Protective effects of docosahexaenoic acid supplementation on cognitive dysfunction and hippocampal synaptic plasticity impairment induced by early postnatal PM2.5 exposure in young rats.

PM2.5 exposure is a challenging environmental issue that is closely related to cognitive development impairment; however, currently, relevant means for prevention and treatment remain lacking. Herein, we determined the preventive effect of docosahexaenoic acid (DHA) supplementation on the neurodevelopmental toxicity induced by PM2.5 exposure. Neonatal rats were divided randomly into three groups: control, PM2.5, and DHA + PM2.5 groups. DHA could ameliorate PM2.5-induced learning and memory dysfunction, as well as reverse the impairment of hippocampal synaptic plasticity, evidenced by enhanced long-term potentiation, recovered synaptic ultrastructure, and increased expression of synaptic proteins. Moreover, DHA increased CREB phosphorylation and BDNF levels and attenuated neuroinflammation and oxidative stress, reflected by lower levels of IBA-1, IL-1ß, and IL-6 and increased levels of SOD1 and Nrf2. In summary, our findings demonstrated that supplementation of DHA effectively mitigated the cognitive dysfunction and synaptic plasticity impairment induced by early postnatal exposure to PM2.5. These beneficial effects may be attributed to the upregulation of the CREB/BDNF signaling pathway, as well as the reduction of neuroinflammation and oxidative stress.

A peptide from the Japanese encephalitis virus failed to induce the production of anti-N-methyl-d-aspartate receptor antibodies via molecular mimicry in mice.

Background: The development of anti-N-methyl-d-aspartate receptor (NMDAR) encephalitis following viral encephalitis, such as Japanese encephalitis, has received increasing attention in recent years. However, the mechanism of anti-NMDAR antibody production following Japanese encephalitis has not been explored. Methods: A peptide from the Japanese encephalitis virus (JEV), which shares a similar amino acid sequence with GluN1, was identified by sequence comparison. We then explored whether active subcutaneous immunization with the JEV peptide could induce the production of anti-NMDAR antibodies and related pathophysiological and behavioral changes in mice. In addition, a published active immune model of anti-NMDAR encephalitis using a GluN1 peptide was used as the positive control. Results: A 6-amino-acid sequence with 83 % similarity between the envelope protein of the JEV (HGTVVI) and GluN1 (NGTHVI) was identified, and the sequence included the N368/G369 region. Active immunization with the JEV peptide induced a substantial and specific immune response in mice. However, anti-NMDAR antibodies were not detected in the serum of mice immunized with the JEV peptide by ELISA, CBA, and TBA. Moreover, mice immunized with the JEV peptide presented no abnormities related to anti-NMDAR antibodies according to western blotting, patch clamp, and a series of behavioral tests. In addition, active immunization with a recently reported GluN1 peptide failed to induce anti-NMDAR antibody production in mice. Conclusions: In this study, the attempt of active immunization with the JEV peptide to induce the production of anti-NMDAR antibodies via molecular mimicry failed. The pathogenesis of anti-NMDAR encephalitis following Japanese encephalitis remains to be elucidated.

Dynamic effects of miR-20a-5p on hippocampal ripple energy after status epilepticus in rats.

To determine the dynamic effects of miR-20a-5p on hippocampal ripple energy in rats after status epilepticus (SE). A lithium pilocarpine (LiCl-PILO)-induced rat model of status epilepticus (SE) was established, and the rats were divided into the normal control (Control, CTL), epileptic control (PILO), valproic acid (VPA + PILO), miR-20a-5p overexpression lentivirus vector (miR + PILO), sponges blocking lentivirus vector (Sponges + PILO), and scramble sequence negative control (Scramble + PILO) groups (n = 6). Electroencephalograms (EEGs) were used to analyze changes in hippocampal ripple energy before and after SE. Quantitative polymerase chain reaction (q-PCR) analysis showed that miR-20a-5p levels gradually increased after miR-20a-5p overexpression lentivirus vector injection into the lateral ventricle, and the miR-20a-5p levels were significantly higher than that in CTL group on days 7 and 36 (P < 0.001). The miR-20a-5p levels decreased significantly on days 7 and 36 after blocking by sponges lentivirus vector injected into the lateral ventricle (P < 0.001). After injection of PILO, the average ripple energy expression in each group gradually increased, and reached the peak before chloral hydrate injection (compared with 1 day before SE, P < 0.05). The ripple energy in the VPA + PILO and Sponges + PILO groups was significantly lower than that in the PILO group at 60 min and 70 min after PILO injection and before chloral hydrate injection (P < 0.05), and maintained lower until 2 h after chloral hydrate injection in VPA + PILO (P < 0.05). Compared with the VPA + PILO group, the mean ripple energy of the Sponges + PILO group had no difference at all time points (P ≥ 0.05). After SE, ripple distribution of space and energy is closely related to the occurrence of epilepsy. Inhibition of miR20a-5p expression can downregulate ripple oscillation energy during seizure.

