nanotubes (CNTs) impregnated with subventricular zone neural progenitor cells (SVZ NPCs) could repair damaged neural tissue following stroke. For this purpose, stroke damaged rats were transplanted with HL CNT-SVZ NPCs, HP CNT-SVZ NPCs, or SVZ NPCs alone for 1, 3, 5, and 8 weeks. Results showed that the HP CNT-SVZ NPC transplants improved rat behavior and reduced infarct cyst volume and infarct cyst area compared with the experimental control and the HL CNT-SVZ NPC and SVZ NPCs alone groups. The transplantation groups showed an increase in the expression of nestin (cell stemness marker) and proliferation which was evident with the increased number of doublecortin and bromodeoxyuridine double-stained immunopositive cells around the lesion site. But, these effects were more prominent in the HP CNT-SVZ NPC group compared with the other transplantation groups. The HP CNT-SVZ NPC and HL CNT-SVZ NPC transplants increased the number of microtubule-associated protein 2 (marker for neurons) and decreased the number of glial fibrillary acidic protein (marker for astroglial cells) positive cells within the injury epicenter. The majority of the transplanted HP CNT-SVZ NPCs collectively broadened around the ischemic injured region and the SVZ NPCs differentiated into mature neurons, attained the synapse morphology (TUJ1, synaptophysin), and decreased microglial activation (CD11b/c [OX-42]). For these reasons, this study provided the first evidence that CNTs can improve stem cell differentiation to heal stroke damage and, thus, deserve further attention.

Menopause, natural or surgical, might facilitate the onset of psychiatric pathologies. Some reports suggest that their severity could increase if the decline of ovarian hormones occurs abruptly and before natural endocrine senescence. Therefore, we compared the effects of ovariectomy on microglia's morphological alterations, the complexity of newborn neurons, and the animal's ability to cope with stress. Young adult (3 months) and middle-aged (15 months) female Wistar rats were subjected to an ovariectomy (OVX) or were sham-operated. After 3 weeks, animals were assigned to one of the following independent groups: (1) young adult OVX + no stress; (2) young adult sham + no stress; (3) young adult OVX + stress; (4) young adult sham + stress; (5) middle-aged OVX + no stress; (6) middle-aged sham + no stress; (7) middle-aged OVX + stress; (8) middle-aged sham + stress. Acute stress was induced by forced swimming test (FST) exposure. Immobility behavior was scored during FST and 30 min after; animals were euthanized, their brains collected and prepared for immunohistochemical detection of Iba-1 to analyze morphological alterations in microglia, and doublecortin (DCX) detection to evaluate the dendrite complexity of newborn neurons. OVX increased immobility behavior, induced microglia morphological alterations, and reduced dendrite complexity of newborn neurons in young adult rats. FST further increased this effect. In middle-aged rats, the main effects were related to the aging process without OVX or stress exposure. In conclusion, surgical menopause favors in young adult rats, but not in middle-aged, the vulnerability to develop immobility behavior, retracted morphology of microglial cells, and decreased dendrite complexity of newborn neurons.

(1) Background: The c-Jun-NH2-terminal protein kinase (JNK) is a mitogen-activated protein kinase involved in regulating physiological processes in the central nervous system. However, the dual genetic deletion of Mkk4 and Mkk7 (upstream activators of JNK) in adult mice is not reported. The aim of this study was to induce the genetic deletion of Mkk4/Mkk7 in adult mice and analyze their effect in hippocampal neurogenesis. (2) Methods: To achieve this goal, Actin-CreERT2 (Cre+/−), Mkk4flox/flox, Mkk7flox/flox mice were created. The administration of tamoxifen in these 2-month-old mice induced the gene deletion (Actin-CreERT2 (Cre+/−), Mkk4∆/∆, Mkk7∆/∆ genotype), which was verified by PCR, Western blot, and immunohistochemistry techniques. (3) Results: The levels of MKK4/MKK7 at 7 and 14 days after tamoxifen administration were not eliminated totally in CNS, unlike what happens in the liver and heart. These data could be correlated with the high levels of these proteins in CNS. In the hippocampus, the deletion of Mkk4/Mkk7 induced a misalignment position of immature hippocampal neurons together with alterations in their dendritic architecture pattern and maturation process jointly to the diminution of JNK phosphorylation. (4) Conclusion: All these data supported that the MKK4/MKK7–JNK pathway has a role in adult neurogenic activity.

