1.Expressions of CCAAT/enhancer-binding Protein Homologous Protein and Calnexin in the Hippocampus of a Mouse Model of Mesial Temporal Lobe Epilepsy.
Zhi-qiang SHA ; Long-ze SHA ; Qi XU
Acta Academiae Medicinae Sinicae 2016;38(3):265-270
Objective To explore the temporal and spatial distribution of CCAAT/enhancer-binding protein homologous protein (CHOP) and calnexin (CNX) in the dentate gyrus of mesial temporal lobe epilepsy (mTLE) mouse model. Methods We used kainic acid (KA) to induce acute phase (12 h and 24 h) mTLE mouse models and performed Western blotting and immunofluorescence to detect the different expressions and distribution pattern of CHOP and CNX in CA3 of the hippocampus. Results Compared with the controls,the expressions of CHOP(F=1.136,P=0.4069) and CNX (F=2.378,P=0.2087) did not increase in CA3 of hippocampus 12 h following KA injection in the acute phase of mTLE mouse models,whereas the expressions in CA1 and CA3 of hippocampus 24 h after injection were significantly higher (F=8.510,P=0.0362;F=6.968,P=0.0497,respectively). As shown by immunofluorescence analysis,CHOP was expressed mainly in CA3 of hippocampus 12 h after KA injection,and increased in CA1 and CA3 24 h after KA administration. Compared with the controls,the expressions of CHOP(F=24.480,P=0.0057) and CNX (F=7.149,P=0.0478) were significantly higher 24 h after KA injection.Conclusions The expression of CHOP increases along with the progression of seizures,indicating the increased level of endoplasmic reticulum stress. An increasing number of CNX,which serves as molecular chaperone,may be needed to facilitate the unfolded protein to complete the folding process.
Animals
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Calnexin
;
metabolism
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Dentate Gyrus
;
metabolism
;
Disease Models, Animal
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Epilepsy, Temporal Lobe
;
chemically induced
;
metabolism
;
Kainic Acid
;
Mice
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Seizures
;
chemically induced
;
metabolism
;
Transcription Factor CHOP
;
metabolism
2.Axonal sprouting of somatostatin positive interneurons in the hippocampus in epileptic rats.
Fang YI ; Bo XIAO ; Ting JIANG ; Lili LONG ; Jinghui LIANG ; Li FENG ; Guoliang LI
Journal of Central South University(Medical Sciences) 2011;36(12):1176-1182
OBJECTIVE:
To investigate the axonal sprouting of somatostatin(SS) positive interneurons in temporal lobe epilepsy.
METHODS:
6-8 week-old healthy male SD rats were divided randomly into an epileptic group (treated by lithium and pilocarpine intraperitoneal injection) and a control group (by lithium and normal sodium intraperitoneal injection). Each group was randomly divided into 5 subgroups at 1,7,15,30, amd 60 d after the injection. Immunohistochemistry method was used to detect the number changes of SS or neuronal nuclei (NeuN) positive neurons in different domains of the hippocampus at different time points in each group, and the coexpression of SS positive interneurons combined with NeuN was detected by double immunofluorescence to observe the dynamic changes and axonal sprouting of SS positive interneurons.
RESULTS:
The number of SS neurons in the experimental group exceeded that in the control group in the CA1 area at 60 d post-status epileptieus SE (P<0.01), and numerous SS positive fibers were seen throughout the layers of the CAl area at 60 d post-SE. NeuN positive neurons in the stratum oriens and stratum radiatum layers in the initiation site of the CA1 area were beyond normal at 60 d post-SE. The number of double labeled SS interneurons gradually rose at 15 d in stratum oriens of CA1, and even exceeded that of the controls in the stratum oriens and stratum radiatum layers of CA1 at 60 d.
CONCLUSION
The numerous SS positive fibers throughout the layers of the CAl area at 60 d post-SE come from the increased interneurons in the stratum oriens and stratum radiatum layers of CA1 area. The pathological axonal sprouting may play an important role in the generation and compensation of temporal lobe epilepsy.