The dysfunctionality of hippocampal synapses may be directly related to PM-induced impairments in spatial learning and memory in juvenile rats.

Epidemiological studies have demonstrated that exposure to air particulate matter (PM) increases the incidence of cardiovascular and respiratory diseases and exerts a significant neurotoxic effect on the nervous system, especially on the immature nervous system. Here, we selected PND28 rats to simulate the immature nervous system of young children and used neurobehavioral methods to examine how exposure to PM affected spatial learning and memory, as well as electrophysiology, molecular biology, and bioinformatics to study the morphology of hippocampus and the function of hippocampal synapses. We discovered that spatial learning and memory were impaired in rats exposed to PM. The morphology and structure of the hippocampus were altered in the PM group. In addition, after exposure to PM, the relative expression of synaptophysin (SYP) and postsynaptic density 95 (PSD95) proteins decreased dramatically in rats. Furthermore, PM exposure impaired long-term potentiation (LTP) in the hippocampal Schaffer-CA1 pathway. Interestingly, RNA sequencing and bioinformatics analysis revealed that the differentially expressed genes (DEGs) were rich in terms associated with synaptic function. Five hub genes (Agt, Camk2a, Grin2a, Snca, and Syngap1) that may play a significant role in the dysfunctionality of hippocampal synapses were identified. Our findings implied that exposure to PM impaired spatial learning and memory via exerting impacts on the dysfunctionality of hippocampal synapses in juvenile rats and that Agt, Camk2a, Grin2a, Snca, and Syngap1 may drive PM-caused synaptic dysfunction.

Severe epilepsy phenotype with SCN1A missense variants located outside the sodium channel core region: Relationship between functional results and clinical phenotype.

PURPOSE: Most SCN1A missense variants located outside the sodium channel core region show a mild phenotype. However, there are exceptions, because of which it is challenging to determine the correlation between genotype and phenotype. In this study, we aimed to determine whether functional study could be used to determine disease severity in cases with such variants, and elucidate possible genotype-phenotype relationships. METHODS: Forty-seven patients with SCN1A missense variants were recruited, and one with a Dravet syndrome phenotype with an SCN1A missense variant (c.3811T>C/ p.W1271R) located outside the core region was screened with electrophysiological tests. We also reviewed functional SCN1A studies on patients with inconsistent phenotypes and genotypes, and studied the relationship between electrophysiological measurements and clinical phenotype. RESULTS: Patch clamp experiments showed that the W1271R variant caused significantly reduced sodium current, decreased channel voltage sensitivity, loss of channel availability, and prolonged recovery time from inactivation compared with wild type (WT), which ultimately caused a change in loss of function (LOF). Twelve cases of severe SCN1A-related epilepsy with missense variants located outside the channel core region were also included from the functional studies. Nine patients with missense SCN1A variants showed complete (3/9) or partial (6/9) physiological LOF. Two missense SCN1A variants caused physiological gain-and-loss of function (G-LOF), and one caused decreased excitability (DE). CONCLUSIONS: Not all missense variants located outside the core region cause a mild phenotype. Although current functional studies in heterologous expression systems do not accurately reflect disease severity caused by SCN1A missense variants, they could be an effective model for generation of data to study the initial effects of SCN1A missense variants.

Anticonvulsant Effect of Carbenoxolone on Chronic Epileptic Rats and Its Mechanism Related to Connexin and High-Frequency Oscillations.

