| Objective:With the wide application of microwave technology,it brings convenience to people.Meanwhile,its adverse reactions have also aroused people's attention.It could injure multiple functions of brain including the synaptic plasticity impairment and cause learning and memory dysfunction,but the specific molecular mechanism remains largely unknown.NMDAR is an ionic excitatory amino acid receptor that regulates synaptic plasticity and affects the process of learning and memory.BDNF is a brain-derived nerve growth factor that plays an important role in the reproduction,growth,differentiation and survival of neurons.However,the regulation mechanism of NMDAR subunits,BDNF and related molecules under different conditions of microwave radiation in learning and memory impairment caused by microwave radiation is not yet clear.Therefore,this study intends to explore the role of NMDAR,BDNF and related molecules in learning and memory impairment caused by microwave radiation,and provide new targets for its prevention and treatment.Materials and methods:(1)Animal model:two kinds of radiation condition were used:In the single radiation condition,140 male wistar rats were exposed to microwave at 0,10,30 and 50mW/cm~2 for 6 min.In the long-term radiation condition,75 male Wistar rats were exposed to microwave at 0,5,10,20 and 30mW/cm~2 for6min/time,3 times/week,for 6 weeks.Morris water maze was used to test the spatial learning and memory function of rats.The hippocampal structure and synapse ultrastructure were observed by H&E staining and transmission electron microscope.(2)Neuron model:In the single radiation condition,the rat primary hippocampal neurons were exposed to microwave at 0,5,10,20,30 and 50mW/cm~2 for 6 min.In the multiple radiation condition,The rat primary hippocampal neurons were exposed to microwave at 0,5,10,20 and 30mW/cm~2 for 6min/time,3 times,each time interval of6min.The neurons viability was detected by CCK-8 assay at 0h,6h,24h,48h,72h after microwave radiation.The density of dendritic spine in neurons were observed by the laser scanning confocal microscopy(LCSM)using Vybrant CM-Dil cell-labeling at0h,6h,24h,48h,72h after microwave radiation.(3)The expression of several key NMDAR signaling molecules in neurons was evaluated by Western Blot,including NR1,NR2A,NR2B,PSD-95,cortactin,BDNF and TrkB.Results:(1)The changes of learning and memory after microwave exposure:At 6h,1d and 2d,rats in single radiation condition showed significant longer escape latency and less number of platform crossings(P<0.01 or P<0.05).(2)The changes of hippocampal structure:In single radiation group,at 6h and 3d after microwave exposure,the hippocampus structure had no obvious change.At 7d after 10mW/cm~2microwave exposure,neurons appeared slightly concentrated anachromasis.After30mW/cm~2 microwave exposure,the numbers of stained neurons increased.After50mW/cm~2 microwave exposure,there was more concentration of deeply stained neurons.At 14d after microwave exposure the changes trended to recover.At 28d and2m,the neurons of radiation group restored to the normal level.(3)The changes of synapse ultrastructure:In the 10 mW/cm~2 group after microwave exposure of 6h-7d,and in the 30-50mW/cm~2 group after microwave exposure of 6h-14d,microwave caused synaptic cleft blurred,postsynaptic density increased and the postsynaptic active region lengthened(P<0.05).At 28d and 2m,the synaptic structure was basically restored to the normal level.(4)Effects of different conditions microwave radiation on the NMDAR,BDNF and related molecules in hippocampus:In the single radiation condition,compared with the control group,the expressions of the correlative proteins did not statistically change after microwave irradiation of 10mW/cm~2exposure at each time points.With the average power density of 30mW/cm~2,the expressions of NR1,NR2A and NR2B were increased significantly at time points of 1d,3d and 7d after microwave exposure(P<0.05);the expression of PSD-95 was increased significantly at time points of 3d and 7d after microwave exposure(P<0.05);the expression of cortactin was increased significantly at the time of 3d after microwave exposure(P<0.05);at 6h and 1d,the expression of BDNF was increased after radiation(P<0.05);the expression of TrkB was increased after radiation at 1d(P<0.05).After50mW/cm~2 microwave radiation,the expressions of NR1,NR2A,NR2B were decreased at each time points(P<0.05);the expression of PSD-95 was decreased at 7d after microwave radiation(P<0.05);the expression of cortactin was decreased at time point of 3d after microwave exposure(P<0.05);at 6h and1d(P<0.05),the expression of BDNF was decreased after radiation;at 1d(P<0.05),the expression of TrkB was decreased after radiation(P<0.05).In the long-term microwave radiation condition,compared with the control group,the expressions of correlative proteins did not statistically change after microwave irradiation of 5mW/cm~2 exposure at each time point.At 14d,in the group of 10mW/cm~2,the expressions of NR1 and PSD-95 were increased(P<0.05).After 20mW/cm~2 microwave radiation,the expression of NR1 was increased at 14d and 28d(P<0.05);the expression of NR2A was increased at 28d(P<0.05),and the expression of NR2B was decreased at 14d and 28d(P<0.05).With the average power density of 30mW/cm~2,the expressions of NR1?NR2A?PSD-95were increased and the expression of NR2B were decreased at 14d and 28d(P<0.05);the expressions of cortactin,BDNF and TrkB were increased at 14d after irradiation(P<0.05).