Background Adult hippocampal neurogenesis (AHN) is restricted under the pathological conditions of neurodegenerative diseases, especially in Alzheimer’s disease (AD). The drop of AHN reduces neural circuit plasticity, resulting in the decrease of the generation of newborn neurons in dentate gyrus (DG), which makes it difficult to recover from learning/memory dysfunction in AD, therefore, it is imperative to find a therapeutic strategy to promote neurogenesis and clarify its underlying mechanism involved. Methods Amyloid precursor protein/presenilin 1 (APP/PS1) mice were treated with photobiomodulation therapy (PBMT) for 0.1 mW/mm2 per day in the dark for 1 month (10 min for each day). The neural stem cells (NSCs) were isolated from hippocampus of APP/PS1 transgenic mice at E14, and the cells were treated with PBMT for 0.667 mW/mm2 in the dark (5 min for each time). Results In this study, photobiomodulation therapy (PBMT) is found to promote AHN in APP/PS1 mice. The latent transforming growth factor-β1 (LTGFβ1) was activated in vitro and in vivo during PBMT-induced AHN, which promoted the differentiation of hippocampal APP/PS1 NSCs into newborn neurons. In particular, behavioral experiments showed that PBMT enhanced the spatial learning/memory ability of APP/PS1 mice. Mechanistically, PBMT-stimulated reactive oxygen species (ROS) activates TGFβ/Smad signaling pathway to increase the interaction of the transcription factors Smad2/3 with Smad4 and competitively reduce the association of Smad1/5/9 with Smad4, thereby significantly upregulating the expression of doublecortin (Dcx)/neuronal class-III β-tubulin (Tuj1) and downregulating the expression of glial fibrillary acidic protein (GFAP). These in vitro effects were abrogated when eliminating ROS. Furthermore, specific inhibition of TGFβ receptor I (TGFβR I) attenuates the DNA-binding efficiency of Smad2/3 to the Dcx promotor triggered by PBMT. Conclusion Our study demonstrates that PBMT, as a viable therapeutic strategy, directs the adult hippocampal APP/PS1 NSCs differentiate towards neurons, which has great potential value for ameliorating the drop of AHN in Alzheimer’s disease mice.

ABSTRACТPurpose: To explore if the total body γ-irradiation at a dose of 0.1 Gy 7 days prior to acute mixed γ, n-irradiation of the head at the dose of 1 Gy can reduce the harmful effects of neutron irradiation on the hippocampal functions, neuroinflammation and neurogenesis.Materials and methods: Mice were exposed to γ-radiation alone, mixed γ,n-radiation or combined γ-rays and γ,n-radiation 7 days after γ-irradiation. Two months post-irradiation, mice were tested in Open Field and in the Morris water maze. The content of microglia, astrocytes, proliferating cells and cytokines TGF-β, TNF-α, IL-1β, GFAP levels, hippocampal BDNF, NT-3, NT-4, NGF mRNA expression were evaluated.Results: Two months after combined irradiation, we observed impaired hippocampus-dependent cognition, which was not detected in mice exposed to γ,n-irradiation. Combined exposure and γ,n-irradiation led to a significant increase in the level of activated microglia and astrocytes in the brains. The level of pro- and anti-inflammatory cytokines in the brain and hippocampal neurotrophine's genes changed differenly after the combined exposure and γ,n-irradiation. The quantity of DCX-positive cells was reduced after γ,n-irradiation exposer alone, but increased after combined irradiation.Conclusions: Our results indicate radio-adaptive responses in brains of mice that were exposed to low-dose gamma irradiation 7 days prior to acute 1 Gy γ,n-irradiation.

Immature neurons are maintained in cortical regions of the adult mammalian brain. In rodents, many of these immature neurons can be identified in the piriform cortex based on their high expression of early neuronal markers, such as doublecortin (DCX) and the polysialylated form of the neural cell adhesion molecule (PSA-NCAM). This molecule plays critical roles in different neurodevelopmental events. Taking advantage of a DCX-CreERT2/Flox-EGFP reporter mice, we investigated the impact of targeted PSA enzymatic depletion in the piriform cortex on the fate of immature neurons. We report here that the removal of PSA accelerated the final development of immature neurons. This was revealed by a higher frequency of NeuN expression, an increase in the number of cells carrying an axon initial segment (AIS), and an increase in the number of dendrites and dendritic spines on the immature neurons. Taken together, our results demonstrated the crucial role of the PSA moiety in the protracted development of immature neurons residing outside of the neurogenic niches. More studies will be required to understand the intrinsic and extrinsic factors affecting PSA-NCAM expression to understand how the brain regulates the incorporation of these immature neurons to the established neuronal circuits of the adult brain.

Levetiracetam (LEV) is a broad-spectrum antiepileptic drug with neuroprotective properties and novel mechanisms of action. Some evidence suggests that LEV may impact adult neurogenesis, but the results are controversial. The present study was aimed to evaluate the effects of LEV on the proliferation and differentiation of rat embryonic neural stem cells (NSCs) and to explore the role of GABANSCs were isolated from rat fetal ganglionic eminence at embryonic day 14.5. The effects of LEV on viability, proliferation, neurosphere formation, and neuronal or astroglial differentiation of NSCs were assessed using resazurin, BrdU incorporation, immunocytochemistry, quantitative real-time PCR, and western blotting. Additionally, we addressed the relationship between treatment with NMDA and GABAThe data showed that LEV (50 μM) significantly increased the number (p 0.01) and diameter of neurospheres (p 0.05), enhanced proliferation (p 0.01), and promoted neuronal differentiation, as revealed by significantly increased expressions of DCX and NeuN. The expressions of astroglial markers, GFAP and Olig2, were markedly reduced. The addition of MK801 (10 μM) significantly diminished neurospheres growth (p 0.001), decreased the number of proliferating cells (p 0.01), and reduced the number of new neurons (p 0.001) but increased the astroglial cells (p 0.001) induced by LEV. Co-treatment with saclofen (25 μM) did not significantly affect LEV-induced NSCs proliferation and differentiation.Our findings suggest that LEV may enhance rat embryonic neurogenesis mainly through an NMDA receptor-mediated mechanism.