Objective: This study was designed to investigate the influence and mechanism of gap junction carbenoxolone (CBX) on dynamic changes in the spectral power of ripples and fast ripples (FRs) in the hippocampus of chronic epileptic rats. Methods: The lithium-pilocarpine (PILO) status epilepticus (SE) model (PILO group) and the CBX pretreatment model (CBX + PILO group) were established to analyze dynamic changes in the spectral power of ripples and FRs, and the dynamic expression of connexin (CX)26, CX32, CX36, and CX43 in the hippocampus of chronic epileptic rats. Results: Within 28 days after SE, the number of spontaneous recurrent seizures (SRSs) in the PILO group was significantly higher than that in the CBX + PILO group. The average spectral power of FRs in the PILO group was significantly higher than the baseline level at 1 and 7 days after SE. The average spectral power of FRs in the PILO group was significantly higher than that in the CBX + PILO group at 1, 7, and 14 days after SE. Seizures induced an increase in CX43 expression at 1 and 7 days after SE, but had no significant effect on CX26, CX36, or CX32. CBX pretreatment did not affect the expression of CXs in the hippocampus of normal rats, but it inhibited the expression of CX43 in epileptic rats. The number of SRSs at 2 and 4 weeks after SE had the highest correlation with the average spectral power of FRs; the average spectral power of FRs was moderately correlated with the expression of CX43. Conclusion: The results of this study indicate that the energy of FRs may be regulated by its interference with the expression of CX43, and thus, affect seizures. Blocking the expression of CX43 thereby reduces the formation of pathological high-frequency oscillations (HFOs), making it a promising strategy for the treatment of chronic epilepsy.

The Role and Mechanism of AMIGO3 in the Formation of Aberrant Neural Circuits After Status Convulsion in Immature Mice.

Leucine rich repeat and immunoglobulin-like domain-containing protein 1 (Lingo-1) has gained considerable interest as a potential therapy for demyelinating diseases since it inhibits axonal regeneration and myelin production. However, the results of clinical trials targeted at Lingo-1 have been unsatisfactory. Amphoterin-induced gene and open reading frame-3 (AMIGO3), which is an analog of Lingo-1, might be an alternative therapeutic target for brain damage. In the present study, we investigated the effects of AMIGO3 on neural circuits in immature mice after status convulsion (SC) induced by kainic acid. The expression of both AMIGO3 and Lingo-1 was significantly increased after SC, with levels maintained to 20 days after SC. Following SC, transmission electron microscopy revealed the impaired microstructure of myelin sheaths and Western blot analysis showed a decrease in myelin basic protein expression, and this damage was alleviated by downregulation of AMIGO3 expression. The ROCK/RhoA signaling pathway was inhibited at 20 days after SC by downregulating AMIGO3 expression. These results indicate that AMIGO3 plays important roles in seizure-induced damage of myelin sheaths as well as axon growth and synaptic plasticity via the ROCK/RhoA signaling pathway.

Effect of gap junction blockers on hippocampal ripple energy expression in rats with status epilepticus. / ç¼éš™è¿žæŽ¥é˜»æ–­å‰‚å¯¹å¤§é¼ ç™«ç—«åŽæµ·é©¬æ¶Ÿæ³¢æŒ¯è¡èƒ½é‡å˜åŒ–çš„å½±å“.

OBJECTIVES: To study the effect of gap junction blockers, quinine (QUIN) and carbenoxolone (CBX), on hippocampal ripple energy expression in rats with status epilepticus (SE). METHODS: A total of 24 rats were randomly divided into four groups: model, QUIN, valproic acid (VPA), and CBX (n=6 each). A rat model of SE induced by lithium-pilocarpine (PILO) was prepared. The QUIN, VPA, and CBX groups were given intraperitoneal injection of QUIN (50 mg/kg), VPA by gavage (200 mg/kg), and intraperitoneal injection of CBX (50 mg/kg) respectively, at 3 days before PILO injection. Electroencephalography was used to analyze the change in hippocampal ripple energy before and after modeling, as well as before and after chloral hydrate injection to control seizures. RESULTS: Ripple expression was observed in the hippocampal CA1, CA3, and dentate gyrus regions of normal rats. After 10 minutes of PILO injection, all groups had a gradual increase in mean ripple energy expression compared with 1 day before modeling, with the highest expression level before chloral hydrate injection in the model, VPA and CBX groups (P<0.05). The QUIN group had the highest expression level of mean ripple energy 60 minutes after PILO injection. The mean ripple energy returned to normal levels in the three intervention groups immediately after chloral hydrate injection, while in the model group, the mean ripple energy returned to normal levels 1 hour after chloral hydrate injection. The mean ripple energy remained normal till to day 3 after SE in the four groups. The changing trend of maximum ripple energy was similar to that of mean ripple energy. CONCLUSIONS: The change in ripple energy can be used as a quantitative indicator for early warning of seizures, while it cannot predict seizures in the interictal period. Gap junction blockers can reduce ripple energy during seizures.