(5)The effect of different conditions microwave irradiation on the activity of primary hippocampal neurons:In the single radiation condition,there were no significant changes in neuronal activity at each time points of 5mW/cm~2 group.At 0h,in 10 and 20mW/cm~2 groups,the neuronal activity decreased(P<0.05).At 0h and 6h,the activities of neurons in 30 and 50mW/cm~2 groups were decreased(P<0.05),and that of 50mW/cm~2 at 24h after raditation was still lowered(P<0.05).There was no significant difference in the neuronal activity at 48h and 72h after radiation.Under multiple radiation conditions,in the 5mW/cm~2 radiation group,there was no significant difference in the activity of hippocampal neurons.At 0h and 6h,in the groups of 10,20,30 and 50mW/cm~2,the activity of neurons decreased after radiation(P<0.05).In the 30 and 50mW/cm~2 groups,the activity were lower than that of group0mW/cm~2 at 24h after radiation(P<0.05).At 48h after radiation of 50mW/cm~2,the activity rate was still lowered(P<0.05).(6)The effect of different conditions microwave irradiation on dendritic spine density of primary hippocampal neurons:under single radiation condition,there were no significant change in dendritic spine density in 5 and 10m W/cm~2 groups at any time after radiation.At 48h and 72h,in the group of 20 and 30mW/cm~2,the dendritic spine density increased after radiation(P<0.05).In the group of 50mW/cm~2,as time went on,there was a downward trend.And the density of dendritic spines was lower than that of group 0mW/cm~2 at48h and 72h(P<0.05).Under multiple radiation condition,there were no significant difference in the dendritic spine density of hippocampal neurons in the 5mW/cm~2group after radiation.In 10,20,30 and 50mW/cm~2 groups,the dendritic spine density decreased at 0h and 6h(P<0.05).And at 24h,the dendritic spine density of 30 and50mW/cm~2 groups were lower than that in 0mW/cm~2 group(P<0.05).At 48h,the dendritic spine density of 50mW/cm~2 group was still lower than that of 0mW/cm~2group(P<0.05).(7)The effect of different conditions microwave irradiation on NMDAR,BDNF and related molecules in primary hippocampal neurons:In the single radiation condition,compared with the control,the expressions of correlative proteins did not statistically change after microwave irradiation of 5,10mW/cm~2exposure at each time points.With the average power density of 20mW/cm~2,the expression of NR1 was increased significantly at the time of 6h after microwave exposure(P<0.05).With the average power density of 30mW/cm~2,the expression of NR1 and NR2B were increased significantly at time points of 6h and 24h after microwave exposure(P<0.05);the expressions of NR2A,BDNF and TrkB were increased significantly at timepoints of 6h after microwave exposure(P<0.05);the expressions of PSD-95 and cortactin were increased significantly at 24h after microwave exposure(P<0.05).After 50mW/cm~2 microwave radiation,the expressions of NR1 and NR2B were decreased at 6h and 24h(P<0.05);the expressions of NR2A,BDNF and TrkB were decreased at 6h(P<0.05);the expression of PSD-95 was decreased at 24 and 48h(P<0.05);the expression of BDNF was decreased at 6h,1d after radiation(P<0.05);the expression of cortactin was decreased after radiation atn48h(P<0.05).In multiple radiation conditions,The expressions of correlative proteins did not statistically change after microwave irradiation of 5mW/cm~2 exposure at each time point.In the group of 10mW/cm~2,the expression of NR1 were increased at 6h(P<0.05).After 20mW/cm~2 microwave radiation,the expressions of NR1 and NR2B were increased at 6h and 24h(P<0.05),the expression of NR2A was increased at 24h(P<0.05).With the average power density of 30mW/cm~2,the expressions of NR1 and NR2B were increased at 6h and 24h(P<0.05);the expressions of NR2A and cortactin were increased at 24h and 48h(P<0.05);the expression of PSD-95 was increased at48h(P<0.05);the expressions of BDNF and TrkB were increased at 6h(P<0.05).With the average power density of 50mW/cm~2,the expression of NR1 was decreased at 6h,24h and 48h(P<0.05);the expressions of NR2A and TrkB were decreased at 6h and24h(P<0.05);the expressions of NR2B and PSD-95 were decreased at 24h and 48h(P<0.05);the expression of BDNF was decreased at 6h;the expression of cortactin was decreased at 48h(P<0.05).Conclusion:(1)Different conditions of microwave radiation could induce the injury of synaptic plasticity in hippocampus and primary cultured hippocampal neurons.(2)There were the discrepancy of NMDAR and its related molecules,BDNF and receptor expression of hippocampus and primary cultured hippocampal neurons in the different radiation dosages of microwave which might participate in the pathophysiological process of synaptic plasticity damage by the different signal transductions.BDNF might regulate the activity of NMDAR after microwave radiation. |
PDF Full Text Request