【作者】 张玲；

【导师】 孔德虎； 黄振信； 祝延；

【作者基本信息】 安徽医科大学 ， 生理学， 2002， 硕士

【摘要】 应用细胞内记录技术，观察银杏叶提取物（EGb）和银杏黄酮对豚鼠离体腹腔神经节（CG）细胞生物电及突触传递的影响，旨在从电生理学和药理学角度对其作用及其机制进行初步探讨。主要结果如下： 1．EGb对CG细胞生物电的影响 1．1 豚鼠CG细胞的静息电位为-50.5±2.5mV（n=200）；膜电阻55.2±3.8MΩ（n=100）。 1．2 EGb50～500ug/ml灌流CG细胞（n=44），其中有12个细胞（27.3％）不产生反应，32个细胞（72.7％）对银杏黄酮敏感。在敏感的CG细胞中，24个细胞（75.0％）为去极化反应，2个细胞（6.3％）出现双相反应，4个细胞（12.5％）为超极化反应，另有2个细胞（6.3％）反应不恒定，即在同样的灌流条件下或出现超极化反应或出现去极化反应。EGb引起的去极化反应表现为缓慢、持久，幅度5.6±0.8mV，时程240.7±50.3Sec，膜电阻可以减小（33.8％）、不变（40.0％）或增大（26.2％），说明EGb对CG细胞作用较为复杂。 1．3 用胆碱受体阻断剂阿托品(10^-4mol/L)、六烃季铵(10^-4mol/L)和肾上腺素受体阻断剂酚妥拉明(10^-4mol/L)、心得安(10^-4mol/L)灌流神经节，对EGb去极化反应不产生影响（n=8），提示这种去极化反应是通过非胆碱非肾上腺素受体途径实现的。低钙/高镁液可以部分阻抑EGb的去极化反应（n=7），使去极化幅度降低。提示EGb既能直接作用于突触后神经元也能影响突触前神经末梢释放某种递质。 2．银杏黄酮对CG细胞生物电和突触传递的影响及其机制的探讨 2．1 银杏黄酮10～100ug/ml灌流CG细胞，产生与EGb相似的结果（n=120）。55个细胞（45.8％）出现膜电位去极化，12个（10.0％）超极化，另有12个(10.0％ 安徽医科大学硕士学位论文 为双相反应，此外还有41个细胞（34．2％）几乎不发生反应。在去极化的细胞其 去极化的幅度7，9土0．7mV，时程245．9土29二h，随着灌流浓度的增加，去极化在 幅度亦随之增大，表现剂量效应关系。 2．2胆碱和肾上腺素受体阻断剂对银杏黄酮引起的去极化反应不产生影响。 低钙／高镁溶液灌流可以部分阻抑银杏黄酮的去极化反应。 2．3在银杏黄酮引起的去极化反应中＊叫20入42个细胞N6．7们膜电阻 减小，平均减小18．2％fZ．5％，27个细胞不变（30．0％），21个细胞增大（23．3％）， 膜电阻平均增大26．7 16．7兄表明银杏黄酮所引起的去极化反应涉及一种以上离 子的跨膜转s；用N－methyl－Dglucamine chloride （NM）代替Na“，以无钠的 克氏液灌流CG，可使银杏黄酮的去极化反应受到抑制，幅度减小；用BaClz代替 Ca’”，以无钙的克氏液灌流 CG，由银杏黄酮引起的去极化反应受到抑制；用 TEA 灌流 CG，可使银杏黄酮的去极化反应幅度增强，推测 Na”儿a’”内流的增加和 K“外 流的减少可能是银杏黄酮去极化产生的原因；将膜电位控制在不同水平＊＜0Xo 观察到 8个细胞抢0们，膜电位愈负，银杏黄酮的去极化反应幅度愈大，时程愈 长，膜逆转电位约在－30mV左右，亦证明银杏黄酮的去极化反应可能与若干离子 的跨膜转运有关。 2．4单脉冲刺激左侧内脏大神经，在比细胞引导出 卜E陀巳用银杏黄酮 10叶00ug加 灌流CG，在引起膜去极化的同时，可使卜EPSP的幅度增大并爆发 锋电位，而用低钙／高镁克氏液灌流 CG，可部分阻抑银杏黄酮易化突触传递的作用 （n＝20）。 3．银杏寅酮与其他递质的相互作用 用5－HT、BOM改变膜的兴奋状态，银杏黄酮可以抑制由5－HT、BOM等引起的 CG细胞的去极化反应。推测EGb和银杏黄酮对交感神经节细胞的作用可能与细胞 当时所处的机能状态有关。更多还原

