Expression of AMPA receptor subunits in hippocampus after status convulsion

Objective

The expression of 2-amino-3-(5-methyl-3-oxo-1, 2-oxazol-4-yl) propanoic acid receptor (AMPAR) subunits in the hippocampus of naive immature and adult rats (IRs, ARs) was investigated after status convulsion (SC).

Methods

Seizures were induced in IRs and ARs with intraperitoneal injections of lithium and pilocarpine. Rats were killed at four time points (3?h, 1?day, 3?days, and 7?days) after SC. The proportion of apoptotic cells was quantified by Annexin V-FITC apoptosis detection. The location and type of apoptotic cells were assessed by using terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling staining. Immunoblotting techniques were used to demonstrate changes in AMPAR subunit expression.

Results

Severe seizures induced neuronal apoptosis in the hippocampus. The proportion of apoptotic cells in IRs was consistently lower than that in ARs after SC. The expressions of four AMPAR subunits in IRs were consistently lower than those in ARs before and after SC. SC for 1?h inhibited the expression of glutamate receptors (GluR1–4) in the hippocampus of IRs and ARs and altered the subunit composition of AMPARs. GluR2 was the predominant AMPAR subunit in the hippocampus of normal ARs, while the GluR2/3 subunits were predominantly expressed 7?days after SC. GluR3/4 subunits were mainly expressed in the hippocampus of normal IRs, which had the lowest levels of GluR2.

Conclusions

Immature brain was more resistant to seizure-induced neural damage. The time course of reduction and recovery differed for each subunit and was dependent on developmental stage. The increased expression of GluR2 could confer early but transient protection in the immature brain after SC.     

A chemical-genetic strategy reveals distinct temporal requirements for SAD-1 kinase in neuronal polarization and synapse formation

Background

β-Amyloid precursor protein (APP) has been reported to play a role in the outgrowth of neurites from cultured neurons. Both cell-surface APP and its soluble, ectodomain cleavage product (APPs-α) have been implicated in regulating the length and branching of neurites in a variety of assays, but the mechanism by which APP performs this function is not understood.

Results

Here, we report that APP is required for proper neurite outgrowth in a cell autonomous manner, both in vitro and in vivo. Neurons that lack APP undergo elongation of their longest neurite. Deletion of APLP1 or APLP2, homologues of APP, likewise stimulates neurite lengthening. Intriguingly, wild-type neurons exposed to APPs-α, the principal cleavage product of APP, also undergo neurite elongation. However, APPs-α is unable to stimulate neurite elongation in the absence of cellular APP expression. The outgrowth-enhancing effects of both APPs-α and the deletion of APP are inhibited by blocking antibodies to Integrin β1 (Itgβ1). Moreover, full length APP interacts biochemically with Itgβ1, and APPs-α can interfere with this binding.

Conclusion

Our findings indicate that APPs-α regulates the function of APP in neurite outgrowth via the novel mechanism of competing with the binding of APP to Itgβ1.     

Effects of synaptic plasticity regulated by 17β-estradiol on learning and memory in rats with Alzheimer’s disease

Objective

To investigate whether estrogen modulates learning and memory and long-term potentiation (LTP) in the hippocampus of rats with Alzheimer’s disease (AD).

Methods

The rats were divided into ovariectomy (OVX) and estrogen replacement therapy (ERT) groups. Rats in the ERT group received OVX, followed by ERT, while rats in the OVX group received only OVX. The rat model of AD was established by injection of 1 μL (10 μg/μL) amyloid-beta peptide 1–40(Aβ_1–40) into the hippocampus. The learning and memory ability and LTP were determined by Morris water maze and electrophysiological method, respectively.

Results

The escape latency in Morris water maze significantly decreased in ERT group compared with that in OVX group (P < 0.05). Besides, rats in ERT group exhibited a significant enhancement of the magnitude of LTP at 30 min after high-frequency stimulation (HFS), compared with that in OVX group (P < 0.01).

Conclusion

ERT can attenuate the cognitive deficits in the rat model of AD, and estrogen can regulate LTP and synaptic remodeling in AD rats.     