Knockdown of Lingo-1 by short hairpin RNA promotes cognitive function recovery in a status convulsion model.

The purpose of this study was to determine the dynamic changes of the Nogo-66 receptor 1 (NgR1) pathway during epileptogenesis and the potential beneficial of leucine-rich repeat and Ig-like domain-containing Nogo receptor interacting protein 1 (Lingo-1) inhibition on epilepsy rats. The hippocampal changes of the NgR1 pathway during epileptogenesis were determined by western blot analysis of multiple proteins, including neurite outgrowth inhibitor protein A (NogoA), myelin-associated glycoprotein (MAG), oligodendrocyte-myelin glycoprotein (OMgp), Lingo-1, ras homolog family member A (RhoA) and phosphorylated RhoA (p-RhoA). Lentivirus-mediated short hairpin RNA (shRNA) was used to knockdown the hippocampal expression of Lingo-1. Novel object recognition (NOR) test and Morris Water Maze (MWM) test were employed to determine the cognitive functions of rats. Hematoxylin and eosin (H&E) staining, protein expressions of RhoA, p-RhoA, and myelin basic protein (MBP), as well as convulsion susceptibility test were additionally performed. Our results showed that the NgR1 pathway was activated during epileptogenesis, characterized by up-regulation of NogoA, MAG, OMgp, and Lingo-1, which was especially significant at the chronic phase of epilepsy. The cognitive function, convulsion susceptibility and hippocampal neuronal survival of rats were impaired at the chronic phase of epileptogenesis but all improved by Lingo-1 inhibition; besides, the hippocampal protein expressions of p-RhoA and MBP were significantly decreased at the chronic phase of SC rats but increased after Lingo-1 inhibition. Our results demonstrated that Lingo-1 shRNA can improve epilepsy-induced cognitive impairment, which may be related with the pro-myelination and neuroprotection effects of Lingo-1 inhibition.

Effect of sodium butyrate regulating IRAK1 (interleukin-1 receptor-associated kinase 1) on visceral hypersensitivity in irritable bowel syndrome and its mechanism.

The current study aimed to investigate the effects of sodium butyrate on the level of colonic protein IRAK1 (interleukin-1 receptor-associated kinase 1) in irritable bowel syndrome (IBS) models as well as revealing the relationship between IRAKI level and visceral sensitivity during the progression of IBS. IBS symptoms were induced using TNBS (2,4,6-trinitrobenzene sulfonic acid) in mice and using IL-33 in HT-29 cells, which were then hanlded with sodium butyrate (100 mM for each mice and 0.05 M for HT-29 cells). The threshold of visceral pain and the expression of IRAKI in mice, and the level of IRAKI in HT-29 cells were detected. The data showed that the level of IRAK1 in IBS mice was higher than that in the control group, while the pre-treatment with sodium butyrate could solidy suppressed the level of IRAK1. Morevoer, it was found that the level of IRAK1 was negatively correlated with the pain threshold. In in vitro assays, the level of IRAK1 was firstly induced by IL-33 stimulation and then suppressed by sodium butyrate pretreatment. Collectively, the level of IRAKI showed an obvioulty positive relation with visceral hypersensitivity in IBS models, and the treatment with sodium butyrate could alleviate visceral hypersensitivity by inhibiting the expression of IRAKI.

Autoimmune encephalitis after Japanese encephalitis in children: A prospective study.