Animals
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Axons
;
metabolism
;
pathology
;
CA1 Region, Hippocampal
;
cytology
;
metabolism
;
physiopathology
;
Efferent Pathways
;
pathology
;
physiology
;
Epilepsy, Temporal Lobe
;
chemically induced
;
metabolism
;
physiopathology
;
Interneurons
;
cytology
;
metabolism
;
pathology
;
Male
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Pilocarpine
;
Random Allocation
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Rats
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Rats, Sprague-Dawley
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Somatostatin
;
metabolism
;
Temporal Lobe
;
metabolism
3.Morphology and differentially expressed proteins in hippocampus of mesial temporal lobe epilepsy model of immature rats induced by pilocarpine.
Liwen WU ; Fei YIN ; Jing PENG ; Fang HE ; Ciliu ZHANG ; Xiaolu DENG ; Guoli WANG
Journal of Central South University(Medical Sciences) 2013;38(6):560-569
OBJECTIVE:
To examine the changes of morphology and differentially expressed proteins in hippocampus at the latent stage of chronic mesial temporal lobe epilepsy (MTLE) in immature rats, and to explore the global mechanism of chronic MTLE at a new point.
METHODS:
MTLE models of immature rats were induced by lithium-pilocarpine. The rats were divided into 2 groups randomly: a control group (n=20) and an MTLE model group (n=20). At the latent stage, nissl and Timm staining were performed to evaluate the cell loss and mossy fiber sprouting. The differentially expressed proteins were separated by 2-dimensional polyacrylamide gel electrophoresis (2-DE) combined with matrix-assisted laser desorption/ ionization time of flight mass spectrometry (MALDI-TOF-MS) technology. Western blot was used to determine the differentially expression levels of partial proteins.
RESULTS:
Neuron loss and abnormal mossy fiber sprouting were obviously observed in the hippocampus in the MTLE model group; 2-DE patterns of hippocampus of the MTLE model group in latent stage and the control group were established. Thirty-one differential proteins were identified by MALDI-TOF-MS, which were categorized into several groups by biological functions: synaptic and neurotransmitter release related proteins, cytoskeletal proteins, cell junctions proteins, energy metabolism and mitochondrial proteins, biological enzymes, cellular structure related proteins, signal regulating molecular and others. The expression levels of partial proteins determined by Western blot were similar to the changes of proteomics.
CONCLUSION
The differentially expressed proteins of synapse-related proteins such as dynamin-1, neurogranin and ubiquitin, which cause the synapse reorganization and mossy fiber terminal sprouting related to the formation of abnormal excitatory network, probably play critic roles in the mechanism of MTLE.
Animals
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Epilepsy, Temporal Lobe
;
chemically induced
;
metabolism
;
pathology
;
Female
;
Hippocampus
;
metabolism
;
pathology
;
Male
;
Pilocarpine
;
Proteins
;
genetics
;
metabolism
;
Proteomics
;
methods
;
Rats
;
Rats, Sprague-Dawley
4.Tenidap is neuroprotective in a pilocarpine rat model of temporal lobe epilepsy.
Xing-Hua TANG ; Xun-Yi WU ; Lan XU ; You-Xin FANG ; Jian-Hong WANG ; Guo-Xing ZHU ; Zhen HONG
Chinese Medical Journal 2013;126(10):1900-1905
BACKGROUNDTenidap is a liposoluble non-steroidal anti-inflammatory drug that is easily distributed in the central nervous system and also inhibits the production and activity of cyclooxygenase-2 (COX-2) and cytokines in vitro. This study aimed to evaluate the neuroprotective effect of tenidap in a pilocarpine rat model of temporal lobe epilepsy (TLE).
METHODSTenidap was administered daily at 10 mg/kg for 10 days following pilocarpine-induced status epilepticus (SE) in male Wistar rats after which prolonged generalized seizures resulted in TLE. After tenidap treatment, spontaneous recurrent seizures (SRSs) were recorded by video monitoring (for 7 hours per day for 14 days). The frequency and severity of the SRSs were observed. Histological and immunocytochemical analyses were used to evaluate the neuroprotective effect of tenidap and detect COX-2 expression, which may be associated with neuronal death.
RESULTSThere were 46.88 ± 10.70 survival neurons in tenidap-SE group, while there were 27.60 ± 5.18 survival neurons in saline-SE group at -2.4 mm field in the CA3 area. There were 37.75 ± 8.78 survival neurons in tenidap-SE group, while there were 33.40 ± 8.14 survival neurons in saline-SE group at -2.4 mm field in the CA1 area. Tenidap treatment significantly reduced neuronal damage in the CA3 area (P < 0.05) and slightly reduced damage in the CA1 area. Tenidap markedly inhibited COX-2 expression in the hippocampus, especially in the CA3 area.