Expression of connexin 30 and connexin 32 in hippocampus of rat during epileptogenesis in a kindling model of epilepsy

Objective Understanding the molecular and cellular mechanisms underlying epileptogenesis yields new insights into potential therapies that may ultimately prevent epilepsy.Gap junctions(GJs) create direct intercellular conduits between adjacent cells and are formed by hexameric protein subunits called connexins(Cxs).Changes in the expression of Cxs affect GJ communication and thereby could modulate the dissemination of electrical discharges.The hippocampus is one of the main regions involved in epileptogenesis and has a wide network of GJs between different cell types where Cx30 is expressed in astrocytes and Cx32 exists in neurons and oligodendrocytes.In the present study,we evaluated the changes of Cx30 and Cx32 expression in rat hippocampus during kindling epileptogenesis.Methods Rats were stereotaxically implanted with stimulating and recording electrodes in the basolateral amygdala,which was electrically stimulated once daily at afterdischarge threshold.Expression of Cx30 and Cx32,at both the mRNA and protein levels, was measured in the hippocampus at the beginning,in the middle(after acquisition of focal seizures),and at the end(after establishment of generalized seizures) of the kindling process,by real-time PCR and Western blot.Results Cx30 mRNA expression was upregulated at the beginning of kindling and after acquisition of focal seizures.Then it was downregulated when the animals acquired generalized seizures.Overexpression of Cx30 mRNA at the start of kindling was consistent with the respective initial protein increase.Thereafter,no change was found in protein abundance during kindling.Regarding Cx32,mRNA expression decreased after acquisition of generalized seizures and no other significant change was detected in mRNA and protein abundance during kindling.Conclusion We speculate that Cx32 GJ communication in the hippocampus does not contribute to kindling epileptogenesis.The Cx30 astrocytic network localized to perivascular regions in the hippocampus is,however,overexpressed at the initiation of kindling to clear excitotoxic molecules from the milieu.     

Granule cell mRNA levels for BDNF, NGF, and NT-3 correlate with neuron losses or supragranular mossy fiber sprouting in the chronically damaged and epileptic human hippocampus

This study determined in temporal lobe epilepsy patients if there were correlations among hippocampal granule cell expression of neurotrophin mRNAs, aberrant supragranular mossy fiber sprouting, and neuron losses. Consecutive surgically resected hippocampi (n=9) and comparison tissue from autopsies (n=3) were studied for:

1. Granule cell mRNA levels usingin situ hybridization for brainderived neurotrophic factor (BDNF), nerve growth factor (NGF), and neurotrophin-3 (NT-3);
2. neo-Timm supragranular mossy fiber sprouting; and
3. Ammon’s horn neuron densities.

Clinically, patients were classified into those with hippocampal sclerosis (HS;n=7) and non-HS cases (i.e., mass lesions and autopsies;n=5). Results showed that compared to non-HS cases, HS patients showed increased granule cell mRNA levels for BDNF, NGF, and NT-3 (p=0.035,p=0.04,p=0.045 respectively; one-tail directional test). Moreover, granule cell BDNF mRNA levels correlated inversely with Ammon’s horn neuron densities (p=0.02) and correlated positively with greater supragranular mossy fiber sprouting (p=0.02). NGF mRNA levels correlated inversely with Ammon’s horn neuron densities (p=0.02), and TN-3 mRNA levels correlated inversely with age at surgery (p=0.04) and correlated positively with greater mossy fiber sprouting (p=0.026). These results indicate in the chronically damaged human hippocampus that granule cells express neurotrophin mRNAs, and mRNA levels correlate with either hippocampal neuron losses or aberrant supragranular mossy fiber sprouting. These data support the hypothesis that in the epileptic human hippocampus, there may be pathophysiologic associations among mossy fiber synaptic plasticity, hippocampal neuron damage, and granule cell mRNA neurotrophin levels.     

