神经节苷脂GM1对大鼠脑缺血的保护作用及其机制的研究
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摘要
脑血管病是多发病，具有很高的死亡率和致残率，但是，临床有一级可信度的治疗脑血管病的药物很少。神经节苷脂GM1有保护脑缺血的作用，本研究探讨GM1的保护脑缺血的作用和机制。本研究将从整体、脑片水平评价GM1对大鼠局灶脑缺血、离体大鼠海马脑片缺氧缺糖的保护作用，分析GM1与神经元缺血缺氧损伤后N-甲基-D-门冬氨酸(N-methyl-D-aspartate，NMDA)受体亚单位和蛋白激酶C表达的关系；并在细胞水平评价内源性神经节苷脂对神经元兴奋性毒性损伤和去血清损伤的保护作用。
第一部分 神经节苷脂GM1保护大鼠局灶性脑缺血再灌注损伤的作用及其机制
目的：在整体水平，观察GM1对大鼠局灶性脑缺血再灌注损伤是否具有保护作用，以及保护作用是否与NMDA受体亚单位有关。方法：采用线栓法制作SD大鼠大脑中动脉阻塞和再灌注模型，脑缺血1h后再灌注72h进行神经体征评分。在脑缺血后不同时间(缺血后5min、1h和2h)腹腔注射GM1 10mg/kg，用TTC染色显示再灌注72h的梗塞灶，计算梗塞灶体积。用免疫印迹技术测定脑缺血后不
浙江大学博士学位论文
同再灌注时间点（4、6、24、48和72 h）梗塞半球的N－甲基－D／ 冬氨酸
（N－methyl－D也spartate，NMDA）受体亚单位 NRI、NRZA和 NRZB蛋白的含量。
结果：（）在脑缺血 lh再灌注 72 h，缺血后 5 min给予 GM的相对脑梗死体积
显著小于缺血后 2 h给药组和对照组0＜0．of，one－way ANOVA人 缺血后 lh给
药组的相对脑梗死体积显著小于缺血后 2 h给药组和对照组0＜0刀5，＊eWay
＊＊OVA人而缺血后Zh给药组的相对的脑梗死体积与对照组无显著差异（P＞0刀5，
Ofl卜W叮 ANOVA）；缺血后 5 iin给药组的矫正的相对脑梗死灶体积显著小于对照
组和缺血后 2 h给药组0＜0．of，one卅ay ANO“人 缺血后 lh给药组的矫正的
相对脑梗死灶体积显著小于对照组和缺血后 2 h给药组（P＜0刀5，OnOway
ANOVA）；而缺血后 2 h给药组和对照组间的矫正的相对脑梗死体积无显著差异0
＞0刀5，oneWay ANOVA人各组大鼠在手术清醒后的神经体征评分无显著差异o＞
0刀5，oneWay ANOVA人脑缺血再灌注 6 h后 TTC染色开始出现梗死灶，逐渐增
大，到再灌注72 h达到高峰。脑缺血再灌注6 h时，甲苯胺蓝染色显示缺血侧皮
层和基底节区神经元变性、缺失。（2）梗死半球的NRI、NRZA和 NMB在再灌
注 6 h出现一过性的高表达；在再灌注 48～72 h有明显下降；脑缺血后 5 min使用
GMI （l mg／kg）可以显著降低再灌注 6 h时 NRI的表达（P＜0刀5 vs对照组，
Tiest入 再灌注 72 h的缺血半球的 NRI的表达显著低于对照组（P＜0刀5，Tiest入
使用 GMI（脑缺血后 5 min、2 h）组的 NRI的表达与对照组无显著差异（P＞0刀5，
Tiest）。结论：神经节昔脂 GM在大鼠局灶性脑缺血后早期用药（＜lh），可以减
少脑梗死体积。而晚期用药（2 h）则不能减少脑梗死体积。GM抑制脑缺血再灌
注后6h出现的NRI的一过性高表达，并防止再灌注72h时NRI的过度降低，
这可能是 GM减小脑梗死体积的机制之一。
Cerebrovascular disease is a source of mortality and profound morbidity that remains pervasive in the modern world. But effective therapeutic drug is few. Monoganglioside (GM1) is reported as a potential therapeutic drug for cerebral ischemia. To determine the protective effect and mechanism(s) of GM1 on cerebral ischemic injury, the effects of GM1 on neural injuries induced by focal cerebral ischemia in rats and by oxygen glucose deprivation in rat hippocampal slices were evaluated in this study. Influence of GM1 on expression of N-methyl-D-aspartate receptor subunits and protein kinase C were analyzed to clarify the possible mechanism. Effect of endogenous ganglioside on exitoxicity and serum deprivation injury on culture neural cell was observed as well.