Consumption of (−)-epicatechin (Epi), a cacao flavanol improves cognition. The aim was to compare the effects of (−)-Epi or its stereoisomer (+)-Epi on mouse frontal cortex-dependent short-term working memory and modulators of neurogenesis. Three-month-old male mice (n = 7 per group) were provided by gavage either water (vehicle; Veh), (−)-Epi, at 1 mg kg−1 or (+)-Epi at 0.1 mg per kg of body weight for 15 days. After treatment, spontaneous alternation was evaluated by Y-maze. Brain frontal cortex was isolated for nitrate/nitrite measurements, Western blotting for nerve growth factor (NGF), microtubule associated protein 2 (MAP2), endothelial and neuronal nitric oxide synthase (eNOS and nNOS) and immunohistochemistry for neuronal specific protein (NeuN), doublecortin (DCX), capillary (CD31) and neurofilaments (NF200). Results demonstrate the stimulatory capacity of (−)-Epi and (+)-Epi on markers of neuronal proliferation as per increases in immunoreactive cells for NeuN (74 and 120% respectively), DCX (70 and 124%) as well as in NGF (34.4, 63.6%) and MAP2 (41.8, 63.8%). Capillary density yielded significant increases with (−)-Epi (∼80%) vs. (+)-Epi (∼160%). CD31 protein levels increased with (−)-Epi (∼70%) and (+)-Epi (∼140%). Effects correlated with nitrate/nitrite stimulation by (–)-Epi and (+)-Epi (110.2, 246.5%) and enhanced eNOS phosphorylation (Ser1177) with (–)-Epi and (+)-Epi (21.4, 41.2%) while nNOS phosphorylation only increased with (+)-Epi (18%). Neurofilament staining was increased in (–)-Epi by 135.6 and 84% with (+)-Epi. NF200 increased with (–)-Epi (116%) vs. (+)-Epi (84.5%). Frontal cortex-dependent short-term spatial working improved with (–)-Epi and (+)-Epi (15, 13%). In conclusion, results suggest that both enantiomers, but more effectively (+)-Epi, upregulate neurogenesis markers likely through stimulation of capillary formation and NO triggering, improvements in memory.

Background/aim Stroke is among the most common causes of disability and death in highly developed countries and China. We sought to study the role of oleanolic acid in cerebral ischemia-reperfusion injury. Methods Middle cerebral artery occlusion (MCAO) was performed to induce cerebral ischemia-reperfusion injury in mice. For the short-term effects of oleanolic acid (OA) against MCAO, mice administrated with OA (6 mg/kg /d) for 3 days before the injury were evaluated the infarct volume, neurological scores, blood brain barrier permeability and oxidative stress level, while for the long-term effects, MCAO mice were injected daily with OA for 6 weeks, followed by assessments of motor function, behavior and cerebral infarction area. Results Pretreatment of oleanolic acid alleviated MCAO-induced ischemia-reperfusion injury as indicated by the significant decreases in cerebral infarction area and neurological symptom score at 24 h post injury, Evans blue leakage, expression of matrix metalloproteinase 9 (MMP9) and occludin, dihydroethidium fluorescence, and block malonaldehyde generation. In the long run, OA significantly reduced brain loss, enhanced the motor function, promoted the recovery of nerve function, and improved the learning and memory ability 9 weeks after the ischemia-reperfusion injury. OA also inhibited astrocytes proliferation and microglia activation, promoted the expression of synapse-related proteins, and increased the number of DCX+ cells in the hippocampus. Conclusions OA exhibits both short-term and long-term protective effects against the cerebral ischemia-reperfusion injury in mice. The short-term protective mechanism is related to the anti-oxidation of blood-brain barrier, while the long-term protective effect lies in neuroglia modulation, promotion of synaptic connection and neuroregeneration.

To observe the effect of moxibustion on learning-memory ability and expression of hippocampal inflammatory factors and microtubule-associated protein doublecortin (DCX, a marker of neuronal regeneration) in vascular dementia (VD) rats, so as to explore its mechanisms underlying improvement of VD.SD rats were randomly divided into normal control, sham operation, VD model, moxibustion and medication groups (Following modeling, the average escape latency was significantly longer in the model group than in the normal control and sham operation groups (Moxibustion can improve learning-memory ability in VD rats, which may be related to its effect in down-regulating the expression of inflammatory factors and up-regulating the expression of DCX to promote neuronal repair and regeneration.

Adolescence represents a crucial period for maturation of brain structures involved in cognition. Early in life unhealthy dietary patterns are associated with inferior cognitive outcomes at later ages; conversely, healthy diet is associated with better cognitive results. In this study we analyzed the effects of a short period of hypercaloric diet on newborn hippocampal doublecortin+ (DCX) immature neurons in adolescent mice. Male mice received high fat diet (HFD) or control low fat diet (LFD) from the 5th week of age for 1 or 2 weeks, or 1 week HFD followed by 1 week LFD. After diet supply, mice were either perfused for immunohistochemical (IHC) analysis or their hippocampi were dissected for biochemical assays. Detailed morphometric analysis was performed in DCX+ cells that displayed features of immature neurons. We report that 1 week-HFD was sufficient to dramatically reduce dendritic tree complexity of DCX+ cells. This effect occurred specifically in dorsal and not ventral hippocampus and correlated with reduced BDNF expression levels in dorsal hippocampus. Both structural and biochemical changes were reversed by a return to LFD. Altogether these studies increase our current knowledge on potential consequences of hypercaloric diet on brain and in particular on dorsal hippocampal neuroplasticity.