Comparative Analysis of Gene Expression Profiles Involved in Calcium Signaling Pathways Using the NLVH Animal Model of Schizophrenia

In this study, we evaluated the expression profile changes of genes that intervene in the calcium signaling pathway, in young and adult Wistar rats, using the animal model of neonatal lesion in ventral hippocampus (NLVH) (a recognized animal model for schizophrenia) and compared to the group of control animals (Sham). Through microarray technology, gene expression profiles were obtained from the three brain areas (nucleus accumbens, prefrontal cortex, and hippocampus) of young male Wistar rats (45 days) and adults (90 days) whether or not subjected to NLVH. The calcium signaling pathway reported a greater number of differentially expressed genes with z-score two values, >?2 (over-expression) and <???2 (under-expression), in the three evaluated areas. The comparative analyses of this approach were performed in juvenile and adult rats with ventral hippocampal lesion in neonate rats (NLVH). NLVH influenced change expressions in various genes involved in Ca²⁺ homeostasis, including Cacna1d, Atp2a2, Adcy2, Ppp3cb, and Ptk2b. The expression of Adcy2, Ppp3cb, and Ptk2b genes changed in both age groups; therefore, the study of gene expression profiles between juvenile and adult rats may help to understand the molecular mechanisms of schizophrenia.     

Upregulation of 5-HT2C receptors in hippocampus of pilocarpine-induced epileptic rats: Antagonism by Bacopa monnieri

Emotional disturbances, depressive mood, anxiety, aggressive behavior, and memory impairment are the common psychiatric features associated with temporal lobe epilepsy (TLE). The present study was carried out to investigate the role of Bacopa monnieri extract in hippocampus of pilocarpine-induced temporal lobe epileptic rats through the 5-HT_2C receptor in relation to depression. Our results showed upregulation of 5-HT_2C receptors with a decreased affinity in hippocampus of pilocarpine-induced epileptic rats. Also, there was an increase in 5-HT_2C gene expression and inositol triphosphate content in epileptic hippocampus. Carbamazepine and B. monnieri treatments reversed the alterations in 5-HT_2C receptor binding, gene expression, and inositol triphosphate content in treated epileptic rats as compared to untreated epileptic rats. The forced swim test confirmed the depressive behavior pattern during epilepsy that was nearly completely reversed by B. monnieri treatment.     

Somatostatin receptor subtype 2 (sst2) is a potential prognostic marker and a therapeutic target in medulloblastoma

Introduction

Neuroectodermal tumors in general demonstrate high and dense expression of the somatostatin receptor subtype 2 (sst₂). It controls proliferation of both normal and neoplastic cells. sst₂ has thus been suggested as a therapeutic target and prognostic marker for certain malignancies.

Methods

To assess global expression patterns of sst ₂ mRNA, we evaluated normal (n?=?353) and tumor tissues (n?=?340) derived from previously published gene expression profiling studies. These analyses demonstrated specific upregulation of sst ₂ mRNA in medulloblastoma (p?<?0.001). sst₂ protein was investigated by immunohistochemistry in two independent cohorts.

Results

Correlation of sst₂ protein expression with clinicopathological variables revealed significantly higher levels in medulloblastoma (p?<?0.05) compared with CNS-PNET, ependymoma, or pilocytic astrocytoma. The non-SHH medulloblastoma subgroup tumors showed particularly high expression of sst₂, when compared to other tumors and normal tissues. Furthermore, we detected a significant survival benefit in children with tumors exhibiting high sst₂ expression (p?=?0.02) in this screening set. A similar trend was observed in a validation cohort including 240 independent medulloblastoma samples.

Conclusion

sst₂ is highly expressed in medulloblastoma and deserves further evaluation in the setting of prospective trials, given its potential utility as a prognostic marker and a therapeutic target.     

成年大鼠脊髓全横断损伤后酪氨酸激酶受体C在脊髓和大脑皮层的表达

目的 研究神经营养因子3(neurotrophin-3,NT-3)的受体-酪氨酸激酶受体C(tyrosine kinase receptor C,TrkC)在脊髓损伤(spinal cord injury,SCI)后神经重塑中的作用.方法 研究脊髓全横断损伤大鼠手术后第1、3、7和14 d时,低位胸髓节段和大脑中央前回...     

