Morphological Changes Of Hippocampal Neurons Research Articles

To explore the protective mechanism of promethazine against hippocampal neuron injury based on network pharmacology and experimental verification.

This study aims to investigate the effect of promethazine (PMZ) on hippocampal neuronal injury through network pharmacology and in vivo experiments. Network pharmacology: The intersection genes of PMZ and Alzheimer Disease (AD) were obtained, and the core genes of PMZ in AD were screened. The intersection genes were enriched by Gene Ontology and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses. In the in vitro experiment, mouse hippocampal neurons (HT22) were divided into control, glutamate (GLU) model, and GLU + PMZ treatment groups. The control group was given a complete culture medium, the model group was given GLU for 24 hours, the treatment group was given PMZ pretreatment for 3 hours, and then GLU was administered for 24 hours. Cell viability was determined, cell morphology was observed by microscopy, reactive oxygen species levels and glutathione content were detected, and protein expression of P53, PTGS2, SLC7A11, and GPX4 was detected by western blotting. Network pharmacology: A total of 317 PMZ targets, 1934 AD genes, 125 intersection genes, and 18 core genes, including P53 and PTGS2. Gene Ontology enrichment analysis showed that the effect of PMZ on AD was mainly related to cell proliferation, inflammation, hypoxia, synaptic structure, plasma membrane, and oxidoreductase activity. Kyoto Encyclopedia of Genes and Genomes results showed neuroactive ligand-receptor interaction, cell senescence, cancer pathway, PI3K-AKT signal pathway, neurodegeneration, and HIF-1 signal pathway. In vitro experiments: PMZ improved the GLU-induced decrease in cell viability and morphological changes in hippocampal neurons. PMZ inhibited reactive oxygen species levels and increased glutathione content in injured hippocampal neurons. Up-regulated of P53, SLC7A11 and GPX4 expression, and inhibited expression of PTGS2. PMZ regulates the SLC7A11-GPX4 antioxidant system to protect hippocampal neurons from oxidative stress injury.

The Mechanism of Propofol in Treating Depression-like Behaviors Based on Network Pharmacology and Experimental Validation

Objective: To explore the potential mechanism of propofol in improving antidepressant-like behavior through network pharmacology and animal experimental validation, providing theoretical and experimental support for the development of novel antidepressant drugs based on propofol. Methods: 1. The targets of propofol and the disease targets of depression were screened through Swiss Target prediction, Super-pred, SEA, Drugbank, CTD, GeneCards, OMIM, and TTD databases. A protein-protein interaction (PPI) network was constructed, and the core targets were screened and visualized using CytoScape 3.10.2 software. Additionally, GO and KEGG enrichment analyses of the intersection targets were performed using the Metascape database. 2. Thirty male C57BL/6 mice were randomly divided into the CON group, LH model group, and PRO treatment group, with 10 mice in each group. The sucrose preference test, forced swim test, and tail suspension test were conducted to determine whether the depression model was successfully established and to assess the improvement effect of propofol on antidepressant-like behavior. 3. The pathological morphological changes of hippocampal neurons in the three groups of mice were observed through HE staining experiments. The expression of mGluR5 protein in the hippocampus was analyzed by Western blot. Results: 1. Network pharmacology analysis indicated that propofol may exert antidepressant effects by acting on core genes such as ALB, ESR1, NFKB1, HSP90AB1, and EGFR. These core target genes were mainly enriched in biological processes such as synaptic transmission and membrane potential regulation; they involved cellular components such as GABA receptor complexes and synaptic membranes; and they included molecular functions such as neurotransmitter receptor activity. Additionally, propofol may exert antidepressant effects through signaling pathways such as neuroactive ligand-receptor interaction, morphine addiction, and GABAergic synapse. 2. Compared with the CON group, the sucrose preference rate of mice in the LH model group significantly decreased, while the immobility time in the forced swim test and tail suspension test significantly increased. Compared with the LH model group, the sucrose preference rate of mice in the PRO treatment group significantly increased, and the immobility time significantly decreased. 3. The HE results showed that compared with the CON group, the number of neurons in the LH group decreased, with loose arrangement, pycnotic and deeply stained nuclei with blurred boundaries. However, the number of necrotic neurons in the PRO group significantly decreased. The Western blot results showed that compared with the CON group, the expression level of mGluR5 protein in the LH model group significantly increased, while it significantly decreased in the PRO treatment group compared with the LH model group. Conclusion: Propofol may exert antidepressant effects by regulating the expression of mGluR5, improving antidepressant-like behavior, and reducing hippocampal neuronal necrosis.