OBJECTIVE: To explore anti-neuronal surface antibodies and identify associated serum predictors of autoimmune encephalitis after Japanese encephalitis (JE). METHODS: This prospective study first detected anti-neuronal surface antibodies and cytokines in the serum and cerebrospinal fluid (CSF) of JE patients within one week of symptom onset. Anti-neuronal surface antibodies and cytokines in the serum were detected on day 21 post-JE. If the patients relapsed during the convalescent phase, we simultaneously detected JE virus RNA and cytokines in the CSF, as well as anti-neuronal surface antibodies in the serum and CSF. RESULTS: All 31 patients were negative for anti-neuronal surface antibodies at the onset of JE in the serum and CSF. During the convalescent phase, five patients developed autoimmune encephalitis (two had anti-N-methyl-d-aspartate receptor [NMDAR] antibodies, one had γ-aminobutyric acid-B receptor [GABABR] antibodies, and two had other antibodies against unknown neuronal surface antigens). Patients who developed autoimmune encephalitis experienced more severe outcomes than those who did not at the one-year follow-up (p = 0.044). The levels of serum CXCL13 and IL-6, as well as CXCL13, BAFF, CXCL10, and MMP-9 in the CSF were increased in the convalescent phase compared to the acute phase in patients who developed autoimmune encephalitis (p < 0.05). CONCLUSION: In addition to anti-NMDAR antibodies, anti-GABABR antibodies and antibodies against unknown neuronal surface antigens can trigger autoimmune encephalitis following JE. Patients who developed autoimmune encephalitis had a poorer prognosis at the one-year follow-up. Serum CXCL13 may represent a predictor of autoimmune encephalitis after JE.

Role of PKA/CREB/BDNF signaling in PM2.5-induced neurodevelopmental damage to the hippocampal neurons of rats.

Exposure to fine particulate matter (PM2.5) is implicated in neurodevelopmental disorders including cognitive decline, attention-deficit/hyperactivity disorder, and autism spectrum disorder. However, the specific molecular mechanisms by which PM2.5 impacts neurodevelopment are poorly understood. Accordingly, in the present study, the role of protein kinase A (PKA)/cAMP response element binding protein (CREB)/brain-derived neurotrophic factor (BDNF) signaling in PM2.5-induced neurodevelopmental damage was investigated using primary cultured hippocampal neurons. When hippocampal neurons cultured for 3 days in vitro (DIV3) were exposed to PM2.5 for 24 h and 96 h, neuronal viability decreased by 18.8% and 32.7% respectively, percentage of TUNEL-positive neurons increased by 78.5% and 64.0% separately, caspase-9 expression increased, lower postsynaptic density and shorter active zones were observed by transmission electron microscopy, expression of synapse-related proteins including postsynaptic density-95 (PSD95), growth associated protein-43 (GAP43), and synaptophysin (SYP) were decreased, and the phosphorylation levels of PKA, CREB, and BDNF expression also decreased. However, the PM2.5-induced neuronal damage could be ameliorated or aggravated to varying degrees by up- or down-regulation of the PKA/CREB/BDNF signaling pathway, respectively. Our results indicate that PM2.5 exposure exerts neurodevelopmental toxicity as indicated by lower viability, apoptosis, and synaptic damage in primary cultured hippocampal neurons, and that the PKA/CREB/BDNF pathways could play a vital role in PM2.5-mediated neurodevelopmental toxicity.

VEGF Modulates Neurogenesis and Microvascular Remodeling in Epileptogenesis After Status Epilepticus in Immature Rats.

Neurogenesis and angiogenesis are widely recognized to occur during epileptogenesis and important in brain development. Because vascular endothelial growth factor (VEGF) is a critical neurovascular target in neurological diseases, its effect on neurogenesis, microvascular remodeling and epileptogenesis in the immature brain after lithium-pilocarpine-induced status epilepticus (SE) was investigated. The dynamic changes in and the correlation between hippocampal neurogenesis and microvascular remodeling after SE and the influence of VEGF or SU5416 injection into the lateral ventricles at different stages after SE on neurogenesis and microvascular remodeling through regulation of VEGF expression were assessed by immunofluorescence and immunohistochemistry. Western blot analysis revealed that the VEGFR2 signaling pathway promotes phosphorylated ERK and phosphorylated AKT expression. The effects of VEGF expression regulation at different stages after SE on pathological changes in hippocampal structure and spontaneous recurrent seizures (SRS) were evaluated by Nissl staining and electroencephalography (EEG). The results showed that hippocampal neurogenesis after SE is related to microvascular regeneration. VEGF promotion in the acute period and inhibition in the latent period after SE alleviates loss of hippocampal neuron, abnormal vascular regeneration and inhibits neural stem cells (NSCs) ectopic migration, which may effectively alleviate SRS severity. Interfering with VEGF via the AKT and ERK pathways in different phases after SE may be a promising strategy for treating and preventing epilepsy in children.