A direct regulatory link between microRNA-137 and <Emphasis Type="Italic">SHANK2</Emphasis>: implications for neuropsychiatric disorders

Background

Mutations in the SHANK genes, which encode postsynaptic scaffolding proteins, have been linked to a spectrum of neurodevelopmental disorders. The SHANK genes and the schizophrenia-associated microRNA-137 show convergence on several levels, as they are both expressed at the synapse, influence neuronal development, and have a strong link to neurodevelopmental and neuropsychiatric disorders like intellectual disability, autism, and schizophrenia. This compiled evidence raised the question if the SHANKs might be targets of miR-137.

Methods

In silico analysis revealed a putative binding site for microRNA-137 (miR-137) in the SHANK2 3′UTR, while this was not the case for SHANK1 and SHANK3. Luciferase reporter assays were performed by overexpressing wild type and mutated SHANK2-3′UTR and miR-137 in human neuroblastoma cells and mouse primary hippocampal neurons. miR-137 was also overexpressed or inhibited in hippocampal neurons, and Shank2 expression was analyzed by quantitative real-time PCR and Western blot. Additionally, expression levels of experimentally validated miR-137 target genes were analyzed in the dorsolateral prefrontal cortex (DLPFC) of schizophrenia and control individuals using the RNA-Seq data from the CommonMind Consortium.

Results

miR-137 directly targets the 3′UTR of SHANK2 in a site-specific manner. Overexpression of miR-137 in mouse primary hippocampal neurons significantly lowered endogenous Shank2 protein levels without detectable influence on mRNA levels. Conversely, miR-137 inhibition increased Shank2 protein expression, indicating that miR-137 regulates SHANK2 expression by repressing protein translation rather than inducing mRNA degradation.To find out if the miR-137 signaling network is altered in schizophrenia, we compared miR-137 precursor and miR-137 target gene expression in the DLPFC of schizophrenia and control individuals using the CommonMind Consortium RNA sequencing data. Differential expression of 23% (16/69) of known miR-137 target genes was detected in the DLPFC of schizophrenia individuals compared with controls. We propose that in further targets (e.g., SHANK2, as described in this paper) which are not regulated on RNA level, effects may only be detectable on protein level.

Conclusion

Our study provides evidence that a direct regulatory link exists between miR-137 and SHANK2 and supports the finding that miR-137 signaling might be altered in schizophrenia.

     

Tetrahydroxy stilbene glucoside reduces the cognitive impairment and overexpression of amyloid precursor protein induced by aluminum exposure

Objective

Excessive aluminum (Al) exposure impairs neurocognitive function in humans and animals. Epidemiologic studies have shown a potential linkage between chronic Al exposure and Alzheimer’s disease. The present study aims to evaluate the effects of tetrahydroxy stilbene glucoside (TSG), the extract from herbal medicine Polygoni Multiflori, on cognitive impairment and the over-expression of hippocampal amyloid precursor protein (APP) induced by chronic exposure to Al in rats

Methods

Rats were treated with 0.3% aluminum chloride (AlCl3) prepared in the drinking water for 90 d. AlCl₃-treated animals were then randomly assigned to receive vehicle, TSG (4 g/kg), or Vitamin E (VE; 40 mg/kg) treatment for 5 months. VE served as a positive control. The effect of TSG was evaluated by passive avoidance task, and APP expression was evaluated by Western blotting.

Results

Following exposure to AlCl₃ for 90 d, animals displayed a striking decrease (>80%) in step-through latency in the passive avoidance task and a significant increase in the expression of APP in the hippocampus. Both TSG and VE significantly ameliorated the performance impairment in the passive avoidance task, and suppressed the over-expression of APP. Moreover, the effects of TSG, but not of VE, were in a time-dependent manner.

Conclusion

TSG may possess therapeutic effects against Alzheimer’s disease.     