We previously introduced the brain erythropoietin (EPO) circle as a model to explain the adaptive ‘brain hardware upgrade’ and enhanced performance. In this fundamental circle, brain cells, challenged by motor-cognitive tasks, experience functional hypoxia, triggering the expression of EPO among other genes. We attested hypoxic cells by a transgenic reporter approach under the ubiquitous CAG promoter, with Hif-1α oxygen-dependent degradation-domain (ODD) fused to CreERT2-recombinase. To specifically focus on the functional hypoxia of excitatory pyramidal neurons, here, we generated CaMKIIα-CreERT2-ODD::R26R-tdTomato mice. Behavioral challenges, light-sheet microscopy, immunohistochemistry, single-cell mRNA-seq, and neuronal cultures under normoxia or hypoxia served to portray these mice. Upon complex running wheel performance as the motor-cognitive task, a distinct increase in functional hypoxic neurons was assessed immunohistochemically and confirmed three-dimensionally. In contrast, fear conditioning as hippocampal stimulus was likely too short-lived to provoke neuronal hypoxia. Transcriptome data of hippocampus under normoxia versus inspiratory hypoxia revealed increases in CA1 CaMKIIα-neurons with an immature signature, characterized by the expression of Dcx, Tbr1, CaMKIIα, Tle4, and Zbtb20, and consistent with accelerated differentiation. The hypoxia reporter response was reproduced in vitro upon neuronal maturation. To conclude, task-associated activity triggers neuronal functional hypoxia as a local and brain-wide reaction mediating adaptive neuroplasticity. Hypoxia-induced genes such as EPO drive neuronal differentiation, brain maturation, and improved performance.

Decreased neurogenesis after brain exposure to ionizing radiation is linked to neurocognitive impairments. Using transgenic mouse models, we previously showed that abrogation of radiation-induced senescence, or apoptosis, can partially rescue neurogenesis in the subventricular and hippocampus regions. Here, we evaluate whether the injection of recombinant epidermal growth factor (rEGF) or mesenchymal stromal cells (MSC) engineered to secrete EGF (MSC-EGF) can preserve neurogenesis. Using doublecortin (Dcx) expression and BrdU incorporation assays, we found that the injection of rEGF into the subventricular zone (SVZ) promotes neurogenesis, despite increasing apoptosis, in the brain of irradiated mice. The effect of rEGF was mostly localized, as Dcx expression was not induced in the hippocampus region and limited in the contralateral SVZ. Surprisingly, the injection of bone marrow-derived MSC alone, or secreting EGF, did not result in increased neurogenesis despite the fact that part of the MSC survived a few weeks after injection. Our results suggest that only a supraphysiological concentration of rEGF can promote neurogenesis, likely through a direct mitogenic effect.

To investigate the mechanism of Chinese medicine Buyang Huanwu decoction (BYHWD) promoting neurogenesis and angiogenesis in ischemic stroke rats.Male SD rats were randomly divided into sham operation group, model group, BYHWD group, antagonist group and antagonist control group with 14 rats in each. Focal cerebral ischemia was induced by occlusion of the right middle cerebral artery for 90 min with intraluminal filament and reperfusion for 14 d in all groups except sham operation group. BYHWD (13 g/kg) was administrated by gastrogavage in BYHWD group, antagonist group and antagonist control group at 24 h after modeling respectively, and BrdU (50 mg/kg) was injected intraperitoneally in all groups once a day for 14 consecutive days. miR-199a-5p antagomir or NC (10 nmol) was injected into the lateral ventricle at d5 after ischemia in antagonist and antagonist control groups, respectively. The neurological deficits were evaluated by the modified neurological severity score (mNSS) and the corner test, and the infract volume was measured by toluidine blue staining. Neurogenesis and angiogenesis were detected by immunofluorescence double labeling method. The expression level of miR-199a-5p was tested by real-time RT-PCR, and the protein expressions of vascular endothelial growth factor (VEGF) and brain-derived neurotrophic factor (BDNF) were determined by Western blotting.BYHWD treatment significantly promoted the recovery of neurological function (Buyang Huanwu decoction promotes neurogenesis and angiogenesis in rats with cerebral ischemia, which may be related to increased protein expression of VEGF and BDNF through upregulating miR-199a-5p.

