大鼠海马CCK阳性、阴性神经元电生理特性对比观察

摘要

目的 观察大鼠海马胆囊收缩素(CCK)阳性(CCK+)、阴性(CCK-)神经元电生理学特性的差异.方法 用CCK启动子-绿色荧光蛋白(GFP)的腺病毒标记CCK+神经元与未被标记的CCK-神经元(分别为CCK+组及CCK-组),观察细胞形态,检测静息状态下的静息电位(Vm)、膜阻抗(Rm)、膜电容(Cm)、时间常数(Tau)；用100 pA(500 ms)去极化电流诱发CCK+神经元和CCK-神经元的动作电位,记录动作电位的频率、时间间隔(ISI)、幅值(Amp)、动作电位达幅值中点的时间(HW)、动作电位上升(10％～90％)的时间(RT)、动作电位下降(90％ ～ 10％)的时间(DT)和发放频率；在灌流外液中加入50 μM AP-5和5μM NBQX,记录细胞自发抑制性突触后电流(sIPSC)的频率.结果 CCK+组细胞长径、短径、胞体面积均大于CCK-组(P均＜0.05).CCK+组细胞静息状态下Rm大于CCK-组,Tau小于CCK-组,P均＜0.05.两组Vm、Cm比较无统计学差异.CCK+组动作电位频率、HW大于CCK-组,ISI、RT小于CCK-组,P均＜0.05.两组Amp比较无统计学差异.CCK+组神经元sIPSC的发生频率高于CCK-组,P＜0.05.结论 CCK+神经元的传递速度快于CCK-神经元.%Objective To observe the electrophysiological characteristics divergences of CCK positive and negative neurons on cultured rat hippocampus.Methods Using the CCK promoter-green fluorescent protein (GFP) of adenovirus labeled neurons as CCK +,and unlabeled as CCK-.The aim was to detect morphology,the resting potential (Vm),membrane resistance (Rm),membrane capacitance (Cm),time constant (Tau) of these two cells; and the neurons action potentials were evoked by 100 pA (500 ms) depolarizing current.Then recording the value:action potential frequency,interevent interval (ISI),amplitude (Amp),the action potential half-width was the time between two points at the midpoint of the peak amplitude from the AP threshold(HW),the action potential rise time (10 ～ 90％,RT),the action potential decay time (90 ～ 10％,DT) and firing frequency.The perfusion extracellular fluid was filled with 50 μM AP-5 and 5 μM NBQX and then the spontaneous inhibitory postsynaptic current (sIPSC) was recorded.Results The long diameter in CCK + cells were (22.85 ± 0.59) μm,short diameter (15.37 ± 0.30) μm,cell body area (347.61 ±12.12) μm2.The value in CCK-group were (20.62 ± 0.51) μm,(14.24 ± 0.43) μm,(303.23 ± 14.30) μm2.The long diameter,short diameter and soma size of the two groups all had significant difference,P ＜0.05.The CCK + group:Vm (-49.16 ±1.94) mV,Cm (42.86 ± 1.78) pF,Rm (601.5 ±35.71) MΩ,Tau (221.08 ±35.63) μs; the CCK-group were (-48.81 ±1.81) mV,(47.85 ±2.33) pF,(504.66 ±31.08) MΩ,(500.32 ±40.27) μs.The Rm and Tau of CCK +neurons compared to CCK-cells,P ＜0.05,but Vm and Cm had no obvious difference between the two group,P ＞0.05.The action potential frequency of CCK + cells were (31.7 ± 1.89) Hz,ISI as (31.94 ± 0.57) ms,Amp (74.07 ±3.16) mV,HW (2.60±0.38) ms,RT (7.44±0.84) ms,DT(3.51 ±0.73) ms; CCK-group (23.8±3.64) Hz,(34.47 ± 1.0) ms,(78.77 ± 2.95) mV,(1.80 ± 0.17) ms,(10.49 ± 1.23) ms,(1.76 ± 0.28) ms.Compared with CCK-group,the frequencies,ISI,HW,RT,DT of CCK + group,P ＜ 0.05,Amp P ＞ 0.05.The sIPSC frequency of two group were (5.33 ±0.73) Hz,and (1.40 ±0.15) Hz,P ＜0.05.Conclusion The speed of transmission of CCK +neurons is faster than CCK-neurons