MiR-351-5p regulation of CPEB3 affecting aluminium-induced learning and memory impairment in SD rats

Aluminium exposure has been found to impair learning and memory abilities; however, the underlying molecular mechanisms remain unclear. In this study we conducted a double luciferase reporter assay to determine whether miR-351-5p regulates cytoplasmic polyadenylation element binding protein (CPEB) 3 mRNA. To this end, we overexpressed and inhibited miR-351-5p via stereotaxic microinjections of adeno-associated virus (AAV) into the hippocampus of Sprague Dawley rats in a sub-chronic aluminium exposure model to examine learning and memory ability using Morris water maze. Ultrastructural electron microscopy and Golgi staining were used to examine morphological changes in hippocampal neurons. In addition, we examined the levels of synaptic plasticity-related proteins (PRPs) and CPEB3 to determine the involvement of the miR-351-5P/CPEB3/PRPs pathway in aluminium neurotoxicity. Sub-chronic aluminium exposure reduced the spatial learning and memory ability of rats. Overexpression of AAV-miR-351-5P in the hippocampus aggravated the impairment of spatial learning and memory abilities of aluminium-treated rats, whereas inhibition of AAV-miR-351-5p expression alleviated it. Western blotting suggested that sub-chronic aluminium exposure increased miR-351-5p levels and reduced the expression of CPEB3 and PRPs in the hippocampus. Treatment with an AAV-miR-351-5p inhibitor partially recovered CPEB3 and PRPs. Double luciferase reporter assay results showed that CPEB3 was a direct target of miR-351-5p, while electron microscopy suggested that aluminium could damage mitochondria and synapses in the CA1 of the hippocampus. Golgi staining results indicated that aluminium could reduce the number of dendritic spines in hippocampal neurons. Inhibition of miR-351-5p restored the synaptic structure and growth of dendritic spines in the hippocampus. The involvement of the miR-351-5P/CPEB3/RPPs pathway in aluminium neurotoxicity was confirmed. Our findings suggest that inhibition of miR-351-5p can alleviate learning and memory impairments by increasing CPEB3 and PRPs.

Astaxanthin attenuates cognitive deficits in Alzheimer’s disease models by reducing oxidative stress via the SIRT1/PGC-1α signaling pathway

ObjectiveOxidative stress plays a pivotal role in neurodegenerative diseases. Astaxanthin (AST) can play a neuroprotective role owing to its long-chain conjugated unsaturated double bond, which imparts potent antioxidant, anti-neuroinflammatory, and anti-apoptotic properties. However, the biological mechanisms underlying these effects remain unknown. Therefore, this study aimed to investigate and validate the protective effect of AST on neuronal senescence and apoptosis caused by oxidative stress induced by Aβ25–35 peptide, with the goal of preventing the onset of cognitive dysfunction.MethodsAlzheimer's disease models comprising ICR mice and PC12 cells were established using Aβ25–35. The Morris water maze test was used to assess mouse behavior. Nissl staining revealed morphological changes in the mouse hippocampal neurons. To elucidate the mechanism of action of AST, ICR mice and PC12 cells were treated with the silent information regulator 1 (SIRT1) inhibitor nicotinamide (NAM). Additionally, immunofluorescence, western blotting, and reverse transcription polymerase chain reaction were used to evaluate changes in the expression of Bcl-2 and Bax in the mouse hippocampus, and SIRT1/PGC-1α signaling pathway proteins were detected. Moreover, the oxidative stress markers in ICR mice and PC12 cells were evaluated. Further, CCK-8 assays, Annexin V/PI double staining, and β-galactosidase activity assays were performed in PC12 cells to evaluate the anti-senescence and apoptotic effects of AST.ResultsIn vivo experiments showed that Aβ25–35 impaired cognitive function, promoted morphological changes in hippocampal neurons, decreased Bcl-2 expression, increased Bax expression, decreased superoxide dismutase and GSH-px levels, and increased reactive oxygen species and malondialdehyde levels. Conversely, AST alleviated the impact of Aβ25–35 in mice, with reversed outcomes. NAM administration reduced SIRT1 and PGC-1α expression in the hippocampus. This decrease was accompanied by cognitive dysfunction and hippocampal neuron atrophy, which were also evident in the mice. Additionally, in vitro experiments showed that Aβ25–35 could promote oxidative stress and induce the senescence and apoptosis of PC12 cells. Nonetheless, AST treatment counteracted this effect by inhibiting oxidative stress and altering the state of PC12 cells. Notably, the Aβ + NAM group exhibited the most significant rates of senescence and apoptosis in PC12 cells following NAM treatment.ConclusionAST can improve cellular senescence and apoptosis mediated by oxidative stress via the SIRT1/PGC-1α signaling pathway and plays a vital role in inhibiting neuronal senescence and apoptosis and enhancing cognitive ability.

MiRNA-384-3p alleviates sevoflurane-induced nerve injury by inhibiting Aak1 kinase in neonatal rats.