Dynamic alteration of dendrites and dendritic spines in the hippocampus and microglia in mouse brain tissues after kainate-induced status epilepticus.

PURPOSE: To study the alteration of microglial subtypes, the representative markers of microglia, and the morphology of dendrites and dendritic spines after acute status epilepticus (SE) and during recurrent seizures. METHODS: A mouse kainate-induced SE model was used. Dendrites and dendritic spines of granule neurons in the dentate gyrus (DG) subregion and pyramidal neurons in the cornu ammonis (CA)1 and cornu ammonis (CA)3 subregions of the hippocampus were visualized by Golgi staining. Synaptic proteins were evaluated by Western blot analysis, and microglia and their markers were evaluated by flow cytometry. RESULTS: Extensive partial spine loss was observed in the dendrites of granule and pyramidal cells in the acute and early chronic stages of SE. In terms of spine loss, the thin and mushroom types predominated. Accompanying the spine loss in these two stages, the proportion of M1 microglia increased significantly with high CX3CR1 expression and low CD200R expression. However, at the transiting stage, the proportion of M2 microglia was increased dramatically, and high expression levels of CXCR3 on all microglia and CD68 on M1 microglia were observed. Morris water maze tests revealed significant learning and memory impairment in the chronic phase of epilepsy. CONCLUSION: Dendritic spines in the hippocampus and microglia in the central nevus system are dynamically altered in epilepsy during the establishment and maintenance of spontaneous seizures. Microglia may contribute to the spine loss and related learning and memory impairment.

[Effect of advanced maternal age on development of hippocampal neural stem cells in offspring rats].

OBJECTIVE: To study the effect of advanced maternal age (AMA) on the development of hippocampal neural stem cells in offspring rats. METHODS: Ten 3-month-old and ten 12-month-old female Sprague-Dawley rats were housed individually with 3-month-old male rats (1:1, n=20), whose offspring rats were assigned to a control group and an AMA group. A total of 40 rats were randomly selected from each group. Immunofluorescence assay and Western blot were used to localize and determine the levels of protein expression of Nestin and doublecortin (DCX) on day 7 as well as neuronal nuclear antigen (NeuN) and glial fibrillary acidic protein (GFAP) on day 28 (n=8 for each marker). Immunofluorescence assay was also used to localize the hippocampal expression of polysialylated isoforms of neural cell adhesion molecule (PSA-NCAM) on day 14 (n=8 for each marker). RESULTS: According to the Western blot results, the AMA group had significantly lower protein expression of DCX than the control group (P<0.05), while there were no significant differences in the protein expression of Nestin, NeuN, and GFAP between the two groups (P>0.05). According to the results of immunofluorescence assay, the AMA group had significantly lower protein expression of Nestin, DCX, and PSA-NCAM in the hippocampal dentate gyrus (DG) region than the control group (P<0.05), while there were no significant differences in the above indices in the hippocampal CA1 and CA3 regions between the two groups (P>0.05). The AMA group had significantly higher expression of NeuN in the hippocampal CA1 region than the control group (P<0.01), while there were no significant differences in the expression of NeuN in the hippocampal DG and CA3 regions between the two groups (P>0.05). The AMA group had significantly lower expression of GFAP in the hippocampal CA1, CA3, and DG regions than the control group (P<0.05). CONCLUSIONS: AMA may cause inhibition of proliferation, survival, and migration of hippocampal neural stem cells. AMA may also affect their differentiation into neurons and astrocytes, which will eventually lead to developmental disorders of hippocampal neural stem cells in offspring rats.

[Toxicity of dibutyl phthalate in primary cultured rat hippocampal neurons and the toxicological mechanism].

