炎症大鼠下丘脑弓状核神经元敏感化的NMDA受体机制
发布时间：2019-03-24 10:53
【摘要】：研究目的：本实验旨在研究在外周组织炎症时下丘脑弓状核（arcuate nucleus ofhypothalamus，ARC）产生中枢敏感化的NMDA受体机制和受体后信号转导途径。 研究方法：建立完全弗氏佐剂（complete Freund’s adjuvant，CFA）炎症痛模型；用3管玻璃微电极在体细胞外记录ARC神经元的自发放电及其对伤害性刺激的反应；微电泳MK-801（NMDA受体非竞争性拮抗剂）、PP2（Src家族蛋白酪氨酸激酶抑制剂）、Chelerythrine（蛋白激酶C抑制剂），观察阻断NMDA受体或受体后激酶Src和PKC的磷酸化对大鼠ARC神经元自发放电和痛诱发放电的影响。 研究结果：（1）CFA炎症组大鼠ARC神经元的自发放电频率明显高于生理盐水对照组（Saline）大鼠，前者的放电频率为2.17±0.22Hz，后者为3.63±0.25Hz，两者比较具有显著的统计学意义（P0.01）；（2）CFA炎症组大鼠ARC中痛兴奋型神经元（PEN, pain-excitatory neuron）的痛诱发放电频率明显高于对照组，前者的放电频率为3.92±0.19Hz，后者为4.95±0.29Hz，两者比较具有显著的统计学意义（P0.01）；（3）微电泳MK-801（3mM，+20~30nA，30s）、PP2（1mM，+20~30nA，10~15min）和Chelerythrine（2mM，，+20~30nA，10~15min）可抑制CFA炎症组大鼠ARC中PENs的自发放电，放电频率分别降低了52.36%（P0.001）、25.5%（P0.05）和25%（P0.05），比较给药前后放电频率，皆具有统计学意义；微电泳相同药物对对照组大鼠的自发放电频率不产生影响。（4）微电泳MK-801（3mM，+20~30nA，30s）、PP2（1mM，+20~30nA，10~15min）和Chelerythrine（2mM，+20~30nA，10~15min）可抑制CFA炎症组大鼠ARC中PENs的痛诱发放电，放电频率分别降低了89.15%（P0.001）、77.09%（P0.001）和54.65%（P0.001）；微电泳相同药物对对照组大鼠ARC中PENs的痛诱发放电频率也产生同样的抑制作用,放电频率分别降低了90.63%（P0.001）、79.18%（P0.001）和47.06%（P0.01）。 研究结论：（1）外周CFA炎症时，ARC作为脊髓上水平痛觉调节的一个高位中枢核团，其中的神经元可产生敏感化现象，表现为ARC中神经元的自发放电增加，PENs对伤害性刺激的反应增强,放电频率增加。（2）NMDA受体、Src和PKC参与了CFA大鼠ARC中PENs的敏感化，表现为MK-801、PP2和Chelerythrine可以抑制ARC中PENs自发放电的增加和对伤害性刺激反应的增强；（3）NMDA受体、Src和PKC参与了CFA大鼠和对照组大鼠对伤害性刺激的反应，表现为MK-801、PP2和Chelerythrine可以抑制其PENs痛诱发放电的频率。
[Abstract]:Objective: to investigate the central sensitive NMDA receptor mechanism and postreceptor signal transduction pathway in hypothalamic arcuate nucleus (arcuate nucleus ofhypothalamus,ARC) during peripheral inflammation. Methods: a complete Freund's adjuvant (complete Freund's adjuvant,CFA) model of inflammatory pain was established and the spontaneous discharges of ARC neurons and their responses to noxious stimuli were recorded in vitro with 3-tube glass microelectrodes. Microelectrophoresis MK-801 (NMDA receptor noncompetitive antagonist), PP2 (Src family protein tyrosine kinase inhibitor), Chelerythrine (protein kinase C inhibitor). To observe the effect of blocking phosphorylation of NMDA receptor or post-receptor kinase Src and PKC on spontaneous and pain-induced discharges of ARC neurons in rats. Results: (1) the spontaneous firing frequency of ARC neurons in the CFA inflammatory group was significantly higher than that in the saline control group (2.17 卤0.22Hz, 3.63 卤0.25Hz), and the spontaneous firing frequency in the inflammatory group was significantly higher than that in the saline control group (2.17 卤0.22Hz, 3.63 卤0.25Hz). There was significant statistical significance between the two groups (P0.01). (2) the frequency of pain-induced discharges of pain-excited neurons (PEN, pain-excitatory neuron) in ARC of rats with CFA inflammation was significantly higher than that of the control group (3.92 卤0.19 Hz, 4.95 卤0.29 Hz, respectively). There was significant statistical significance between the two groups (P0.01). (3) Micro-electrophoresis of MK-801 (3mm, 20x30nA, 30s), PP2 (1mm, 20x30nA, 10min) and Chelerythrine (2mm, 20mM, 20k30nA, 10min) inhibited the spontaneous discharge of PENs in ARC of CFA inflammatory rats, and the firing frequency was decreased by 52.36% (P0.001), respectively, and the spontaneous discharge of PENs in ARC was decreased by 52.36% (P0.001) in the inflammatory group of CFA, and the spontaneous discharge frequency was decreased by 52.36% (P0.001). 25.5% (P0.05) and 25% (P0.05). The same drugs had no effect on the spontaneous discharge frequency of the control rats. (4) MK-801 (3mm, 20k30nA, 30s), PP2 (1mm, 20mA, 10min) and Chelerythrine (2mm, 20mA, 30nA), respectively, had no effect on the spontaneous discharge frequency of the control rats. 10~15min could inhibit the pain-induced discharge of PENs in ARC of rats with CFA inflammation, and the firing frequency was decreased by 89.15% (P0.001), 77.09% (P0.001) and 54.65% (P0.001), respectively. The same drugs showed the same inhibitory effect on the pain-induced discharge frequency of PENs in the ARC of the control rats, which decreased by 90.63% (P0.001), 79.18% (P0.001) and 47.06% (P0.01), respectively, and decreased by 90.63% (P0.001), 79.18% (P0.001) and 47.06% (P0.01) respectively. Conclusion: (1) in peripheral CFA inflammation, ARC is a high central nucleus of pain regulation at the level of spinal cord, in which neurons can be sensitized, and the spontaneous discharges of neurons in ARC are increased. The response of PENs to noxious stimuli was enhanced and the discharge frequency was increased. (2) NMDA receptor, Src and PKC were involved in the sensitization of PENs in ARC of CFA rats, which showed MK-801,. PP2 and Chelerythrine could inhibit the increase of spontaneous discharge of PENs and the enhancement of nociceptive stimulation in ARC. (3) NMDA receptors, Src and PKC participated in the responses of CFA rats and control rats to noxious stimuli, which showed that MK-801,PP2 and Chelerythrine could inhibit the frequency of PENs pain-induced discharges.