CONCLUSIONTenidap conferred neuroprotection to the CA3 area in a pilocarpine-induced rat model of TLE by inhibiting COX-2 expression.
Animals ; Cyclooxygenase 2 ; metabolism ; Epilepsy, Temporal Lobe ; chemically induced ; drug therapy ; metabolism ; Indoles ; therapeutic use ; Male ; Neuroprotective Agents ; therapeutic use ; Pilocarpine ; toxicity ; Rats ; Rats, Wistar
5.c-JUN Expression and Apoptotic Cell Death in Kainate-Induced Temporal Lobe Epilepsy.
Min Cheol LEE ; Jin Lee RHO ; Myung Kyu KIM ; Young Jong WOO ; Jae Hyoo KIM ; Sang Chae NAM ; Jung Jin SUH ; Woong Ki CHUNG ; Jai Dong MOON ; Hyung Ihl KIM
Journal of Korean Medical Science 2001;16(5):649-656
Following kainate (KA)-induced epilepsy, rat hippocampal neurons strongly ex-press immediate early gene (IEG) products, i.e., c-FOS and c-JUN, and neural stress protein, HSP72. Prolonged expression of c-JUN and c-FOS 48 hr after cerebral ischemia has been underwent delayed neuronal death. However, it is not yet clear whether IEGs actually assume the essential roles in the cell death process or simply as a by-product due to external stimuli because of the prolonged expression of c-FOS, more than one week, on intact CA2 neurons of the hippocampus in a KA-induced epilepsy model. This study investigated the relationships between prolonged expression of c-JUN and hippocampal neuronal apoptosis in a KA-induced epilepsy model. Epileptic seizure was induced in rats by a single microinjection of KA (1g/l) into the left amygdala. Characteristic seizures and hippocampal neuronal injury were developed. The expression of c-JUN was evaluated by immunohistochemistry, and neuronal apoptosis by in situ end labeling. The seizures were associated with c-JUN expression in the hippocampal neurons, of which the level showed a positive correlation with that of apoptosis. Losses of hippocampal neurons, especially in the CA3 region, were partly caused by apoptotic cell death via a c-JUN-mediated signaling pathway. This is thought to be an important component in the pathogenesis of hippocampal neuronal injury via KA-induced epilepsy.
Animal
;
*Apoptosis
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Epilepsy, Temporal Lobe/chemically induced/*metabolism/pathology
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Hippocampus/*chemistry/pathology
;
Immunohistochemistry
;
Kainic Acid/*toxicity
;
Male
;
Proto-Oncogene Proteins c-jun/*analysis
;
Rats
;
Rats, Wistar
6.Number changes and axonal sprouting of neuropeptide Y interneurons in the hippocampus of pilocarpine-induced rats.
Zhiguo WU ; Lili LONG ; Bo XIAO ; Si CHEN ; Fang YI
Journal of Central South University(Medical Sciences) 2009;34(2):93-98
OBJECTIVE:
To investigate the role of neuropeptide Y(NPY) positive interneurons in the generation and compensation of temporal lobe epilepsy.
METHODS:
Pilocarpine-induced rat model was founded. Immunohistochemistry was used to observe the number changes and axonal sprouting of NPY interneurons at different time points in the hippocampus of rats.
RESULTS:
After lithium-chloride and pilocarpine administration, 92.9% rats were induced status epilepticus (SE) successfully, and the mortality rate was 19.2%. In the experimental group, the number of NPY positive neurons decreased in the hilus of the hippocampus, and was least on 7 d after the SE (P<0.01). In the chronic phase, the number of hilus NPY neurons partially recovered, but was still less than the number in the control group on 60 d after the SE (P<0.05). No evident changes of the number of NPY neurons existed in CA domains (P>0.05) except the loss of them in CA3 area on 7 d after the SE (P>0.05). Increased NPY positive fibers could be seen in the molecular layer of the dentate gyrus on 30 d after the SE.