Part Ⅰ Protective effect and mechanism of monosialoganglionside (GM1) on
injury induced by focal cerebral ischemia and reperfusion in rats
Purpose: In the in vivo experiments, we determined whether GM1 has a protective effect on injury induced by focal cerebral ischemia and reperfusion and the possible
mechanism in rats. Methods: Middle cerebral artery (MCA) occlusion was performed to induce focal brain ischemia / reperfusion model by an intraluminal filament in SD rats. GM1 was given i.p. at different time point (5 min, 1 h and 2 h after ischemia). MCA was occluded for 1 h and the brain was reperfused for 12 h at that moment neurologic deficiency scores were assessed, and infarct volume was measured by TTC staining. Expression levels of NMD A receptor subunit NR1, NR2A and NR2B were detected by Western blot at reperfusion time points (4, 6, 24, 48 and 72 h) after cerebral ischemia. Results: (1) Relative infarct volume of was significantly smaller in the group administered 5 min after ischemia than that 2 h after ischemia or control (P < 0.01, one-way ANOVA). Relative infarct volume of in group administered 1 h after ischemia was significantly smaller than that of group 2 h after ischemia or control (P < 0.05, one-way ANOVA). There was no significant difference between relative infarct volume of group administered 2 h after ischemia and that of control (P > 0.05, one-way ANOVA). Adjusted relative infarct volume of group administered 5 min after ischemia was significantly smaller than that of group 2 h after ischemia and control (P < 0.01, one-way ANOVA). Adjusted relative infarct volume of group administered 1 h after ischemia was significantly smaller than that of 2 h after ischemia or control (P < 0.05, one-way ANOVA). There was not significant difference between adjusted relative infarct volume of group administered at 2 h after ischemia and that of control (P > 0.05, one-way ANOVA). TTC staining showed cerebral infarction at 6 h after reperfusion, and infarct volume gradually increased, and reached the peak at 12 h after reperfusion. Toluidine blue staining 6 h after reperfusion showed nerve cells degenerated and being missed in the cortical and basoganglic area of ischemic hemisphere. (2) Expression levels of NR1, NR2A and NR2B were elevated temporally 6 h after reperfusion, and came below normal levels at 48 ~ 72 h after reperfusion. GM1 given i.p. 5 min after ischemia significantly suppressed the expression of NR1 6 h after reperfusion (P < 0.05 vs control, T-test). NR1 expression of ischemic hemisphere 72 h after reperfusion was significantly lower than that of the control (P < 0.05, T-test), There was no significant difference between expression of NR1 in ischemic hemispheres of control and those accepted GM1 (5 min and 2 h after ischemia, 10 mg/kg, i.p.) (P > 0.05, T-test).
Conclusion: Early used GM1 (< 1 h after ischemia) significantly reduces the cerebral infarct volume, but late used GM1 (2 h after ischemia) does not. One of the possible mechanisms for GM1 reducing cerebral infarct volume may be probably that early used GM1 can suppress temporally over-expression of NRl in the ischemic hemisphere 6 h after reperfusion and depressed-expression of NRl 72 h after reperfusion.