Effects of nerve growth factor in experimental model of focal microgyria

Aim

The effects on neural repair of intraparenchymal nerve growth factor (NGF) administration were evaluated in neonate Wistar rats with experimentally induced focal microgyria.

Methods

A freezing focal polymicrogyric lesion was induced on the frontal cortex in 35 newborn Wistar rats on postnatal day?1. NGF was administered in 15 cases, with 20 pups as controls. Animals were sacrificed at 72?h and 7?days after NGF administration. Real-time PCR was used for the quantification of the expression of TrkA, p75, and doublecortin (DCX) at the level of the cortical lesion in seven different groups of animals: control 72?h (n?=?5), control 7?days (n?=?5), microgyria 72?h (n?=?5), microgyria 7?days (n?=?5), microgyria + NGF 72?h (n?=?5), microgyria + NGF 7?days (n?=?5), and control + NGF (n?=?5).

Results

A significant increase in TrkA expression was found in the microgyria + NGF 7?days group compared to the others. TrkA upregulation was already visible 72?h after NGF administration. Unlike TrkA, p75 expression increased in animals subjected to the experimental focal microgyria and decreased markedly after NGF administration. DCX expression in injured animals was observed to increase strongly 7?days after NGF administration compared with other groups.

Conclusions

NGF administration interferes with neural repair mechanisms at the polymicrogyric lesion site by means of TrkA and DCX upregulation which possibly counteracts the process of apoptosis caused by the brain injury.     

Expression Analysis of Vitamin D Signaling Pathway Genes in Epileptic Patients

Vitamin D deficiency has been detected in epileptic patients. Vitamin D participates in neuroprotection, brain cell proliferation, and differentiation. Consequently, vitamin D supplementation has been suggested as an alternative treatment in epileptic patients. We aimed at assessment of vitamin D signaling pathway in epileptic patients. In the present study, we evaluated vitamin D serum concentration as well as expression of vitamin D receptor (VDR) gene and genes encoding for vitamin D activating enzyme 1-alpha-hydroxylase (CYP27B1) and deactivating enzyme 24-hyroxylase (CYP24A1) in epileptic patients compared with healthy individuals. We found significant lower levels of vitamin D in epileptic patients compared with healthy subjects. Expression analyses showed significant downregulation of VDR expression in peripheral blood of epileptic patients compared with healthy subjects (relative expression (REx)?=?0.16, P?<?0.001). However, there was no significant difference in CYP24A1 expression between epileptic patients and normal subjects. CYP27B1 expression analysis showed significant upregulation in male patients aged between 30 and 40 (REx?=?5.43, P?=?0.013). After using two-way ANCOVA for adjusting the effects of sex and age, there was a statistically significant difference in the VDR expression values between patient and control groups (P?<?0.001). Spearman’s correlation analysis showed no significant correlation between genes expression levels and patients’ age or vitamin D serum concentrations. However, we found significant correlations between VDR expression levels and CYP24A1/ CYP27B1 expression levels in epileptic patients (r?=?0.435 and P?<?0.001; r?=?0.26 and P?=?0.02 respectively). There was also a significant correlation between the expression levels of CYP24A1 and CYP27B1 (r?=?0.349 and P?=?0.001). Our study shows a possible role for VDR in the pathogenesis of epilepsy.     

Selective regulation of nerve growth factor expression in developing cutaneous tissue by early sensory innervation

Background

In the developing vertebrate peripheral nervous system, the survival of sympathetic neurons and the majority of sensory neurons depends on a supply of nerve growth factor (NGF) from tissues they innervate. Although neurotrophic theory presupposes, and the available evidence suggests, that the level of NGF expression is completely independent of innervation, the possibility that innervation may regulate the timing or level of NGF expression has not been rigorously investigated in a sufficiently well-characterized developing system.