OBJECTIVE: To investigate the neurotoxicity and toxicological mechanism of dibutyl phthalate (DBP) in primary cultured rat hippocampal neurons. METHODS: Primary rat hippocampal neurons cultured for 4 days were exposed to 1 g/L DBP for 24, 48, or 96 h. Immunofluorescence assay and transmission electron microscopy (TEM) were used to observe the morphological changes of the axons and the ultrastructure of DBP-treated neurons. The action potential (AP) of the hippocampal neurons was measured with patch-clamp electrophysiology. CCK-8 assay was used to detect the viability of the hippocampal neurons, and Western blotting was performed to determine the mRNA and protein expressions of brain-derived neurotrophic factor (BDNF), neuropeptide Y (NPY) and estrogen receptor ß (ERß). High-performance liquid chromatography-tandem mass spectrometry (HPLC-MS) was employed to detect the release of the neurotransmitter GABA. RESULTS: After exposure to DBP for 96 h, the cellular network of the hippocampal neurons became sparse, and the neurons showed significantly decreased axonal length (P < 0.01) and presented with round cell nuclei, chromatin aggregation and cytoplasmic vacuolization. Patch-clamp electrophysiology revealed depolarization drift and increased frequency of discharge in the exposed neurons (P < 0.01). The neurons with DBP exposure for 24, 48 and 96 h all showed significantly decreased cell viability (P < 0.01). DBP exposure for 48 and 96 h significantly lowered the protein expressions of ERß, BDNF and NPY, and a 96-h exposure significantly reduced the release of the neurotransmitter GABA in the neurons (P < 0.05). CONCLUSIONS: DBP exposure causes morphological and functional damages of primary cultured rat hippocampal neurons. DBP-induced neurotoxicity is probably associated with GABA-mediated blockage of the ERß-BDNF-NPY signaling communication.

TRPV1 Contributes to the Neuroprotective Effect of Dexmedetomidine in Pilocarpine-Induced Status Epilepticus Juvenile Rats.

To investigate the antiepileptic and neuroprotective effects of dexmedetomidine (Dex) in pilocarpine- (Pilo-) induced status epilepticus (SE) juvenile rats, rats were randomly assigned to the following six groups (n = 20): normal, normal+Dex, SE, SE+Cap, SE+Dex, and SE+Dex+Cap. The rats were treated with either diazepam (i.p., an antiepileptic drug) or Dex after the onset of SE. The Morris water maze was used to assess rat cognitive behavior. Flow cytometry was used to detect the concentrations of Ca2+, mitochondrial membrane potential, and reactive oxygen species. Transmission electron microscopy was performed to evaluate specimens of brain tissue. The levels of caspase 3 and TRPV1 were examined by western blot and immunohistochemistry (IHC). Treatment with Dex significantly decreased the escape latency of the SE rats (P < 0.05). Capsaicin, a TRPV1 agonist, delivery aggravated the performance of SE rats. Pathological changes in SE rat were attenuated by Dex and deteriorated by capsaicin. Swollen mitochondria and abnormal endoplasmic reticulum were found in SE rats and were then aggravated by capsaicin and reversed by Dex. Moreover, our data showed that Dex significantly restrained calcium overload, ROS production, and mitochondrial membrane potential loss, all of which were induced by Pilo and capsaicin (P < 0.05). Dex decreased the apoptotic rate in the Model SE group (P < 0.05) and TRPV1 and caspase 3 expression in the Dex treatment group (P < 0.05). Interestingly, all these effects of Dex were partially counteracted by the TRPV1 agonist, capsaicin (P < 0.05). Our study showed that Dex exerted a neuroprotective effect in Pilo-induced SE rats by inhibiting TRPV1 expression and provided information for therapy to SE patients.

The implications of hippocampal neurogenesis in adolescent rats after status epilepticus: a novel role of notch signaling pathway in regulating epileptogenesis.

Seizure-induced neurogenesis has a widely recognized pro-epileptogenic function. Given the critical role of Notch signaling during the maintenance and neurogenesis of neural stem cells, we hypothesized that Notch may affect epileptogenesis and its progression through its role in neurogenesis in the adolescent rat brain. We used the lithium-pilocarpine-induced epilepsy model in adolescent Sprague-Dawley rats in order to evaluate hippocampal neurogenesis and epileptogenesis following the onset of status epilepticus (SE). We used western blotting analyses and qPCR to measure levels of Notch signaling at different phases after seizures and immunofluorescence to detect the proliferation and differentiation of neural stem cells after seizure. Following the administration of DAPT, a Notch γ-secretase inhibitor, into the lateral ventricles, we observed a suppression of abnormal neurogenesis in the acute phase and a reduction of gliosis in the chronic phase after SE. Accordingly, the frequency and duration of spontaneous seizures in chronic phase were decreased. Our results clarify the basic concept regarding the involvement of Notch signaling in the regulation of hippocampal neurogenesis and epileptogenesis, thereby potentially offering a novel and alternative treatment for epilepsy.