第一部分 神经节苷脂GM1保护大鼠局灶性脑缺血再灌注损伤的作用及其机制
目的：在整体水平，观察GM1对大鼠局灶性脑缺血再灌注损伤是否具有保护作用，以及保护作用是否与NMDA受体亚单位有关。方法：采用线栓法制作SD大鼠大脑中动脉阻塞和再灌注模型，脑缺血1h后再灌注72h进行神经体征评分。在脑缺血后不同时间(缺血后5min、1h和2h)腹腔注射GM1 10mg/kg，用TTC染色显示再灌注72h的梗塞灶，计算梗塞灶体积。用免疫印迹技术测定脑缺血后不
浙江大学博士学位论文
同再灌注时间点（4、6、24、48和72 h）梗塞半球的N－甲基－D／ 冬氨酸
（N－methyl－D也spartate，NMDA）受体亚单位 NRI、NRZA和 NRZB蛋白的含量。
结果：（）在脑缺血 lh再灌注 72 h，缺血后 5 min给予 GM的相对脑梗死体积
显著小于缺血后 2 h给药组和对照组0＜0．of，one－way ANOVA人 缺血后 lh给
药组的相对脑梗死体积显著小于缺血后 2 h给药组和对照组0＜0刀5，＊eWay
＊＊OVA人而缺血后Zh给药组的相对的脑梗死体积与对照组无显著差异（P＞0刀5，
Ofl卜W叮 ANOVA）；缺血后 5 iin给药组的矫正的相对脑梗死灶体积显著小于对照
组和缺血后 2 h给药组0＜0．of，one卅ay ANO“人 缺血后 lh给药组的矫正的
相对脑梗死灶体积显著小于对照组和缺血后 2 h给药组（P＜0刀5，OnOway
ANOVA）；而缺血后 2 h给药组和对照组间的矫正的相对脑梗死体积无显著差异0
＞0刀5，oneWay ANOVA人各组大鼠在手术清醒后的神经体征评分无显著差异o＞
0刀5，oneWay ANOVA人脑缺血再灌注 6 h后 TTC染色开始出现梗死灶，逐渐增
大，到再灌注72 h达到高峰。脑缺血再灌注6 h时，甲苯胺蓝染色显示缺血侧皮
层和基底节区神经元变性、缺失。（2）梗死半球的NRI、NRZA和 NMB在再灌
注 6 h出现一过性的高表达；在再灌注 48～72 h有明显下降；脑缺血后 5 min使用
GMI （l mg／kg）可以显著降低再灌注 6 h时 NRI的表达（P＜0刀5 vs对照组，
Tiest入 再灌注 72 h的缺血半球的 NRI的表达显著低于对照组（P＜0刀5，Tiest入
使用 GMI（脑缺血后 5 min、2 h）组的 NRI的表达与对照组无显著差异（P＞0刀5，
Tiest）。结论：神经节昔脂 GM在大鼠局灶性脑缺血后早期用药（＜lh），可以减
少脑梗死体积。而晚期用药（2 h）则不能减少脑梗死体积。GM抑制脑缺血再灌
注后6h出现的NRI的一过性高表达，并防止再灌注72h时NRI的过度降低，
这可能是 GM减小脑梗死体积的机制之一。
Cerebrovascular disease is a source of mortality and profound morbidity that remains pervasive in the modern world. But effective therapeutic drug is few. Monoganglioside (GM1) is reported as a potential therapeutic drug for cerebral ischemia. To determine the protective effect and mechanism(s) of GM1 on cerebral ischemic injury, the effects of GM1 on neural injuries induced by focal cerebral ischemia in rats and by oxygen glucose deprivation in rat hippocampal slices were evaluated in this study. Influence of GM1 on expression of N-methyl-D-aspartate receptor subunits and protein kinase C were analyzed to clarify the possible mechanism. Effect of endogenous ganglioside on exitoxicity and serum deprivation injury on culture neural cell was observed as well.