【学位授予单位】：苏州大学
【学位级别】：硕士
【学位授予年份】：2012
【分类号】：R338
[Abstract]:Objective: to investigate the central sensitive NMDA receptor mechanism and postreceptor signal transduction pathway in hypothalamic arcuate nucleus (arcuate nucleus ofhypothalamus,ARC) during peripheral inflammation. Methods: a complete Freund's adjuvant (complete Freund's adjuvant,CFA) model of inflammatory pain was established and the spontaneous discharges of ARC neurons and their responses to noxious stimuli were recorded in vitro with 3-tube glass microelectrodes. Microelectrophoresis MK-801 (NMDA receptor noncompetitive antagonist), PP2 (Src family protein tyrosine kinase inhibitor), Chelerythrine (protein kinase C inhibitor). To observe the effect of blocking phosphorylation of NMDA receptor or post-receptor kinase Src and PKC on spontaneous and pain-induced discharges of ARC neurons in rats. Results: (1) the spontaneous firing frequency of ARC neurons in the CFA inflammatory group was significantly higher than that in the saline control group (2.17 卤0.22Hz, 3.63 卤0.25Hz), and the spontaneous firing frequency in the inflammatory group was significantly higher than that in the saline control group (2.17 卤0.22Hz, 3.63 卤0.25Hz). There was significant statistical significance between the two groups (P0.01). (2) the frequency of pain-induced discharges of pain-excited neurons (PEN, pain-excitatory neuron) in ARC of rats with CFA inflammation was significantly higher than that of the control group (3.92 卤0.19 Hz, 4.95 卤0.29 Hz, respectively). There was significant statistical significance between the two groups (P0.01). (3) Micro-electrophoresis of MK-801 (3mm, 20x30nA, 30s), PP2 (1mm, 20x30nA, 10min) and Chelerythrine (2mm, 20mM, 20k30nA, 10min) inhibited the spontaneous discharge of PENs in ARC of CFA inflammatory rats, and the firing frequency was decreased by 52.36% (P0.001), respectively, and the spontaneous discharge of PENs in ARC was decreased by 52.36% (P0.001) in the inflammatory group of CFA, and the spontaneous discharge frequency was decreased by 52.36% (P0.001). 25.5% (P0.05) and 25% (P0.05). The same drugs had no effect on the spontaneous discharge frequency of the control rats. (4) MK-801 (3mm, 20k30nA, 30s), PP2 (1mm, 20mA, 10min) and Chelerythrine (2mm, 20mA, 30nA), respectively, had no effect on the spontaneous discharge frequency of the control rats. 10~15min could inhibit the pain-induced discharge of PENs in ARC of rats with CFA inflammation, and the firing frequency was decreased by 89.15% (P0.001), 77.09% (P0.001) and 54.65% (P0.001), respectively. The same drugs showed the same inhibitory effect on the pain-induced discharge frequency of PENs in the ARC of the control rats, which decreased by 90.63% (P0.001), 79.18% (P0.001) and 47.06% (P0.01), respectively, and decreased by 90.63% (P0.001), 79.18% (P0.001) and 47.06% (P0.01) respectively. Conclusion: (1) in peripheral CFA inflammation, ARC is a high central nucleus of pain regulation at the level of spinal cord, in which neurons can be sensitized, and the spontaneous discharges of neurons in ARC are increased. The response of PENs to noxious stimuli was enhanced and the discharge frequency was increased. (2) NMDA receptor, Src and PKC were involved in the sensitization of PENs in ARC of CFA rats, which showed MK-801,. PP2 and Chelerythrine could inhibit the increase of spontaneous discharge of PENs and the enhancement of nociceptive stimulation in ARC. (3) NMDA receptors, Src and PKC participated in the responses of CFA rats and control rats to noxious stimuli, which showed that MK-801,PP2 and Chelerythrine could inhibit the frequency of PENs pain-induced discharges.
【学位授予单位】：苏州大学
【学位级别】：硕士
【学位授予年份】：2012
【分类号】：R338
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