CONCLUSION
NPY interneurons have different sensitivities to the injuries induced by seizures at different time points and domains. Loss of NPY interneurons plays an important role in the generation of temporal lobe epilepsy, while axonal sprouting of them may play a significant role in the compensation of temporal lobe epilepsy.
Animals
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Epilepsy, Temporal Lobe
;
chemically induced
;
pathology
;
Hippocampus
;
metabolism
;
pathology
;
Interneurons
;
metabolism
;
pathology
;
Male
;
Neuropeptide Y
;
metabolism
;
Pilocarpine
;
Random Allocation
;
Rats
;
Rats, Sprague-Dawley
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Retrograde Degeneration
;
pathology
;
Status Epilepticus
;
chemically induced
;
pathology
7.Eph in the mechanism of mossy fiber axon sprouting in dentate gyrus in rats with chronic temporal lobe epilepsy.
Wei-ping LIU ; Bo XIAO ; Shu-yu LI ; Xiao-qin LU
Journal of Central South University(Medical Sciences) 2008;33(8):657-662
OBJECTIVE:
To investigate the relationship among mossy fiber axon sprouting(MFS), synaptic reorganization, and the alteration of expression of Eph A5 and ephrin A3 in the dentate gyrus in rats with pilocarpine-induced chronic temporal lobe epilepsy.
METHODS:
Mossy fiber sprouting and synaptic formation in rats were observed by Neo-Timm staining, after the acute status epilepticus and chronic spontaneous temporal lobe epilepsy induced by lithium-chloride and pilocarpine. In situ hybridization was used to detect ephrin A3 mRNA and an immunohistochemical staining was applied to determine Eph A5 protein.
RESULTS:
In entorhinal cortex, only Eph A5 mRNA and protein expressed, which significantly decreased on Day 7 after pilocarpine induced status epilepticus(P<0.01),and resumed to normal levels on Day 30 (P>0.05). In the dentate granule cells, ephrin A3 mRNA reduced obviously on Day 7 after pilocarpine-induced status epilepticus (P<0.01), and returned to normal levels on Day 30 (P>0.05).
CONCLUSION
The down-regulation of Eph A5 mRNA and protein in entorhinal cortex and dentate gyrus, and ephrin A3 mRNA in dentate gyrus after status epilepticus may be part of the endogenous molecular mechanism of mossy fiber sprouting to the inner molecular layer of dentate gyrus.
Animals
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Axons
;
physiology
;
Chromosome Pairing
;
physiology
;
Dentate Gyrus
;
physiopathology
;
Down-Regulation
;
Ephrin-A3
;
biosynthesis
;
Epilepsy, Temporal Lobe
;
chemically induced
;
metabolism
;
physiopathology
;
RNA, Messenger
;
biosynthesis
;
genetics
;
Rats
;
Receptor, EphA5
;
biosynthesis
;
Status Epilepticus
;
chemically induced
;
metabolism
;
physiopathology
8.The effect of low frequency transcranial magnetic stimulation on neuropeptide-Y expression and apoptosis of hippocampus neurons in epilepsy rats induced by pilocarpine.
Yong-Li WANG ; Yu ZHAI ; Xiao-Lin HUO ; Jian-Ning ZHANG
Chinese Journal of Surgery 2007;45(24):1685-1687
OBJECTIVETo analyze the effect of low frequency transcranial magnetic stimulation (LF-TMS) on changing neuropeptide-Y (NPY) expression and apoptosis of hippocampus neurons in epilepsy rats induced by pilocarpine (PLO).
METHODSThirty male Sprague Dawley rats (240 g +/- 20 g) were randomly divided into 2 groups. I group simply celiac injected pilocarpine. II group celiac injected PLO after LF-TMS. Pathological item included HE staining, NPY immunohistochemical staining and apoptosis staining.
RESULTSHE staining revealed neurons of hippocampus were obviously death and cell's structure was destroyed in PLO group. The PLO + LF-TMS group was less injured and destroyed. Using One-Way ANOVA, NPY immunohistochemical staining shown the positive cell number was increased at all areas of hippocampus in PLO group contrasting with the low positive cell number in the PLO + LF-TMS group. In PLO group the number of apoptosis cell at hippocampus areas was significant higher than the PLO + LF-TMS group.