Fetal alcohol spectrum disorder is the main preventable cause of intellectual disability in the Western world. Although binge drinking is the most studied prenatal alcohol exposure pattern, other types of exposure, such as the Mediterranean, are common in specific geographic areas. In this study, we analyze the effects of prenatal alcohol exposure in binge and Mediterranean human drinking patterns on placenta and brain development in C57BL/6J mice. We also assess the impact of prenatal treatment with the epigallocatechin-3-gallate antioxidant in both groups. Study experimental groups for Mediterranean or binge patterns: (1) control, (2) ethanol, (3) ethanol + epigallocatechin-3-gallate. Brain and placental tissue were collected on gestational Day 19. The molecular pathways studied were fetal and placental growth, placental angiogenesis (VEGF-A, PLGF, VEGF-R), oxidative stress (Nrf2), and neurodevelopmental processes including maturation (NeuN, DCX), differentiation (GFAP) and neural plasticity (BDNF). Prenatal alcohol exposure resulted in fetal growth restriction and produced imbalances of placental angiogenic factors. Moreover, prenatal alcohol exposure increased oxidative stress and caused significant alterations in neuronal maturation and astrocyte differentiation. Epigallocatechin-3-gallate therapy ameliorated fetal growth restriction, attenuated alcohol-induced changes in placental angiogenic factors, and partially rescued neuronal nuclear antigen (NeuN), (doublecortin) DCX, and (glial fibrillary acidic protein) GFAP levels. Any alcohol consumption (Mediterranean or binge) during pregnancy may generate a fetal alcohol spectrum disorder phenotype and the consequences may be partially attenuated by a prenatal treatment with epigallocatechin-3-gallate.

Aim Neurogenesis is an age-related process that is closely associated with neurological disorders. In the present study, we aimed to investigate neurogenesis in both adult and 3-week-old genetic absence epilepsy rats from Strasbourg (GAERS) to determine if newly formed neurons within the dentate gyrus (DG) form synaptic contacts with GABAergic (gamma aminobutyric acid) and glutamatergic nerve terminals and compared to the control (non-GAERS) Wistar rats. Material and methods Brain tissue was processed for electron microscopic assessment. Thin sections from the hippocampal DG were double-labelled for anti-GABA or anti-VGLUT1 (vesicular glutamate transporter 1) and anti-doublecortin (DCX) antibodies using immunogold methodology and examined with the transmission electron microscope for morphological changes and to quantify the density of gold labeling. Results DCX immunoreactivity was demonstrated within axon terminals, dendrites and somata in all groups. DCX and GABA or VGLUT1 were found to be co-localized in the axon terminals in all groups. We observed that DCX-immunoreactive (-ir) profiles formed synaptic contacts with GABAergic and glutamatergic terminals. The percentage of DCX labeling in dendrites, compared to axons, and the percentage of DCX-ir terminal profiles forming asymmetrical synapses, compared to those forming symmetrical synapses, were increased in all groups compared to the control group. DCX immunoreactivity in the 21-day-old GAERS group was found to be increased compared to the Wistar group. Conclusion We conclude that newly born neurons are incorporated into the local hippocampal network in both the GAERS and the control Wistar rats. The results suggest that the neurogenesis taking place in the hippocampus may also be involved in the mechanism underlying absence seizures in GAERS.

New neurons born in the adult brain undergo a critical period soon after migration to their site of incorporation. During this time, the behavior of the animal may influence the survival or culling of these cells. In the songbird song system, earlier work suggested that adult-born neurons may be retained in the song motor pathway nucleus HVC with respect to motor progression toward a target song during juvenile song learning, seasonal song restructuring, and experimentally manipulated song variability. However, it is not known whether the quality of song per se, without progressive improvement, may also influence new neuron survival. To test this idea, we experimentally altered song acoustic structure by unilateral denervation of the syrinx, causing a poor quality song. We found no effect of aberrant song on numbers of new neurons in HVC, suggesting that song quality does not influence new neuron culling in this region. However, aberrant song resulted in the loss of left-side dominance in new neurons in the auditory region caudomedial nidopallium (NCM), and a bilateral decrease in new neurons in the basal ganglia nucleus Area X. Thus new neuron culling may be influenced by behavioral feedback in accordance with the function of new neurons within that region. We propose that studying the effects of singing behaviors on new neurons across multiple brain regions that differentially subserve singing may give rise to general rules underlying the regulation of new neuron survival across taxa and brain regions more broadly.

Objective Parenchymal arterioles (PAs) regulate perfusion of the cerebral microcirculation, and impaired PA endothelium-dependent dilation occurs in dementia models mimicking chronic cerebral hypoperfusion (CCH). Epoxyeicosatrienoic acids (EETs) are vasodilators; their actions are potentiated by soluble epoxide hydrolase (sEH) inhibition. We hypothesized that chronic sEH inhibition with trifluoromethoxyphenyl-3 (1-propionylpiperidin-4-yl) urea (TPPU) would prevent cognitive dysfunction and improve PA dilation in a hypertensive CCH model. Methods Bilateral carotid artery stenosis (BCAS) was used to induce CCH in twenty-week-old male stroke-prone spontaneously hypertensive rats (SHSRP) that were treated with vehicle or TPPU for 8 weeks. Cognitive function was assessed by novel object recognition. PA dilation and structure were assessed by pressure myography, and mRNA expression in brain tissue was assessed by qRT-PCR. Results TPPU did not enhance resting cerebral perfusion, but prevented CCH-induced memory deficits. TPPU improved PA endothelium-dependent dilation but reduced the sensitivity of PAs to a nitric oxide donor. TPPU treatment had no effect on PA structure or biomechanical properties. TPPU treatment increased brain mRNA expression of brain derived neurotrophic factor, doublecortin, tumor necrosis factor-alpha, sEH, and superoxide dismutase 3, CONCLUSIONS: These data suggest that sEH inhibitors may be viable treatments for cognitive impairments associated with hypertension and CCH.