ObjectiveSevoflurane is a common anesthetic and is widely used in pediatric clinical surgery to induce and maintain anesthesia through inhalation. Increasing studies have revealed that sevoflurane has neurotoxic effects on neurons, apoptosis, and memory impairment. miR‐384 is involved in the process of neurological diseases. However, the role of miRNA‐384‐3p in sevoflurane‐induced nerve injury is not clear. This study focused on exploring the roles and mechanisms of miRNA‐384‐3p in sevoflurane‐induced nerve injury.MethodsSeven‐day‐old rats were exposed to 2.3% sevoflurane to induce nerve injury. The morphological changes in neurons in the hippocampal CA1 region were detected by HE staining and Nissl staining. Neuronal apoptosis was detected by TUNEL and Western blot assays. Spatial memory and learning ability were detected by the Morris water maze assay. The target gene of miRNA‐384‐3p was verified through a luciferase reporter assay. A rescue experiment was used to confirm the miRNA‐384‐3p pathway in sevoflurane‐induced nerve injury.ResultsSevoflurane reduced miRNA‐384‐3p expression in the rat hippocampus. miRNA‐384‐3p alleviated sevoflurane‐induced morphological changes in hippocampal neurons and apoptosis of neurons in the hippocampal CA1 region. Meanwhile, miRNA‐384‐3p attenuated the decline in spatial memory and learning ability induced by sevoflurane. miRNA‐384‐3p alleviated sevoflurane‐induced nerve injury by inhibiting the expression of adaptor‐associated kinase 1 (Aak1).ConclusionOur findings revealed the role and mechanism of miRNA‐384‐3p in sevoflurane‐induced nerve injury, suggesting that miRNA‐384‐3p could be a novel and promising strategy for reducing sevoflurane‐induced neurotoxicity.

Simvastatin Alleviates Vascular Cognitive Impairment Caused by Lacunar Cerebral Infarction Through Protein Kinase B/Nuclear Factor Erythroid 2–Related Factor 2 (AKT/Nrf2) Signaling Pathway

Lacunar cerebral infarction (LACI) is one of the main causes of vascular cognitive impairment (VCI). Herein, this study explored the potential effect of Simvastatin (Sim) on VCI secondary to LACI and Akt/Nrf2 signaling transduction and apoptosis. We established a rat model of VCI and the animals were administered with Sim (40 mg/kg and 80 mg/kg) every day for 28 days. After that, the cognition and memory abilities of rats were assessed together with analysis of morphological changes of hippocampal neurons by immunohistochemistry staining and level of anti-apoptotic related proteins and Akt and Nrf2 signaling proteins by western blot. Compared with normal saline (control group), Sim administration significantly improved the capacity spatial learning and relieved the memory impairment with an improvement in morphological defects. Importantly, Sim treatment restored the p-Akt, t-Nrf2, n-Nrf2 and HO-1 expression along with up-regulation of Bcl-2 and down-regulation of Bax. In conclusion, Sim improves cognitive and morphological disorders induced by LACI possibly through regulating Akt/Nrf2 signaling pathway. These evidence might promote the development of Sim-based treatment for VCI and LACI.

The Effects of Intraoperative Hypothermia on Postoperative Cognitive Function in the Rat Hippocampus and Its Possible Mechanisms.

Intraoperative hypothermia is a common complication during operations and is associated with several adverse events. Postoperative cognitive dysfunction (POCD) and its adverse consequences have drawn increasing attention in recent years. There are currently no relevant studies investigating the correlation between intraoperative hypothermia and POCD. The aim of this study was to assess the effects of intraoperative hypothermia on postoperative cognitive function in rats undergoing exploratory laparotomies and to investigate the possible related mechanisms. We used the Y-maze and Morris Water Maze (MWM) tests to assess the rats’ postoperative spatial working memory, spatial learning, and memory. The morphological changes in hippocampal neurons were examined by haematoxylin-eosin (HE) staining and hippocampal synaptic plasticity-related protein expression. Activity-regulated cytoskeletal-associated protein (Arc), cyclic adenosine monophosphate-response element-binding protein (CREB), S133-phosphorylated CREB (p-CREB [S133]), α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid receptor 1 (AMPAR1), and S831-phosphorylated AMPAR1 (p-AMPAR1 [S831]) were evaluated by Western blotting. Our results suggest a correlation between intraoperative hypothermia and POCD in rats and that intraoperative hypothermia may lead to POCD regarding impairments in spatial working memory, spatial learning, and memory. POCD induced by intraoperative hypothermia might be due to hippocampal neurons damage and decreased expression of synaptic plasticity-related proteins Arc, p-CREB (S133), and p-AMPAR1 (S831).

