repository
Graduate School of Ajou University
Department of Neuroscience and Technology Course
3. Theses(Master)
Glutamate로 유도된 뇌신경세포 사멸과정에서 Na+/H+ exchanger-1의 역할
| DC Field | Value | Language |
|---|---|---|
| dc.contributor.advisor | 문창현 | - |
| dc.contributor.author | Lee, Bo Kyung | - |
| dc.date.accessioned | 2019-10-21T07:14:22Z | - |
| dc.date.available | 2019-10-21T07:14:22Z | - |
| dc.date.issued | 2010-08 | - |
| dc.identifier.other | 10884 | - |
| dc.identifier.uri | https://dspace.ajou.ac.kr/handle/2018.oak/17636 | - |
| dc.description | 학위논문(박사)--아주대학교 일반대학원 :신경과학기술과정,2010. 8 | - |
| dc.description.tableofcontents | ABSTRACT TABLE OF CONTENTS LIST OF FIGURES LIST OF TABLES ABBREVIATION I. INTRODUCTION A. The role of Na+/H+ exchanger (NHE) B. The role of NHE-1 during brain ischemia C. The regulation of NHE-1 during brain ischemia D. The role of Protein kinase Cs (PKC) during brain ischemia E. The regulation of NHE-1 during PKC activation F. Aims of study II. MATERIALS AND METHODS A. Materials B. Cell cultures 1. Primary culture of cortical neurons 2. Cell lines C. Measurements of pHi and NHE activity D. Cell death analysis 1. Lactate dehydrogenase (LDH) assay 2. TUNEL assay 3. Caspase-3 activity 4. Trypan blue staining 5. Annexin V –FITC and propidium iodide (PI) staining 6. MTT assay E. Image analysis 1. Measurement of intracellular Ca2+ 2. Measurement of mitochondrial Ca2+ 3. Measurement of mitochondrial membrane potential (ΔΨm) 4. Measurement of ROS release F. Separation of cytosolic and mitochondrial fractions for cytochrome c release G. Gel electrophoresis and Western blotting H. RNA preparation and RT-PCR I. Analysis of NHE-1 phosphorylation by immunoprecipitation J. Transfection and reagents K. Small interfering RNA treatment L. Statistical analysis III. RESULTS A. NHE-1 inhibitory potency of NHE-1 inhibitors B. Acidosis-mediated NHE-1 activation during glutamate exposure in neurons C. The effects of NHE-1 inhibitors on glutamate-induced neuronal apoptotic and necrotic cell death D. The effects of NHE-1 inhibitors on cytosolic and mitochondrial Ca2+ overload induced by glutamate E. The role of cytosolic Ca2+ and mitochondrial Ca2+ during glutamate excitotoxicity F. The effects of NHE-1 inhibitors on glutamate-induced reduction in mitochondrial membrane potential and cytochrome c release G. The effects of NHE-1 inhibitors on glutamate-induced ROS accumulation H. The effects of FCCP on glutamate-induced neuronal cell death I. The expression of NHE-1 in neurons induced by glutamate exposure J. The change of activation of PKCs and MAPKs during glutamate exposure in neurons K. The change of NHE-1 activity during glutamate stimulation in neurons L. The role of p90RSK during glutamate-induced NHE-1 activation in cortical neurons M. The role of NHE-1 phosphorylation in glutamate-induced neuronal death N. The signaling of NHE-1 activation induced by PKC-β activation O. The signaling of NHE-1 activation induced by PKC-β activation in COS-1 cells P. The signaling of NHE-1 activation induced by PKC-β activation in SK-N-MC cells Q. The relationship between acidosis and NHE-1 phosphorylation during glutamate excitotoxicity in cortical neuronal cells IV. DISCUSSION V. CONCLUSION REFERENCES 국문요약 | - |
| dc.language.iso | eng | - |
| dc.publisher | The Graduate School, Ajou University | - |
| dc.rights | 아주대학교 논문은 저작권에 의해 보호받습니다. | - |
| dc.title | Glutamate로 유도된 뇌신경세포 사멸과정에서 Na+/H+ exchanger-1의 역할 | - |
| dc.title.alternative | The role of Na+/H+ exchanger-1 activation in cortical neurons during glutamate excitotoxicity | - |
| dc.type | Thesis | - |
| dc.contributor.affiliation | 아주대학교 일반대학원 | - |
| dc.contributor.alternativeName | Bo Kyung Lee | - |
| dc.contributor.department | 일반대학원 신경과학기술과정 | - |
| dc.date.awarded | 2010. 8 | - |
| dc.description.degree | Master | - |
| dc.identifier.localId | 568755 | - |
| dc.identifier.url | http://dcoll.ajou.ac.kr:9080/dcollection/jsp/common/DcLoOrgPer.jsp?sItemId=000000010884 | - |
| dc.subject.keyword | Na+/H+ exchanger-1 | - |
| dc.subject.keyword | 신경세포 | - |
| dc.subject.keyword | Glutamate | - |
| dc.subject.keyword | 세포사멸 | - |
| dc.description.alternativeAbstract | During brain ischemia, glutamate has excitotoxic effects to neurons through excessive activation of its receptor. Na+/H+ exchangers-1 (NHE-1) activity has been known to play a critical role in neuronal injury during glutamate excitotoxicity. It has been recently reported that inhibition of NHE-1 during ischemia/reperfusion injury has protective effects in brain. However, the protective mechanism of NHE-1 inhibition has not been clearly elucidated during glutamate excitotoxicity. In the present study, inhibition of NHE-1 with the potent inhibitors, cariporide and KR-33028 reduced glutamate-induced apoptotic and necrotic cell death in cortical neuronal cells. During glutamate exposure, dual peaks of cytosolic and mitochondrial Ca2+ rise were observed, and NHE-1 inhibitors did not suppress the initial peak whereas reduced the secondary peak. NHE-1 inhibitors also inhibited dissipation of mitochondrial membrane potential, cytochrome c release and ROS accumulation following glutamate exposure, suggesting its inhibitory effect on glutamate-mediated mitochondrial death pathways. In this study, I further demonstrated that the NHE-1 activity following glutamate was regulated by protein kinase C-β (PKC-β) in neuronal cells. The activation of PKC-β was significantly increased during glutamate stimulation, and NHE-1 activity and its phosphorylation during glutamate exposure was attenuated by the PKC-β inhibition with Go6976. I also examined the extracellular signal-related kinases (ERK)-p90kDa ribosomal S6 kinase (p90RSK) signaling pathway. At glutamate exposure, phosphorylated NHE-1 was increased with a concurrent elevation of phosphorylation of ERK and p90RSK. Inhibition of ERK and p90RSK activity with U0126 and SL0101 abolished activity and phosphorylation of NHE-1, respectively. Moreover, neuroprotection was observed with Go6976, U0126 and SL0101 following glutamate stimulation, respectively. Taken together, these results suggest that activation of PKC-β-ERK-p90RSK pathways following glutamate phosphorylates NHE-1 and increases its activity, which subsequently contributes to neuronal apoptotic and necrotic cell death via mitochondrial death pathway. I conclude that targeted inhibition of PKC-β-ERK-p90RSK pathway and reduction of NHE-1 activity in response to agonists such as glutamate is a novel strategy to brain ischemic injury. | - |
- Appears in Collections:
- Graduate School of Ajou University > Department of Neuroscience and Technology Course > 3. Theses(Master)
- Files in This Item:
- There are no files associated with this item.
Items in DSpace are protected by copyright, with all rights reserved, unless otherwise indicated.
-
- License
- STATISTICS
- Total Visit :3,767,882
- Total Download :1,819
- Today View :4,383