Graphical abstract The dentate gyrus (DG) in rodent hippocampus is a site of neurogenesis. Newly developing neurons can be identified through immunostaining for doublecortin (DCX). We have found that genetic inactivation of the kinase activity of MSK1 (in red) results in an increase in the number of DCX-positive (DCX+) cells. This suggests that MSK1 negatively regulates neurogenesis in the subgranular zone, potentially to limit the number of new neurons in the dentate gyrus and maintain the stability of neuronal networks., Highlights • Neurogenesis in the subgranular zone of the hippocampal dentate gyrus was examined. • Ki-67 staining was not affected by the loss of MSK1 kinase activity. • DCX staining was increased in the MSK1 kinase dead mutant mouse. • MSK1 may negatively regulate neurogenesis to maintain stable neuronal networks., Neurogenesis in the subgranular zone (SGZ) of the adult hippocampus can be stimulated by a variety of means, including via exposure of experimental animals to an enriched environment that provides additional sensory, social, and motor stimulation. Tangible health and cognitive benefits accrue in enriched animals, including the amelioration of signs modelling psychiatric, neurological and neurodegenerative conditions that affect humans, which may in part be due to enhanced production of neurons. A key factor in the neuronal response to enrichment is the release of brain-derived neurotrophic factor (BDNF) and the activation of the Mitogen-Activated Protein Kinase (MAPK) cascade, which can lead to the stimulation of neurogenesis. Mitogen- and Stress-Activated protein Kinase 1 (MSK1) is a nuclear enzyme downstream of BDNF and MAPK that regulates transcription. MSK1 has previously been implicated in both basal and stimulated neurogenesis on the basis of studies with mice lacking MSK1 protein. In the present study, using mice in which only the kinase activity of MSK1 is lacking, we show that the rate of cellular proliferation in the SGZ (Ki-67 staining) is unaffected by the MSK1 kinase-dead (KD) mutation, and no different from controls levels after five weeks of enrichment. However, compared to wild-type mice, the number of doublecortin (DCX)-positive cells was greater in both standard-housed and enriched MSK1 KD mice. These observations suggest that, while MSK1 does not influence the basal rate of proliferation of neuronal precursors, MSK1 negatively regulates the number of cells destined to become neurons, potentially as a homeostatic control on the number of new neurons integrating into the dentate gyrus.

Failure of extinction of fear-conditioned traumatic memory is the main pathology behind post-traumatic stress disorder (PTSD). Functional and structural dysfunctions in the olfactory system are implicated by studies in PTSD patients. However, little is known regarding the neurobiological networks of trauma‑related odor sensitivity in PTSD. Male Wistar rats were exposed with a female cat for 10 min and long-term stress was evaluated by behavioral tests, containing open field (OF) and elevated plus maze (EPM). To prove the PTSD model, the serum level of cortisol was evaluated and compared with the control group. Local field potential (LFP) was applied to compare the electrophysiology of the OB in two groups. To assess neurogenesis, the expression of nestin, and doublecortin were evaluated. Data from EPM revealed a significant increase in spent time in the closed arms in PTSD group. We observed a significant reduction in OF parameters in terms of the total distance traveled, the time spent in the central zone, and the number of crossing the central zone in PTSD group compared to the control group. The mean serum cortisol level was significantly higher in the PTSD group than the control group. In LFP recording, the slope and the amplitude of field excitatory postsynaptic potential (fEPSP) in the PTSD group were significantly higher than that of the control group. Our results also showed that the mRNA expression level of nestin as a neural progenitor marker and doublecortin, as an immature neuron marker, significantly decreased in the PTSD group compared to the control group. This study has shown that PTSD can disrupt the OB function through decreasing neurogenesis. More information on PTSD and OB would help us to establish a greater degree of accuracy on this matter.

Background Adult neurogenesis observed both in the subventricular zone (SVZ) and hippocampus may be regulated and modulated by several endogenous factors, xenobiotics and medications. Classical and atypical antipsychotic drugs are able to affect neuronal and glial cell proliferation in the rat brain. The main purpose of this structural study was to determine whether chronic chlorpromazine treatment affects adult neurogenesis in the canonical sites of the rat brain. At present, the clinical application of chlorpromazine is rather limited; however, it may still represent an important model in basic neuropharmacological and toxicological studies. Methods The number of neural progenitors and immature neurons was enumerated using immunofluorescent detection of Sox2, Musashi1 and doublecortin (DCX) expression within SVZ. Results Chlorpromazine has a depressive effect on the early phase of adult neurogenesis in the rat subventricular zone (SVZ), as the mean number of Sox-2 immunoexpressing cells decreased following treatment. Conclusion Collectively, these results may suggest that long-term treatment with chlorpromazine may decrease neurogenic stem/progenitor cell formation in the rat SVZ and may affect rostral migratory stream formation.