Constructing Brain Aβ-Targeting Nanoparticles Loaded with EGCG for Treating Alzheimer's Disease in Mice

To construct a nanodelivery system surface-modified with RD2 peptide (polypeptide sequence PTLHTHNRRRRR) for brain tissue penetration and β-amyloid (Aβ) binding. Epigallocatechin-3-gallate (EGCG) was selected for encapsulation in the targeted delivery system and its therapeutic potential for Alzheimer's disease (AD) was investigated. EGCG-load nanoparticles (NP/EGCG), NP/EGCG with RD2 peptide surface modification (RD2-NP/EGCG), as well as RD2 peptide-modified blank nanoparticles (RD2-NP) were prepared and characterized. Thioflavin T assay was done to assess the ability of RD2-NP to bind with Aβ and ex vivo imaging was conducted to evaluate the distribution of RD2-NP in brain lesion sites. The AD mice model was established by injecting oligomeric Aβ 42 in the bilateral hippocampi of ICR mice. Then AD mice were administered intravenously through the tail vein with normal saline, EGCG solution, NP/EGCG or RD2-NP/EGCG for 28 d, respectively, and the Morris water maze tests were performed to assess the spatial memory of mice. Subsequently, RT-PCR method was used to determine the mRNA levels of tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) in the hippocampus of the mice, and the morphological changes of hippocampal neurons were observed with Nissl staining. Additionally, the pathological changes of heart, liver, spleen, lung, and kidney were characterized by hematoxylin-eosin (HE) staining. The particle diameter of the prepared RD2-NP/EGCG was (204.83±2.80) nm and the zeta potential was -23.88 mV. The encapsulation efficiency and drug loading capacity were 94.39% and 5.90%, respectively. The RD2 peptide modification has no significant effect on the physiochemical properties of the nanoparticles. RD2-NP had good Aβ binding ability, and it could be concentrated in hippocampus and cerebral cortex, the most common Aβ deposition sites. The four-week RD2-NP/EGCG treatment significantly decreased the expression of the pro-inflammatory cytokine TNF-α and IL-1β, restored neuronal losses and hippocampal damage, and ameliorated spatial memory impairment in AD model mice. Moreover, treatment with the RD2-NP/EGCG did not present organ toxicity. Surface modified RD2 peptide nanodelivery system can efficiently deliver drugs to AD lesions and improve the therapeutic effect of EGCG on AD.

Vitamin D Alleviates Cognitive Dysfunction by Activating the VDR/ERK1/2 Signaling Pathway in an Alzheimer’s Disease Mouse Model

Objective: Vitamin D (Vit D), a steroid hormone, has been linked to cognitive impairment and dementia, such as Alz­heimer’s disease (AD). 1, 25(OH)₂D₃ is the biologically active form of Vit D, which has been shown to have neuroprotective effects. This compound is being evaluated as an emerging therapeutic treatment in models of AD. Material and Methods: The present study was designed to investigate whether Vit D could alleviate cognitive impairment in an AD rat model by regulating the VDR/ERK1/2 signaling pathway. Adult male APPswe/PS1ΔE9 rats (n = 40) were randomly divided into 2 groups: the AD group and the Vit D + AD group (20 mice per group), and 40 C57BL/6J age-matched mice were separated into the control (CON) group and the Vit D + CON group (20 mice per group). The Morris water maze and object recognition tests were used to evaluate learning and memory functions of the mice. Hematoxylin and eosin staining was used to evaluate morphological changes in hippocampal neurons. Western blotting was used to evaluate the proteins responsible for these changes. Results: We found that Vit D improved learning and memory abilities and morphological defects in hippocampal neurons. Vit D decreased the gene expression of caspase-3 and Bax and increased the expression of Bcl-2. Vit D also increased the protein expression of VDR and p-ERK1 protein in AD mice. Conclusion: This study provides new clues about the mechanism by which Vit D exerts neuroprotective effects in an AD mouse model.

Effect of electroacupuncture on cyclin-dependent kinase 5 and Tau protein in hippocampus of SAMP8 mice