【Abstract】 Using the intracellular recording technique and the administration of super-fusion, this work was going to investigate the effects of EGb and Flavonoids on bioelectric and synaptic transmitted of the neurons in celiac ganglia of guinea-pig in vitro. Eventually we hope to discover how EGb and its active composes will do on the sympathetic ganglia. The main experimental results are as follows: 1. The bioelectric effects of EGb on the neurons of CG1.1 The mean of resting potential of CG cell in guinea-pig is -50.5\2.5mV(n=200) and the membrane resistance 55.2 ± 3.8M .1.2 Superfusion of celiac ganglia with EGb(50~500mg/L) evoked a concentration-dependent membrane depolarization in the majority(75.0%) of the neurons(n=44), with the mean amplitude and the average duration are 5.6\0.8mV and 240.7\50Sec respectively. Among the cells tested, 12.5% of the cells was induced hyperpolarization and 6.3% biphasic response which consisted of an initial hyperpolarization phase and a followed depolarization. There are also 6.3% of the cells observed which show both depolarization and hyperpolarization at the different time under the same conditions. The neurons Rm could be changed during the depolarization accured with 33.8% of the cells decreased, 26.2% increased and unchanged in the rest 40.0%. These results imply that the EGb has complicated effects on the neurons of CG.1.3 Depolarizations of EGb could not be affected by cholinocepor and adrenoceptor inhibitors (n=8). Depolarizations of EGb could be blocked partially by low Ca2±/high Mg2"1" solution (n=7).2. The bioelectric and synaptic transmission effect of Flavonoids and the possiblemechanism2.1 Superfiision of celiac ganglia with Flavonoids (10~100mg/L) evoked the same effects as EGb did (n=120) on CG neurons. Among these cells 55(45.8%) presented depolarization , 12(10.0%) biphasic response and 41(34.2%) no responses. The average amplitude and duration of depolarization caused by Flavonoids are 7.9\0.7mv and 245.6 \29.2Sec, respectively. The depolarization effects caused by Flavonoids also has the character of concentration-dependent. The results above suggest that the effects of EGb on CG neurons is mainly caused by Flavonoids.2.2 Superfusion of celiac ganglia with 10-4 mol/L cholinoceptor (Atropine and Hexamethonium )and adrenoceptor inhibitor (Phentolamin and Propranolo) had no effects on the depolarization evoked by Flavonoids; Superfusion of celiac ganglia with Low Ca2±/High Mg2"1" solution, the depolarization caused by Flavonoids would be attenuated.2.3 Among 42 of the depolarization cells, the Rm could be decreased by 18.2\ 2.5% in 46.7%(n=27), unchanged in 30.0%, increased by 26.7\6.7 in the rest 23.3%. These results reveal that more than one kind of ion involves in the depolarization. Substituting Na* with NMG and then Superfusion of celiac ganglia with this Na± free solution would attenuate the depolarization caused by Flavonoids; Substituting Ca± with BaCl2 then superfusion of celiac ganglia with this Ca2± free solution would attenuate the depolarization caused by Flavonoids; Superfusion of celiac ganglia with K± inhibitor TEA, would augment the amplitude of depolarization caused by Flavonoids; These result suggest that Na± Ca2± K* are all the possible ions which involves in the depolarization induced by Flavonoids. The amplitude of depolarization evoked by Flavonoids changed as the membrane potential were controlled at different level (n=10). The depolarizations were attenuated in the majority cells (80%) when membrane potential decreased. The reversal potential for the event was about -30mV. These data futher elucidate that several kinds of ions , including Na\ Ca2±> K±, contribute to the formation of depolarization of Flavonoids.2.4 f-EPSP was initiated by stimulating left lesser splanchnic nerve with currentpulses. Then superfusion of celiac ganglia with Flavonoids (10~100ug/ml) would converted subthreshold PSP to action potentials, The effects更多还原