Results

To address this important question, we studied the influence of innervation on the regulation of NGF mRNA expression in the embryonic mouse maxillary process in vitro and in vivo. The maxillary process receives its innervation from predominantly NGF-dependent sensory neurons of the trigeminal ganglion and is the most densely innervated cutaneous territory with the highest levels of NGF in the embryo. When early, uninnervated maxillary processes were cultured alone, the level of NGF mRNA rose more slowly than in maxillary processes cultured with attached trigeminal ganglia. In contrast to the positive influence of early innervation on NGF mRNA expression, the levels of brain-derived neurotrophic factor (BDNF) mRNA and neurotrophin-3 (NT3) mRNA rose to the same extent in early maxillary processes grown with and without trigeminal ganglia. The level of NGF mRNA, but not BDNF mRNA or NT3 mRNA, was also significantly lower in the maxillary processes of erbB3 ^-/- mice, which have substantially fewer trigeminal neurons than wild-type mice.

Conclusions

This selective effect of initial innervation on target field NGF mRNA expression provokes a re-evaluation of a key assertion of neurotrophic theory that the level of NGF expression is independent of innervation.     

Histological studies on the telencephalon of Hynobius leechii at the metamorphosis phase and the adult phase

Objective

To investigate the telencephalon developmental characteristics of Hynobius leehii, and enrich the research data of comparable neurobiology and nervous system development of amphibian.

Methods

HE staining and Nissl staining methods were used to study the telencephalon histological structure of Hynobius leechii at both the metamorphosis and the adult phases, and to explore the developmental phases of telencephalon.

Results

The olfactory bulb could be roughly divided into 6 layers from lateral to medial. The lateral cerebral ventricles at the metamorphosis phase were smaller than those at the adult phase, and there were no clear borderlines between the primordial pallium and the primordial hippocampus, or between the primordial pallium and the primordial piriform area. Moreover, the cells in the primordial piriform area were more closely distributed than those in the primordial hippocampus or the primordial pallium. Compared with those at the adult phase, cells in nucleuses at the metamorphosis phase were larger in number and more closely distributed.

Conclusion

The telencephalon of Hynobius leehii at the metamorphosis phase has generally formed the adult structure, however, it is still at a transition state of differentiation to maturity during the development of Hynobius leehii.     

Contemplating Genetic Feedback Regarding Lung Cancer Susceptibility

Background and Purpose

We examined three theoretical models (self-enhancement theory, consistency theory, and a combined model) for understanding how expectations and test result favorability influence smokers' desire for a retest following hypothetical genetic test results.

Method

College smokers (N?=?128) read a brochure describing a biomarker for lung cancer (the GSTM1 gene) then reported whether they thought they had the gene (indicating lower lung cancer risk) or were missing the gene (indicating higher lung cancer risk). Participants then reported whether they would get retested if they received favorable GSTM1 results versus unfavorable GSTM1 results.

Results

Participants were most likely to want a retest, suggesting rejection of the results, if they expected favorable news yet received unfavorable news.

Conclusion

The findings supported the combined model such that smokers expressed greatest interest in a retest when they imagined genetic risk feedback that challenges both enhancement and consistency motives.     

铜离子对MES23.5细胞的毒性作用机制与线粒体功能障碍有关

目的探讨铜离子对MES23．5多巴胺能细胞的毒性作用及可能机制。方法用MTF法检测铜离子对细胞存活率的影响;用半定量逆转录聚合酶链反应（RT-PCR）,Western blotting以及高效液相色谱电化学检测法（HPLC-ECD）检测细胞内酪氨酸羟化酶（tyrosine hydroxlase,TH）mRNA、蛋白的表达以及多巴胺含量的改变：用流式细胞仪检测线粒体跨膜电位的改变。结果100μmol／L和200／2mol／L铜离子对细胞存活率没有影响,400μmol／L和800μmol／L铜离子可以造成细胞存活率降低（P〈0．01）。200μmol／L铜离子孵育细胞24h,THmRNA、蛋白表达量以及多巴胺含量较正常对照组降低（P〈0．01,P〈0．01,P〈0．05）。200μmol／L铜离子孵育细胞24h,线粒体跨膜电位较正常对照组明显降低（P〈0．01）。结论铜离子对MES23．5多巴胺能细胞具有毒性作用,降低细胞功能的表达,其机制可能与线粒体功能障碍有关。