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目的:探讨加味五子衍宗方总黄酮成分对Aβ25-35作用后的大鼠海马CA1区锥体神经元电压门控钙通道电流的作用。方法:应用全细胞膜片钳技术记录海马神经元ICa电流,采用内向电流的幅度作为观察指标,观察模型药物及干预药物对其的作用。采用出生7d的乳鼠取海马后制成脑片,选取立体感强、状态好的锥体细胞形成全细胞膜片钳记录,从-50mV到+50mV范围(步进为5mV)一系列去极化电压脉冲刺激(脉冲宽度500ms),以激发出电压依赖性的内向钙离子电流。记录电流后加入Aβ25-35后,再加入总黄酮进行干预。结果:Aβ25-35淀粉蛋白可以通过增强ICa可引起胞内钙超载,正常幅度平均为-(157.1±19.9)pA,Aβ25-35作用后增大为-(323.2±23.4)pA,125,250,500mg.L-1的总黄酮作用之后,分别下降为-(257.9±31.6),-(196.4±29.8),-(169.3±34.0)pA。结论:引起胞内钙离子超载可能是Aβ25-35产生神经毒性作用的机制之一,加味五子衍宗方总黄酮成分可通过抑制钙离子内流对神经元起一定保护作用。
OBJECTIVE: To investigate the effects of total flavonoids in Jiawei Wuzi Yanzongfang on the voltage-gated calcium channel currents of pyramidal neurons in hippocampal CA1 region after Aβ25-35 treatment. Methods: Whole cell patch clamp technique was used to record the ICa current of hippocampal neurons. The amplitude of inward current was used as the observation index to observe the effect of model drug and intervention drug. A 7-day-old neonate was used to take a hippocampus and a brain slice was made. A pyramidal cell with strong stereoscopic effect and a good condition was selected to form a whole-cell patch clamp recording. A series of depolarizations was performed from -50 mV to +50 mV range (stepping to 5 mV). Voltage pulse stimulation (pulse width 500 ms) to excite voltage-dependent inward calcium current. Recorded current after adding Aβ25-35, then add total flavonoids to intervene. RESULTS: Aβ25-35 amyloid protein could induce intracellular calcium overload by enhancing ICa. The normal amplitude was -(157.1±19.9) pA, after Aβ25-35 was increased to -(323.2±23.4)pA, 125, 250, 500 mg.L. After the total flavonoids of -1 were reduced to -(257.9±31.6), -(196.4±29.8), -(169.3±34.0)pA, respectively. Conclusion: Inducing intracellular calcium overload may be one of the mechanisms of neurotoxicity induced by Aβ25-35. Modified Fragrance of Wuzi Yanzongfang can protect the neurons by inhibiting calcium influx.