To observe the effect of electroacupuncture (EA) and manual acupuncture (MA) on learning-memory ability, changes of ultrastructure of neurons and expression of CDK5 and Tau proteins in hippocampus of SAMP8 mice，so as to explore its mechanisms underlying improvement of Alzheimer's disease (AD)．. A total of 45 male SAMP8 mice were randomly divided into model, EA and MA groups, with 15 mice in each group. The other 15 SAMR1 mice were used as the normal group. In the EA group, EA (2 Hz, 1 mA) was applied to bilateral "Shenshu"(BL23) and manual acupuncture was applied to "Baihui"(GV20) for 20 min. In the MA group, MA was applied to GV20 and bilateral BL23 for 20 min. Both group were treated once a day for 31 days, and with an interval of one day between every two 7 days. Morris water maze was performed to assess the animals' learning-memory ability. The morphological changes of hippocampal neurons were observed under transmission electron microscopy. The expression levels of CDK5, p25 and Tau-5 proteins in the hippocampus were detected by immunohistochemistry and Western blot, separately. ①Compared with the normal group, the average escape latency of Morris water maze test was prolonged in the model group(P<0.05, P<0.01), duration of swimming in the original platform quadrant and the number of original platform crossing were significantly shorter and less respectively (P<0.01). Compared with the model group, the average escape latency in the EA group was shortened (P<0.05, P <0.01), the duration of swimming in the original platform quadrant and the number of original platform crossing were significantly prolonged and increased (P<0.01); The average escape latency in the MA group was shortened (P<0.05, P <0.01)，and the duration of swimming in the original platform quadrant was prolonged (P<0.05). Compared with the EA group, the average escape latency of the MA group was prolonged (P<0.05), the duration of swimming in the original platform quadrant was shortened(P<0.05). ②Transmission electron microscopy revealed that the neurons in the hippocampal CA1 area had irregular shape and vague structure, reduction in size and number of mitochondria accompanied with swelling, and malformed changes of mitochondrial crest in the model group, which was relatively milder in both EA and MA groups. ③The expression levels of hippocampal Tau-5, p25 and CDK5 proteins were significantly up-regulated in the model group in contrast to the normal group (P<0.01, P<0.05), and obviously down-regulated in both EA and MA groups relevant to the model group (P<0.05, P<0.01). Compared with the EA group, the expression levels of p25 and CDK5 proteins were significantly increased in the MA group (P<0.05). EA of BL23 can improve the learning-memory ability in SAMP8 mice, which is associated with its effect in down-regulating the expression of hippocampal CDK5, p25 and Tau-5 proteins.

ADENOSINE A1 RECEPTOR AGONIST PROTECTS AGAINST HIPPOCAMPAL NEURONAL INJURY AFTER LITHIUM CHLORIDE-PILOCARPINE-INDUCED EPILEPSY.

Adenosine A1 receptor (AA1R) is widely present in the central nervous system, exerting brain protective antiepileptic effects, mainly by binding corresponding G proteins. We evaluated the neuroprotective effects of AA1R on hippocampal neuronal injury after lithium chloride-pilocarpine-induced epilepsy in rats. A total of 60 male SD rats were randomly divided into four groups (n = 15/group): normal control, epilepsy, epilepsy + AA1R antagonist (DPCPX), and epilepsy + AA1R agonist (2-CAdo). An epilepsy model was established through kindling by lithium chloride-pilocarpine. The four groups were observed on days 1, 14, and 30. Pathological and morphological changes of hippocampal neurons were observed by HE staining; apoptosis was detected by TUNEL assay. Caspase-3 and GABA receptor expressions were detected by Western blot. In the hippocampal CA3 area of the epilepsy group, the cellular structure was not neatly arranged, and some neurons were swelling, thick, and incomplete. Compared with the epilepsy group at the same time point, cells in the epilepsy + DPCPX group had an increased distortion, disorganization, edema, cytoplasmic vacuoles, and degeneration. In the epilepsy + 2-CAdo group, cell arrangement was regular and orderly, and structural damages were lessened. Compared with the normal control group at the same time point, the epilepsy group underwent evident neuronal apoptosis, with a significantly higher apoptotic index (AI) (p < 0.05). Compared with the epilepsy group, the neuronal apoptosis of the epilepsy + DPCPX group was boosted, and the AI significantly increased (p < 0.05). The neuronal apoptosis of the epilepsy + 2-CAdo group was inhibited, and the AI significantly decreased (p < 0.05). Compared with the epilepsy group, the caspase-3 expression levels of the epilepsy + DPCPX group on days 14 and 30 were significantly upregulated (p < 0.05), but those of the epilepsy + 2-CAdo group were significantly downregulated (p < 0.05). AA1R abated cell edema and reduced apoptosis, exerting neuroprotective effects on hippocampal neuronal injury after lithium chloride-pilocarpine-induced epilepsy.

Neuroprotective effects of curcumin against rats with focal cerebral ischemia-reperfusion injury.

The aim of the present study was to investigate the protective effects of curcumin and its effect on the methyl ethyl ketone/extracellular signal regulated kinase/cAMP‑response element binding protein (MEK/ERK/CREB) pathway. The study was conducted in vivo and in vitro as follows: In vivo: Focal cerebral ischemia‑reperfusion (IR) models of rats were made with the plug‑line method. Adult male Sprague‑Dawley rats were divided into four groups: Sham operation control group, IR and curcumin‑treatment groups (100 mg/kg and IC, 300 mg/kg). The effects of curcumin on neurological deficit scores, brain water content and infarct volumes were identified. Transmission electron microscope was utilized to observe morphological changes of hippocampal neurons; hematoxylin and eosin staining was used to observe morphological changes of brain tissue; and the terminal deoxynucleotidyl transferase (TdT)‑mediated dUTP nick end labeling method detected neurons apoptosis of hippocampal CA1. Finally, western blot analysis detected the expression of phosphorylated (p)‑MEK, p‑ERK, p‑CREB, B‑cell lymphoma‑2 (Bcl‑2) and Bcl‑2 associated X protein (Bax). In vitro: An oxygen‑glucose deprivation/reoxygenation method was used on primary cultured astrocytes to make cerebral ischemia‑reperfusion models in vitro. Astrocytes were randomly divided into five groups: Normoxia, oxygen‑glucose deprivation/reoxygenation (OGD/Reoxy), OGD/Reoxy + curcumin (5, 10, 20 µmol/l). The cell viability and toxicity were assessed by MTT and lactate dehydrogenase release assay, and levels of p‑MEK, p‑ERK and p‑CREB proteins were analyzed by the western blotting method. Curcumin was demonstrated to improve nerve damage symptoms and infarct volume, reduce brain water content, relieve neuronal apoptosis and also increase the expression of p‑MEK, p‑ERK, p‑CREB, Bcl‑2 and reduce Bax levels in vivo and in vitro. In conclusion curcumin can mitigate focal cerebral ischemia‑reperfusion injuries and this effect may be carried out through the MEK/ERK/CREB pathway.