Part Ⅰ Protective effect and mechanism of monosialoganglionside (GM1) on
injury induced by focal cerebral ischemia and reperfusion in rats
Purpose: In the in vivo experiments, we determined whether GM1 has a protective effect on injury induced by focal cerebral ischemia and reperfusion and the possible
mechanism in rats. Methods: Middle cerebral artery (MCA) occlusion was performed to induce focal brain ischemia / reperfusion model by an intraluminal filament in SD rats. GM1 was given i.p. at different time point (5 min, 1 h and 2 h after ischemia). MCA was occluded for 1 h and the brain was reperfused for 12 h at that moment neurologic deficiency scores were assessed, and infarct volume was measured by TTC staining. Expression levels of NMD A receptor subunit NR1, NR2A and NR2B were detected by Western blot at reperfusion time points (4, 6, 24, 48 and 72 h) after cerebral ischemia. Results: (1) Relative infarct volume of was significantly smaller in the group administered 5 min after ischemia than that 2 h after ischemia or control (P < 0.01, one-way ANOVA). Relative infarct volume of in group administered 1 h after ischemia was significantly smaller than that of group 2 h after ischemia or control (P < 0.05, one-way ANOVA). There was no significant difference between relative infarct volume of group administered 2 h after ischemia and that of control (P > 0.05, one-way ANOVA). Adjusted relative infarct volume of group administered 5 min after ischemia was significantly smaller than that of group 2 h after ischemia and control (P < 0.01, one-way ANOVA). Adjusted relative infarct volume of group administered 1 h after ischemia was significantly smaller than that of 2 h after ischemia or control (P < 0.05, one-way ANOVA). There was not significant difference between adjusted relative infarct volume of group administered at 2 h after ischemia and that of control (P > 0.05, one-way ANOVA). TTC staining showed cerebral infarction at 6 h after reperfusion, and infarct volume gradually increased, and reached the peak at 12 h after reperfusion. Toluidine blue staining 6 h after reperfusion showed nerve cells degenerated and being missed in the cortical and basoganglic area of ischemic hemisphere. (2) Expression levels of NR1, NR2A and NR2B were elevated temporally 6 h after reperfusion, and came below normal levels at 48 ~ 72 h after reperfusion. GM1 given i.p. 5 min after ischemia significantly suppressed the expression of NR1 6 h after reperfusion (P < 0.05 vs control, T-test). NR1 expression of ischemic hemisphere 72 h after reperfusion was significantly lower than that of the control (P < 0.05, T-test), There was no significant difference between expression of NR1 in ischemic hemispheres of control and those accepted GM1 (5 min and 2 h after ischemia, 10 mg/kg, i.p.) (P > 0.05, T-test).
Conclusion: Early used GM1 (< 1 h after ischemia) significantly reduces the cerebral infarct volume, but late used GM1 (2 h after ischemia) does not. One of the possible mechanisms for GM1 reducing cerebral infarct volume may be probably that early used GM1 can suppress temporally over-expression of NRl in the ischemic hemisphere 6 h after reperfusion and depressed-expression of NRl 72 h after reperfusion.
引文
1. Duchemin AM, Ren Q, Mo L, Neff NH, Hadjiconstantinou M. GM1 ganglioside induces phosphorylation and activation of Trk and Erk in brain. J Neurochem 2002; 81 (4):696-707
2. Komatsumoto S, Greenberg JH, Hickey WF, Reivich M.Effect of the ganglioside GM1 on neurologic function, electroencephalogram amplitude, and histology in chonic middle cerebral artery occlusion in cats. Stroke 1988; 19(8): 1027-35.
3. Kharlamov A, Guidotti A, Costa E, Hayes R, Armstrong D. Semisynthetic sphingolipids prevent protein kinase C translocation and neuronal damage in the perifocal area following a photochemically induced thombotic brain cortical lesion. J Neurosci 1993; 13(6):2483-2494.
