经颅磁刺激对海马原代神经元突起生长的影响

The Influences of Transcranial Magnetic Stimulation on the Growth of Hippocampus Primary Neuronal Processes

目的观察经颅磁刺激(transcranialmagnetic stimulation,TMS)对体外培养的海马原代神经元突起生长及形态的影响,并初步探讨磁刺激对细胞生长发育的影响机制。方法体外培养的海马原代神经元分为正常培养组(C组),假刺激组(S组),40%最大刺激强度组(M1组),60%最大刺激强度组(M2组),培养细胞24 h后开始磁刺激,每天固定时间刺激,刺激的频率为1 Hz,磁刺激最大输出强度为1.9 T,每天刺激3次,每次20序列,每个序列间隔1 s,每次间隔1 m in,连续刺激5 d,刺激源距离细胞1.0 cm。刺激结束后进行免疫荧光染色,观察原代神经元突起形态学改变,比较突起生长数目的改变及SYN-I表达的改变。结果 TMS 5 d后观察细胞生长状态良好,细胞与细胞间联系较多,部分细胞表现为多突起生长。M1、M2组2个突起的神经元占总神经元的比例分别为(45.6±14.9)%和(45.2±15.6)%,较C组和S组明显增多,差异有统计学意义(P<0.05);3个及3个以上突起的神经元占总神经元的比例分别为(30.3±10.8)%和(29.3±11.5)%,较C组和S组亦明显增多,差异有统计学意义(P<0.05)。M2组的SYN-ImRNA的表达较C、S两组增高,差异有统计学意义(P<0.05)。结论 TMS可以促进体外培养的海马原代神经元突起的生长。

Objective Application of transcranial magnetic stimulation（TMS） on primary cultured neurons,to observe the impact of TMS on the growth of neurons processes and morphology of synaptic and the mechanism.Methods The routine methods were used to extract hippocampus primary culture from Kunming mouse 1 day after birth.The hippocampus neuron were planted in culture dish of 35mm There are four groups：control group（C）,sham stimulation group（S）,40% of maximum intensity of stimulation（M1）,60% of maximum intensity of stimulation（M2）.The cells was stimulated at the same time each day.Stimulus is divided into three series.Each series has 20 sequences.Interval of each sequence is 1 second.Interval of each series is 1minute,continuous stimulation 5 days.Stimulation frequency of 1 Hz,stimulus intensity reference to experimental groups.Cellular immunofluorescence staining of MAP-2 was executed in the fifth day after stimulation.The growth state of cells were observed by fluorescence microscope and counted.Expression of Synapsin I mRNA were detected by using RT-RCR technology.Results Contact between processes and processes were significantly increased in each group after 5 days.Some contact formed like net.The percentage of two processes neurons：The percentage of two processes of neurons in M1 and M2 groups（M1 group（45.6±14.9）%;M2 group（45.2±15.6）%）were more than that in C and S groups（P0.05）.The percentage of three and above of processes of neurons：The percentage of three and above of processes of neurons in M1 and M2 groups（M1 group（30.3±10.8）%;M2 group（29.3±11.5）%） were significantly more than that in C and S groups（P0.05）.Expression of SYN-I mRNA in hippocampus primary neurons：Compared with C and S groups,the relative optical densities（ROD） of SYN-I bands in M2 group became higher（P0.05）.There was no statistically significant difference between M1 and other groups（P0.05）.Conclusion In this study,the result was sugeusted that the growth of neurons processes and morphology of synaptic was improved by transcranial magnetic stimulation in hippocampus primary neurons.