Huntington’s disease (HD) is a neurodegenerative disorder with a progressive loss of medium spiny neurons in the striatum and aggregation of mutant huntingtin in the striatal and cortical neurons. Currently, there are no rational therapies for the treatment of the disease. Cerebral dopamine neurotrophic factor (CDNF) is an endoplasmic reticulum (ER) located protein with neurotrophic factor (NTF) properties, protecting and restoring the function of dopaminergic neurons in animal models of PD more effectively than other NTFs. CDNF is currently in phase I–II clinical trials on PD patients. Here we have studied whether CDNF has beneficial effects on striatal neurons in in vitro and in vivo models of HD. CDNF was able to protect striatal neurons from quinolinic acid (QA)-induced cell death in vitro via increasing the IRE1α/XBP1 signalling pathway in the ER. A single intrastriatal CDNF injection protected against the deleterious effects of QA in a rat model of HD. CDNF improved motor coordination and decreased ataxia in QA-toxin treated rats, and stimulated the neurogenesis by increasing doublecortin (DCX)-positive and NeuN-positive cells in the striatum. These results show that CDNF positively affects striatal neuron viability reduced by QA and signifies CDNF as a promising drug candidate for the treatment of HD.

The feasibility of delivering mitochondria intranasally so as to bypass the blood–brain barrier in treating Parkinson's disease (PD), was evaluated in unilaterally 6-OHDA-lesioned rats. Intranasal infusion of allogeneic mitochondria conjugated with Pep-1 (P-Mito) or unconjugated (Mito) was performed once a week on the ipsilateral sides of lesioned brains for three months. A significant improvement of rotational and locomotor behaviors in PD rats was observed in both mitochondrial groups, compared to sham or Pep-1-only groups. Dopaminergic (DA) neuron survival and recovery > 60% occurred in lesions of the substantia nigra (SN) and striatum in Mito and P-Mito rats. The treatment effect was stronger in the P-Mito group than the Mito group, but the difference was insignificant. This recovery was associated with restoration of mitochondrial function and attenuation of oxidative damage in lesioned SN. Notably, P-Mito suppressed plasma levels of inflammatory cytokines. Mitochondria penetrated the accessory olfactory bulb and doublecortin-positive neurons of the rostral migratory stream (RMS) on the ipsilateral sides of lesions and were expressed in striatal, but not SN DA neurons, of both cerebral hemispheres, evidently via commissural fibers. This study shows promise for intranasal delivery of mitochondria, confirming mitochondrial internalization and migration via RMS neurons in the olfactory bulb for PD therapy.

Methotrexate (MTX) is a chemotherapeutic drug commonly used to treat cancers that has an adverse effect on patients' cognition. Metformin is a primary treatment for type 2 diabetes mellitus that can pass through the blood-brain barrier. Metformin has neuroprotective actions, which can improve memory. In the present study, we examined the ability of metformin in MTX chemotherapy-generated cognitive and hippocampal neurogenesis alterations. Male Sprague-Dawley rats were allocated into control, MTX, metformin, preventive, and throughout groups. MTX (75 mg/kg/day) was given intravenously on days 7 and 14 of the study. Metformin (200 mg/kg/day) was injected intraperitoneally for 14 days. Some of the MTX-treated rats received co-treatment with metformin once a day for either 14 (preventive) or 28 days (throughout). After treatment, memory ability was evaluated using novel object location and novel object recognition tests. Ki67 (proliferating cells), BrdU (survival cells), and doublecortin (immature neurons, DCX) positive cells in the subgranular zone (SGZ) of the hippocampal dentate gyrus were quantified. We found that reductions of cognition, the number of proliferating and survival cells and immature neurons in the SGZ were ameliorated in the co-treatment groups, which suggests that metformin can prevent memory and hippocampal neurogenesis impairments induced by MTX in adult rats.

Introduction Recent animal studies showed that isoflurane exposure may lead to the disturbance of hippocampal neurogenesis and later cognitive impairment. However, much less is known about the effect of isoflurane exposure on the neurons generated form tertiary dentate matrix, even though a great increase of granule cell population during the infantile period is principally derived from this area. Methods To label the new cells originated from the tertiary dentate matrix, the mice were injected with BrdU on postnatal day 6 (P6). Then, the mice were exposed to isoflurane for 4 hr at 1, 8, 21, and 42 days after BrdU injection, and the brains were collected 24 hr later. The loss of newly generated cells/neurons with different developmental stage was assessed by BrdU, BrdU + DCX, BrdU + NeuN, or BrdU + Prox‐1 staining, respectively. Results We found that the isoflurane exposure significantly decreased the numbers of nascent cells (1 day old) and mature neurons (42 days old), but had no effect on the immature (8 days old) and early mature neurons (8 and 21 days old, respectively). Conclusion The results suggested isoflurane exposure exerts the neurotoxic effects on the tertiary dentate matrix‐originated cells with an age‐defined pattern in mice, which partly explain the cognitive impairment resulting from isoflurane exposure to the young brain., Isoflurane exposure exerts neurotoxic effects on the tertiary dentate matrix‐originated cells with an age‐defined pattern in mice. The isoflurane significantly decreased the numbers of nascent cells and mature neurons driven from the tertiary dentate matrix.