4. Simon RP, Chen J, Graham SH.GM1 ganglioside treatment of focal ischemia: a dose-response and microdialysis study. J Pharmacol Exp Ther 1993; 265(1): 24-29.
5. Phillis JW, O'Regan MH.GM1 ganglioside inhibits ischemic release of amino acid neurotransmitters from rat cortex. Neuroreport 1995; 6(15):2010-2012.
6. Hicks D, Heidinger V, Mohand-Said S, Sahel J, Dreyfus H.Growth factors and gangliosides as neuroprotective agents in excitotoxicity and ischemia. Gen Pharmacol 1998; 30(3):265-273.
7. Avrova NF, Shestak KI, Zakharova IO, Sokolova TV, Tyurina YY, Tyurin VA. The use of antioxidants to prevent glutamate-induced derangement of calcium ion metabolism in rat cerebral cortex synaptosomes. Neurosci Behav Physiol 2000; 30(5):535-541.
8. Biggo G. Exicitatory Aminio Acids and Brain Ischemia. Oxford: Pergaman Press, 1990: 93-102.
9. Brandoli C, Sanna A, De Bernardi MA, Follesa P, Brooker G, Mocchetti I.Brain-derived neurotrophic factor and basic fibroblast growth factor downregulate NMDA receptor function in cerebellar granule cells.J Neurosci 1998; 18(19):7953-7961.
10. Ferrari G, Fabris M, Fiori MG, Gabellini N, Volonte C. Gangliosides prevent the
inhibition by K-252a of NGF responses in PC12 cells. Brain Res Dev Brain Res. 1992; 65(1):35-42.
11. Skaper SD, Katoh-Semba R, Varon S. GM1 ganglioside accelerates neurite outgrowth from primary peripheral and central neurons under selected culture conditions. Brain Res. 1985; 355(1): 19-26.
12. Fusco M, Oderfeld-Nowak B, Vantini G, Schiavo N, Gradkowska M, Zaremba M, Leon A. Nerve growth factor affects uninjured, adult rat septohippocampal cholinergic neurons. Neuroscience. 1989; 33 (1):47-52.
13. Cuello AC, Garofalo L, Kenigsberg RL, Maysinger D. Gangliosides potentiate in vivo and in vitro effects of nerve growth factor on central cholinergic neurons. Proc Natl Acad Sci U S A. 1989; 86(6):2056-2060.
14. Bederson JB, Pitts LH, Tsuji M, et al. Rat middle cerebral artery occlusion: evaluation of the model and development of a neurologic examination. Stroke 1986; 17(3): 472-476.
15．王伯沄，李玉松，黄高昇，张远强．病理学技术．人民卫生出版社，2000年6 月第一版，P17．
16. Luo J, Wang Y, Yasuda RP, Dunah AW, Wolfe BB. The majority of N-methyl-D-aspartate receptor complexes in adult rat cerebral cortex contain at least thee different subunits (NR1/NR2A/NR2B). Mol Pharmacol 1997; 51(1):79-86.
17. Luo J, Bosy TZ, Wang Y, Yasuda RP, Wolfe BB. Ontogeny of NMDA R1 subunit protein expression in five regions of rat brain. Brain Res Dev Brain Res 1996; 92(1):10-17.
18. Wang YH, Bosy TZ, Yasuda RP, Grayson DR, Vicini S, Pizzorusso T, Wolfe BB. Characterization of NMDA receptor subunit-specific antibodies: distribution of NR2A and NR2B receptor subunits in rat brain and ontogenic profile in the cerebellum. J Neurochem 1995; 65(1): 176-183.
19. Milani D, Guidolin D, Facci L, Pozzan T, Buso M, Leon A, Skaper SD. Excitatory amino acid-induced alterations of cytoplasmic free Ca2+ in individual cerebellar granule neurons: role in neurotoxicity. J Neurosci Res. 1991; 28(3):434-41.