Effects of early postnatal exposure to fine particulate matter on emotional and cognitive development and structural synaptic plasticity in immature and mature rats.

INTRODUCTION: Fine particulate matter (PM2.5) is closely associated with many neurological disorders including neurodegenerative disease, stroke, and brain tumors. However, the toxic effects of PM2.5 on neurodevelopment remain unclear. In this study, we aimed to determine the neurotoxic effects of early postnatal exposure to PM2.5 in immature and mature rats. METHODS: We exposed neonatal rats to PM2.5 (2 or 10 mg/kg body weight) through intranasal instillation from postnatal day (PND) 3-15, once a day. Emotional and cognitive development were evaluated using the elevated plus maze, forced swimming, and Morris water maze tests. Hippocampal tissue was collected and subjected to transmission electron microscopy observation and western blot analysis. RESULTS: Rats had lower body weight after exposure to high dose of PM2.5. The behavioral test results indicated that high-dose PM2.5 exposure led to increased anxiety-like symptoms in immature and mature rats, apparent depressive-like behaviors in mature rats, and impaired spatial learning and memory abilities in immature rats, and low-dose PM2.5 exposure increased anxiety-like behaviors in immature rats. Further, high-dose PM2.5 exposure contributed to fewer synapses, thinner postsynaptic density, and shorter active zone in immature and mature rats, and also decreased expressions of synaptophysin (SYP), growth associated protein-43 (GAP43), and postsynaptic density-95 (PSD95) in immature rats, SYP and PSD95 in mature rats. Moreover, low-dose PM2.5 exposure diminished the expression of PSD95 in immature rats. In addition, high-dose PM2.5 exposure reduced brain-derived neurotrophic factor (BDNF) expression and cAMP response element binding protein (CREB) phosphorylation in both immature and mature rats, and low-dose PM2.5 exposure lessened BDNF expression and CREB phosphorylation in immature rats. CONCLUSIONS: Our findings indicate that PM2.5 impairs emotional and cognitive development by disrupting structural synaptic plasticity, possibly via the CREB/BDNF signaling pathway.

Levetiracetam Protects Against Cognitive Impairment of Subthreshold Convulsant Discharge Model Rats by Activating Protein Kinase C (PKC)-Growth-Associated Protein 43 (GAP-43)-Calmodulin-Dependent Protein Kinase (CaMK) Signal Transduction Pathway.

BACKGROUND Subclinical epileptiform discharges (SEDs) are defined as epileptiform electroencephalographic (EEG) discharges without clinical signs of seizure in patients. The subthreshold convulsant discharge (SCD) is a frequently used model for SEDs. This study aimed to investigate the effect of levetiracetam (LEV), an anti-convulsant drug, on cognitive impairment of SCD model rats and to assess the associated mechanisms. MATERIAL AND METHODS A SCD rat model was established. Rats were divided into an SCD group, an SCD+ sodium valproate (VPA) group, and an SCD+ levetiracetam (LEV) group. The Morris water maze was used to evaluate the capacity of positioning navigation and space exploration. The field excitatory post-synaptic potentials (fEPSPs) were evaluated using a bipolar stimulation electrode. NCAM, GAP43, PS95, and CaMK II levels were detected using Western blot and RT-PCR, respectively. PKC activity was examined by a non-radioactive method. RESULTS LEV shortens the latency of platform seeking in SCD rats in positioning navigation. fEPSP slopes were significantly lower in the SCD group, and LEV treatment significantly enhanced the fEPSP slopes compared to the SCD group (P<0.05). The NCAM and GAP-43 levels were increased and PSD-95 levels were increased in SCD rats (P<0.05), which were improved by LEV treatment. The PKC activity and CaMK II levels were decreased in SCD rats and LEV treatment significantly enhanced PKC activity and increased CaMK II levels. CONCLUSIONS Cognitive impairment in of SCD model rats may be caused by decreased PKC activity, low expression of CaMK II, and inhibition of LTP formation. LEV can improve cognitive function by activating the PKC-GAP-43-CaMK signal transduction pathway.