Effect of inhibition of P38MAPK phosphorylation on learning and memory and autophagy in hippocampus of rats after cerebral ischemic postconditioning

Objective To investigate the effect of ischemic postconditioning on autophagy and learning and memory impairment in rats with ischemia-reperfusion injury by P38MAPK. Methods Ninty-six rats were randomly divided into sham group (Sham group), cerebral ischemia reperfusion group (CIR group), cerebral ischemic postconditioning group (CIP group) and cerebral ischemic postconditioning combined with SB203580 group (CIP + SB203580 group), and 24 rats in each group. The rat model of cerebral ischemia was established by Pulsinelli four-vessel occlusion. The learning and memory abilities of rats were measured by Morris water maze. HE staining were used to detect the morphological changes of hippocampal neurons. The phosphorylation of P38MAPK and Beclin-1, LC3-Ⅱexpression were observed by immunohistochemistry. Results Compared with the Sham group, the number of crossing the platform decreased(24 h: (3.04±0.20)times), and the escape latency was longer in CIR group(24 h: (58.38±1.52) s) (all P<0.05). The number of survival neurons reduced (24 h: 70.93±1.86), and the expression of P38MAPK, LC3-II, Beclin-1 in immunohistochemistry were increased in CIR group(all P<0.05). Compared with CIR group, the number of crossing the platform at each time point increased (24 h: (5.46±0.50)times), the escape latency was shorter (24 h: (52.42±1.53)s), the number of survival neurons increased at each time point(24 h: (83.07±5.30)) and the expression level of P38MAPK decreased in the CIP group, while the expression level of LC3-II, Beclin-1 increased (all P<0.05). Compared with the CIP group, the number of crossing the platform((24 h: (7.13±0.33)times), the escape latency was shorter (24 h: (48.04±1.39)s), the number of survival neurons increased at each time point(24 h: (91.40±1.74)), and the expression of P38MAPK was down-regulated, while the expression of LC3-II, Beclin-1 were up-regulated in CIP + SB203580 group(all P<0.05). Conclusion Ischemic postconditioning can improve learning and memory impairment in rats with ischemia-reperfusion injury by P38MAPK regulating autophagy. Key words: P38MAPK; Ischemic postconditioning; Learning and memory; Autophagy; Rat

Neural basis of major depressive disorder: Beyond monoamine hypothesis.

The monoamine hypothesis has been accepted as the most common hypothesis of major depressive disorder (MDD) for a long period because of its simplicity and understandability. Actually, most currently used antidepressants have been considered to act based on the monoamine hypothesis. However, an important problem of the monoamine hypothesis has been pointed out as follows: it fails to explain the latency of response to antidepressants. In addition, many patients with MDD have remained refractory to currently used antidepressants. Therefore, monoamine-alternate hypotheses are required to explain the latency of response to antidepressants. Such hypotheses have been expected to contribute to identifying hopeful new therapeutic targets for MDD. Past studies have revealed that the volume of the hippocampus is decreased in patients with MDD, which is likely caused by the failure of the hypothalamic-pituitary-adrenal axis and following elevation of glucocorticoids. Two hypotheses have been proposed to explain the volume of the hippocampus: (i) the neuroplasticity hypothesis; and (ii) the neurogenesis hypothesis. The neuroplasticity hypothesis explains how the hippocampal volume is decreased by the morphological changes of hippocampal neurons, such as the shortening length of dendrites and the decreased number and density of spines. The neurogenesis hypothesis explains how the hippocampal volume is decreased by the decrease of neurogenesis in the hippocampal dentate gyrus. These hypotheses are able to explain the latency of response to antidepressants. In this review, we first overview how the neuroplasticity and neurogenesis hypotheses have been developed. We then describe the details of these hypotheses.

Involvement of upregulation of miR-210 in a rat epilepsy model.