20. Cai Z, Rhodes PG. Intrauterine hypoxia-ischemia alters expression of the NMDA receptor in the young rat brain. Neurochem Res 2001 May;26(5):487-495.
21. Kang TC, Hwang IK, Park SK, An S J, Yoon DK, Moon SM, Lee YB, Sohn HS, Cho SS, Won MH. Chronological changes of N-methyl-D-aspartate receptors and excitatory amino acid carrier 1 immunoreactivities in CA1 area and subiculum after transient forebrain ischemia. J Neurocytol 2001; 30(12):945-955.
22. Won MH, Kang T, Park S, Jeon G, Kim Y, Seo JH, Choi E, Chung M, Cho SS. The alterations of N-Methyl-D-aspartate receptor expressions and oxidative DNA damage in the CA1 area at the early time after ischemia-reperfusion insult. Neurosci Lett 2001; 301(2):139-142.
23. Friedman LK, Ginsberg MD, Belayev L, Busto R, Alonso OF, Lin B, Globus MY. Intraischemic but not postischemic hypothermia prevents non-selective hippocampal downregulation of AMPA and NMDA receptor gene expression after global ischemia. Brain Res Mol Brain Res 2001; 86(1-2):34-47.
24. Valdes-Gonzalez T, Morita Y, Suzuki K, Ido T. Excitotoxicity induces changes in rat brain gangliosides. Neurosci Res 2001; 39(2): 197-203.
25. Resink A, Villa M, Boer GJ, Mohler H, Balazs R. Agonist-induced down-regulation of NMDA receptors in cerebellar granule cells in culture. Eur J Neurosci 1995; 7(8): 1700-1706.
2. Komatsumoto S, Greenberg JH, Hickey WF, Reivich M.Effect of the ganglioside GM1 on neurologic function, electroencephalogram amplitude, and histology in chonic middle cerebral artery occlusion in cats. Stroke 1988; 19(8): 1027-35.
3. Kharlamov A, Guidotti A, Costa E, Hayes R, Armstrong D. Semisynthetic sphingolipids prevent protein kinase C translocation and neuronal damage in the perifocal area following a photochemically induced thombotic brain cortical lesion. J Neurosci 1993; 13(6):2483-2494.
4. Simon RP, Chen J, Graham SH.GM1 ganglioside treatment of focal ischemia: a dose-response and microdialysis study. J Pharmacol Exp Ther 1993; 265(1): 24-29.
5. Phillis JW, O'Regan MH.GM1 ganglioside inhibits ischemic release of amino acid neurotransmitters from rat cortex. Neuroreport 1995; 6(15):2010-2012.
6. Hicks D, Heidinger V, Mohand-Said S, Sahel J, Dreyfus H.Growth factors and gangliosides as neuroprotective agents in excitotoxicity and ischemia. Gen Pharmacol 1998; 30(3):265-273.
7. Avrova NF, Shestak KI, Zakharova IO, Sokolova TV, Tyurina YY, Tyurin VA. The use of antioxidants to prevent glutamate-induced derangement of calcium ion metabolism in rat cerebral cortex synaptosomes. Neurosci Behav Physiol 2000; 30(5):535-541.
8. Biggo G. Exicitatory Aminio Acids and Brain Ischemia. Oxford: Pergaman Press, 1990: 93-102.
9. Brandoli C, Sanna A, De Bernardi MA, Follesa P, Brooker G, Mocchetti I.Brain-derived neurotrophic factor and basic fibroblast growth factor downregulate NMDA receptor function in cerebellar granule cells.J Neurosci 1998; 18(19):7953-7961.
10. Ferrari G, Fabris M, Fiori MG, Gabellini N, Volonte C. Gangliosides prevent the
inhibition by K-252a of NGF responses in PC12 cells. Brain Res Dev Brain Res. 1992; 65(1):35-42.