CONCLUSIONSUsing the PLO evoked epilepsy model, LF-TMS alleviated neurons injury at hippocampus area, so LF-TMS might playing an important role in resisting the progressing of epilepsy. The positive cell number of NPY increased at all areas of hippocampus, which indicated the close relation between NPY and epilepsy. NPY might have some function on resisting epilepsy.
Animals ; Apoptosis ; Disease Models, Animal ; Epilepsy, Temporal Lobe ; chemically induced ; metabolism ; pathology ; therapy ; Hippocampus ; metabolism ; pathology ; Male ; Neurons ; metabolism ; pathology ; Neuropeptide Y ; metabolism ; Pilocarpine ; toxicity ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Transcranial Magnetic Stimulation ; methods
9.Calcium/Calmodulin Kinase II Activity of Hippocampus in Kainate-Induced Epilepsy.
Min Cheol LEE ; Sung Soo BAN ; Young Jong WOO ; Seung U KIM
Journal of Korean Medical Science 2001;16(5):643-648
This study investigated calcium/calmodulin kinase II (CaMKII) activity related to long-standing neuronal injury of the hippocampus in kainate (KA)-induced experimental temporal lobe epilepsy. Epileptic seizure was induced by injection of KA (1 g/L) dissolved in phosphate buffer (0.1 M, pH 7.4) into the left amygdala. Clinical seizures, histopathologic changes and CaMKII activity of the hippocampus were evaluated. Characteristic early limbic and late seizures were developed. Hippocampal CaMKII activity increased significantly 4 and 8 weeks after intra-amygdaloid injection of KA, when late seizures developed. The histopathologic changes of the hippocampus included swelling of neuronal cytoplasm with nuclear pyknosis and loss of neurons in CA3 during this period. The increased activity of CaMKII may correlate with appearance of distant damage in the hippocampus. The above results indicate that intra-amygdaloid injection of KA produces excitatory signals for ipsilateral CA3 neurons in the hippocampus and that subsequently increased levels of CaMKII in postsynaptic neurons induce neuronal injury via phosphorylation of N-methyl-D-aspartate type glutamate receptor.
Animal
;
Ca(2+)-Calmodulin Dependent Protein Kinase/*metabolism
;
Epilepsy, Temporal Lobe/chemically induced/*enzymology/pathology
;
Hippocampus/*enzymology/pathology
;
Kainic Acid/*toxicity
;
Long-Term Potentiation/drug effects
;
Male
;
Rats
;
Rats, Wistar
10.Induction of Bis, a Bcl-2-binding protein, in reactive astrocytes of the rat hippocampus following kainic acid-induced seizure.
Mun Yong LEE ; Seong Yun KIM ; Jeong Sun CHOI ; Yun Sik CHOI ; Mi Hee JEON ; Jung Hee LEE ; In Kyung KIM ; Jeong Hwa LEE
Experimental & Molecular Medicine 2002;34(2):167-171
The expression of Bis (also called Bag-3), a Bcl-2-binding protein, was investigated in the rat kainic acid (KA) model of temporal lobe epilepsy. Western blot analysis showed a significant increase in the expression levels of Bis protein in the hippocampus following the systemic administration of KA. Bis immunoreactivity increased preferentially in the CA1 and CA3 regions, as well as in the hilar region of the dentate gyrus. Experiments with double immunofluorescence revealed that, in KA-administered rats, the cells expressing Bis were GFAP-expressing reactive astrocytes. The increase in Bis immunoreactivity was accompanied by increased Bcl-2 in reactive astrocytes in the striatum radiatum, whereas Bcl-2 immunoreactivity in pyramidal neurons was not affected. These results of the co-expression of Bis and Bcl-2 in reactive astrocytes in this seizure model suggest that Bis might modulate the glial reaction under excitotoxic brain injury, probably by interacting with Bcl-2.
Animals
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Apoptosis
;
Astrocytes/*metabolism
;
Blotting, Western
;
Carrier Proteins/*biosynthesis
;
Disease Models, Animal
;
Epilepsy, Temporal Lobe/chemically induced/*metabolism
;
Fluorescent Antibody Technique
;
Hippocampus/*metabolism/pathology
;
Kainic Acid
;
Male
;
Neurons/physiology
;
Protein Binding
;
Proto-Oncogene Proteins c-bcl-2/metabolism
;
Rats
;
Rats, Sprague-Dawley