Epilepsy is a common type of neurological disorder with complex etiology. The mechanisms are still not clear. MicroRNAs are endogenous noncoding RNAs with many physiological activities. Multiple microRNAs were abnormally expressed in status epilepticus, including miR-210. In this study, we applied lithium chloride and pilocarpine to induce epileptic activity and aimed to disclose the potential mechanisms. Our data showed that miR-210 was significantly upregulated in hippocampus one day after modeling (P<0.05 vs control) and the high expression of miR-210 lasted for at least 30 days. By contrast, γ-aminobutyric acid (GABA) level significantly decreased concurrently after modeling (P<0.05 vs control). To question whether miR-210 could be a potential therapeutic target for epilepsy, miR-210 inhibitor was administrated through intrahippocampal injection after epilepsy modeling. Our data showed that morphological changes of hippocampal neurons and apoptosis triggered by epilepsy were mitigated by miR-210 inhibition. More importantly, the expressions of GABA-related proteins, including GABAA receptor α1, glutamate decarboxylase, and GABA transporter 1, were significantly elevated after epilepsy modeling in both mRNA and protein levels 3 days postmodeling (P<0.05 vs control), which were mitigated by miR-210 inhibitor treatment (P<0.05 vs model). In addition, epilepsy-induced upregulation of GABA transaminase was alleviated by miR-210 inhibitor. Taken together, these data implicated potential roles of miR-210 in lithium chloride–pilocarpine-induced epilepsy model and miR-210 could serve as a potential therapeutic target in status epilepticus.

Structural plasticity in hippocampal cells related to the facilitative effect of intracranial self-stimulation on a spatial memory task.

Posttraining intracranial self-stimulation (SS) in the lateral hypothalamus facilitates the acquisition and retention of several implicit and explicit memory tasks. Here, intracellular injections of Lucifer yellow were used to assess morphological changes in hippocampal neurons that might be specifically related to the facilitative posttraining SS effect upon the acquisition and retention of a distributed spatial task in the Morris water maze. We examined the structure, size and branching complexity of cornus ammonis 1 (CA1) cells, and the spine density of CA1 pyramidal neurons and granular cells of the dentate gyrus (DG). Animals that received SS after each acquisition session performed faster and better than Sham ones--an improvement that was also evident in a probe trial 3 days after the last training session. The neuromorphological analysis revealed an increment in the size and branching complexity in apical CA1 dendritic arborization in SS-treated subjects as compared with Sham animals. Furthermore, increased spine density was observed in the CA1 field in SS animals, whereas no effects were observed in DG cells. Our results support the hypothesis that the facilitating effect of SS on the acquisition and retention of a spatial memory task could be related to structural plasticity in CA1 hippocampal cells.

Improved Survival and Neurological Outcomes after Cardiopulmonary Resuscitation in Toll-like Receptor 4-mutant Mice

Background:Toll-like receptor 4 (TLR4) is a crucial receptor in the innate immune system and noninfectious immune responses. It has been reported that TLR4 participates in the pathological course of ischemia/reperfusion (I/R) injury. However, the role of TLR4 in the process of I/R injury after cardiac arrest (CA) and cardiopulmonary resuscitation (CPR) is still unknown. In this study, we investigated the effects of TLR4 mutation on survival and neurological outcome in a mouse model of CA/CPR.Methods:A model of potassium-induced CA was performed on TLR4-mutant mice (C3H/HeJ) and wild-type mice (C3H/HeN). After 3 min of untreated CA, resuscitation was attempted with chest compression, ventilation, and intravenous epinephrine. Behavioral tests were performed on mice on day 3 after CPR. The morphological changes in hippocampal neurons were assessed by light and electron microscopy. Expressions of TLR4 and intercellular adhesion molecule-1 (ICAM-1) were detected by Western blot. Levels of tumor necrosis factor-α (TNF-α) and myeloperoxidase (MPO) were measured with enzyme-linked immunosorbent assay (ELISA).Results:On day 3 after resuscitation the overall mortality was 33.33% in C3H/HeJ group compared with 53.33% in C3H/HeN group (P < 0.05). And there was much higher central tendency in C3H/HeJ group than C3H/HeN group during open field test (P < 0.05). Meanwhile, the percentage of nonviable neurons was 21.16% in C3H/HeJ group compared with 53.11% in C3H/HeN group (P < 0.05). And there were significantly lower levels of hippocampal TNF-α and MPO in C3H/HeJ mice (TNF-α: 6.85±1.19 ng/mL, MPO: 0.33±0.11 U/g) than C3H/HeN mice (TNF-α: 11.36±2.12 ng/mL, MPO: 0.54±0.17 U/g) (all P < 0.01). CPR also significantly increased the expressions of TLR4 and ICAM-1 in C3H/HeN group. However, the expression of ICAM-1 was much lower in C3H/HeJ group than in C3H/HeN group after CPR (P < 0.01).Conclusion:TLR4 signaling is involved in brain damage and in inflammation triggered by CA/CPR.