11. Skaper SD, Katoh-Semba R, Varon S. GM1 ganglioside accelerates neurite outgrowth from primary peripheral and central neurons under selected culture conditions. Brain Res. 1985; 355(1): 19-26.
12. Fusco M, Oderfeld-Nowak B, Vantini G, Schiavo N, Gradkowska M, Zaremba M, Leon A. Nerve growth factor affects uninjured, adult rat septohippocampal cholinergic neurons. Neuroscience. 1989; 33 (1):47-52.
13. Cuello AC, Garofalo L, Kenigsberg RL, Maysinger D. Gangliosides potentiate in vivo and in vitro effects of nerve growth factor on central cholinergic neurons. Proc Natl Acad Sci U S A. 1989; 86(6):2056-2060.
14. Bederson JB, Pitts LH, Tsuji M, et al. Rat middle cerebral artery occlusion: evaluation of the model and development of a neurologic examination. Stroke 1986; 17(3): 472-476.
15．王伯沄，李玉松，黄高昇，张远强．病理学技术．人民卫生出版社，2000年6 月第一版，P17．
16. Luo J, Wang Y, Yasuda RP, Dunah AW, Wolfe BB. The majority of N-methyl-D-aspartate receptor complexes in adult rat cerebral cortex contain at least thee different subunits (NR1/NR2A/NR2B). Mol Pharmacol 1997; 51(1):79-86.
17. Luo J, Bosy TZ, Wang Y, Yasuda RP, Wolfe BB. Ontogeny of NMDA R1 subunit protein expression in five regions of rat brain. Brain Res Dev Brain Res 1996; 92(1):10-17.
18. Wang YH, Bosy TZ, Yasuda RP, Grayson DR, Vicini S, Pizzorusso T, Wolfe BB. Characterization of NMDA receptor subunit-specific antibodies: distribution of NR2A and NR2B receptor subunits in rat brain and ontogenic profile in the cerebellum. J Neurochem 1995; 65(1): 176-183.
19. Milani D, Guidolin D, Facci L, Pozzan T, Buso M, Leon A, Skaper SD. Excitatory amino acid-induced alterations of cytoplasmic free Ca2+ in individual cerebellar granule neurons: role in neurotoxicity. J Neurosci Res. 1991; 28(3):434-41.
20. Cai Z, Rhodes PG. Intrauterine hypoxia-ischemia alters expression of the NMDA receptor in the young rat brain. Neurochem Res 2001 May;26(5):487-495.
21. Kang TC, Hwang IK, Park SK, An S J, Yoon DK, Moon SM, Lee YB, Sohn HS, Cho SS, Won MH. Chronological changes of N-methyl-D-aspartate receptors and excitatory amino acid carrier 1 immunoreactivities in CA1 area and subiculum after transient forebrain ischemia. J Neurocytol 2001; 30(12):945-955.
22. Won MH, Kang T, Park S, Jeon G, Kim Y, Seo JH, Choi E, Chung M, Cho SS. The alterations of N-Methyl-D-aspartate receptor expressions and oxidative DNA damage in the CA1 area at the early time after ischemia-reperfusion insult. Neurosci Lett 2001; 301(2):139-142.
23. Friedman LK, Ginsberg MD, Belayev L, Busto R, Alonso OF, Lin B, Globus MY. Intraischemic but not postischemic hypothermia prevents non-selective hippocampal downregulation of AMPA and NMDA receptor gene expression after global ischemia. Brain Res Mol Brain Res 2001; 86(1-2):34-47.
24. Valdes-Gonzalez T, Morita Y, Suzuki K, Ido T. Excitotoxicity induces changes in rat brain gangliosides. Neurosci Res 2001; 39(2): 197-203.
25. Resink A, Villa M, Boer GJ, Mohler H, Balazs R. Agonist-induced down-regulation of NMDA receptors in cerebellar granule cells in culture. Eur J Neurosci 1995; 7(8): 1700-1706.