Background Women are more vulnerable than men to the neurotoxicity and severe brain damage caused by chronic heavy alcohol use. In addition, brain damage due to chronic heavy alcohol use may be associated with sex-dependent epigenetic modifications. This study aimed to identify microRNAs (miRNAs) and their target genes that are differentially expressed in the hippocampi of male and female animal models in response to alcohol. Methods After chronic alcohol administration (3~3.5 g/kg/day) in male (control, n = 10; alcohol, n = 12) or female (control, n = 10; alcohol, n = 12) Sprague-Dawley rats for 6 weeks, we measured body weights and doublecortin (DCX; a neurogenesis marker) concentrations and analyzed up- or downregulated miRNAs using GeneChip miRNA 4.0 arrays. The differentially expressed miRNAs and their putative target genes were validated by RT-qPCR. Results Alcohol attenuated body weight gain only in the male group. On the other hand, alcohol led to increased serum AST in female rats and decreased serum total cholesterol concentrations in male rats. The expression of DCX was significantly reduced in the hippocampi of male alcohol-treated rats. Nine miRNAs were significantly up- or downregulated in male alcohol-treated rats, including upregulation of miR-125a-3p, let-7a-5p, and miR-3541, and downregulation of their target genes (Prdm5, Suv39h1, Ptprz1, Mapk9, Ing4, Wt1, Nkx3-1, Dab2ip, Rnf152, Ripk1, Lin28a, Apbb3, Nras, and Acvr1c). On the other hand, 7 miRNAs were significantly up- or downregulated in alcohol-treated female rats, including downregulation of miR-881-3p and miR-504 and upregulation of their target genes (Naa50, Clock, Cbfb, Arih1, Ube2g1, and Gng7). Conclusions These results suggest that chronic heavy alcohol use produces sex-dependent effects on neurogenesis and miRNA expression in the hippocampus and that sex differences should be considered when developing miRNA biomarkers to diagnose or treat alcoholics.

Rosmarinus officinalis, commonly known as rosemary, is a medicinal herb that presents significant biological properties such as antimicrobial, antioxidant, anti-inflammatory, anti-diabetic and anti-depressant activities. Recent findings correlate impaired adult neurogenesis, which is crucial for the maintenance of synaptic plasticity and hippocampal functioning, synaptic regulation with the pathological hallmarks of Alzheimer's disease (AD). These observations call for the need to developing compounds that promote neurogenesis and alleviates deficits in cognition and synaptic regulation.The present study was conducted to determine the proneurogenic effects of R. officinalis and its active compounds (ursolic acid and rosmarinic acid) in comparison to Donepezil in an AβBALB/c mice were divided into ten groups. Half were injected with AβThe results show a protective effect by rosmarinic acid and ursolic acid in reversing the deficits in spatial and recognition memory as well as changes in anxiety induced by AβOur findings indicate the potential of R. officinalis and its active compounds as therapeutic agents against Aβ

The aim of this work was to evaluate the physicochemical and biological properties of docetaxel (DCX) loaded chitosan nanocapsules (CS Nc) functionalized with the monoclonal antibody Chi-Tn (CS-PEG-ChiTn mAb Nc) as a potential improvement treatment for cancer therapy. The Tn antigen is highly specific for carcinomas, and this is the first time that such structure is targeted for drug delivery. The nanocapsules (Ncs), formed as a polymeric shell around an oily core, allowed a 99.9% encapsulation efficiency of DCX with a monodispersity particle size in the range of 200 nm and a high positive surface charge that provide substantial stability to the nanosystems. Release profile of DCX from Ncs showed a sustained and pH dependent behavior with a faster release at acidic pH, which could be favorable in the intracellular drug delivery. We have designed PEGylated CS Nc modified with a monoclonal antibody which recognize Tn antigen, one of the most specific tumor associated antigen. A biotin-avidin approach achieved the successful attachment of the antibody to the nanocapsules. Uptake studies and viability assay conducted in A549 human lung cancer cell line in vitro demonstrate that ChiTn mAb enhance nanoparticles internalization and cell viability reduction. Consequently, these ChiTn functionalized nanocapsules are promising carriers for the active targeting of DCX to Tn expressing carcinomas.

In recent years, our understanding of neural circuits associated with depression has increased. Although inherited factors are known to influence individual differences in the risk for this disorder, it has been difficult to identify specific genes that moderate circuit functions affecting depression. Genome-wide association studies have identified genetic variants of Cntn1 that are linked to major depressive disorders. Cntn1, a subset of the neural cell adhesion protein and immunoglobulin supergene family, participates in cell contact formation and axonal growth control and plays a role in degenerative and inflammatory disorders. However, neuronal substrates that mediate Cntn1 action on depression-like phenotypes and involved mechanisms are unclear. Here, we exploited chronic unpredictable stress (CUS) exposure and found that CUS treatment significantly increased hippocampal Cntn1 messenger RNA and protein expression in both mice and rats, but not in the medial prefrontal cortex, which presented a region-specific regulation. Using an adeno-associated virus-based approach to directly overexpress Cntn1 via stereotactic injection, we demonstrated that Cntn1 overexpression in the hippocampus triggered anxiety- and depression-like phenotypes in addition to microglia activation or phagocytosis in the hippocampus, resulting in upregulation of pro-inflammatory cytokine (IL1α, IL6, and Ccl2) mRNA expression and downregulation of anti-inflammatory cytokine (IL4 and CD206) mRNA expression, determined using real-time quantitative PCR, thus impairing hippocampal immature neurons in the dentate gyrus, determined using immunohistochemical staining for doublecortin, a specific marker for immature neurons. Collectively, our results identified Cntn1 as a novel risk gene involved in regulating anxiety and depression via functional actions in the hippocampus that is correlated with microglial activation or phagocytosis and reduced hippocampal immature neurons. These results may provide a better understanding of the pathophysiological mechanisms underlying the risk of depression-related disorders.