Neuritin reverses deficits in murine novel object associative recognition memory caused by exposure to extremely low-frequency (50 Hz) electromagnetic fields.

Animal studies have shown that electromagnetic field exposure may interfere with the activity of brain cells, thereby generating behavioral and cognitive disturbances. However, the underlying mechanisms and possible preventions are still unknown. In this study, we used a mouse model to examine the effects of exposure to extremely low-frequency (50 Hz) electromagnetic fields (ELF MFs) on a recognition memory task and morphological changes of hippocampal neurons. The data showed that ELF MFs exposure (1 mT, 12 h/day) induced a time-dependent deficit in novel object associative recognition memory and also decreased hippocampal dendritic spine density. This effect was observed without corresponding changes in spontaneous locomotor activity and was transient, which has only been seen after exposing mice to ELF MFs for 7-10 days. The over-expression of hippocampal neuritin, an activity-dependent neurotrophic factor, using an adeno-associated virus (AAV) vector significantly increased the neuritin level and dendritic spine density. This increase was paralleled with ELF MFs exposure-induced deficits in recognition memory and reductions of dendritic spine density. Collectively, our study provides evidence for the association between ELF MFs exposure, impairment of recognition memory, and resulting changes in hippocampal dendritic spine density. Neuritin prevented this ELF MFs-exposure-induced effect by increasing the hippocampal spine density.

The Ability of NMDA-Type Glutamate Receptor Blockers to Prevent the Development of Pentylenetetrazole Kindling and Morphological Changes to Pyramidal Neurons in the Mouse Hippocampus

Experiments on mice addressed the link between convulsive syndrome and morphological changes in hippocampal neurons occurring on development of pentylenetetrazole (PTZ) kindling. Kindling was induced by i.p. PTZ (35 mg/kg) three times a week for one month. By the end of this period, 70% of the mice responded to administration of PTZ with severe clonic or clonic-tonic seizures. Hippocampal sections (stratum pyramidale, field CA1, Nissl staining) from convulsive mice showed large numbers of altered cells (24.7 ± 2.1%). Most of these were pyramidal neurons. These hyperchromic neurons had reduced body sizes, loss of turgor, wrinkling of the cell body, and deformation of dendritic processes. These dark-type changes were present in 2.3 ± 2.1% of neurons in the hippocampus of intact mice and mice resistant to the convulsogenic effect of PTZ (30% of the population). Immunohistochemical studies demonstrated normal expression of NeuN (Fox3) protein in all hippocampal cells, including dark hyperchromic neurons. This is evidence that neurons did not die en masse and were relatively viable. Prophylactic s.c. administration of NMDA receptor blockers (0.5 mg/kg memantine, 0.1 mg/kg IEM-1921, or 1 mg/kg IEM-1958) decreased the proportion of mice developing PTZ kindling from 70% to 40%. The proportion of altered neurons in the 60% of mice given NMDA blockers and not developing PTZ kindling or convulsions in the presence of blockers was 0.1 ± 0.06%, which was the same as in intact mice. Conversely, the hippocampus of mice demonstrating convulsions despite simultaneous administration of NMDA blockers showed 24.0 ± 5.6% hyperchromic neurons. These results provide evidence that pathologically altered neurons appeared after convulsive seizures in animals after PTZ kindling and that blockade of NMDA glutamate receptors could weaken both the development of convulsive syndrome and the concomitant morphological changes to hippocampal neurons.

Acute melatonin treatment alters dendritic morphology and circadian clock gene expression in the hippocampus of Siberian hamsters.

In the hippocampus of Siberian hamsters, dendritic length and dendritic complexity increase in the CA1 region whereas dendritic spine density decreases in the dentate gyrus region at night. However, the underlying mechanism of the diurnal rhythmicity in hippocampal neuronal remodeling is unknown. In mammals, most daily rhythms in physiology and behaviors are regulated by a network of circadian clocks. The central clock, located in the hypothalamus, controls melatonin secretion at night and melatonin modifies peripheral clocks by altering expression of circadian clock genes. In this study, we examined the effects of acute melatonin treatment on the circadian clock system as well as on morphological changes of hippocampal neurons. Male Siberian hamsters were injected with melatonin in the afternoon; 4 h later, mRNA levels of hypothalamic and hippocampal circadian clock genes and hippocampal neuron dendritic morphology were assessed. In the hypothalamus, melatonin treatment did not alter Period1 and Bmal1 expression. However, melatonin treatment increased both Period1 and Bmal1 expression in the hippocampus, suggesting that melatonin affected molecular oscillations in the hippocampus. Melatonin treatment also induced rapid remodeling of hippocampal neurons; melatonin increased apical dendritic length and dendritic complexity in the CA1 region and reduced the dendritic spine density in the dentate gyrus region. These data suggest that structural changes in hippocampal neurons are regulated by a circadian clock and that melatonin functions as a nighttime signal to coordinate the diurnal rhythm in neuronal remodeling.