The Role Of Cross-regulation Of BDNF And NMDAR In Abnormal Synaptic Transmission Of Hippocampal Neurons Induced By Microwave Radiation

Objective: A certain dose of microwave radiation can cause learning and memory damage.Synaptic plasticity is the neurobiological basis of learning and memory.Abnormal synaptic transmission is an important manifestation of microwave radiation-induced learning and memory damage.BDNF and NMDAR are signal molecules transmitted by synapses.They regulate each other and play an important role in the generation of LTP and the formation of learning and memory.Therefore,this study uses the mutual regulation of BDNF and NMDAR as an entry point to explore the mechanism of abnormal synaptic transmission in hippocampal neurons caused by microwave radiation,and provides a basis for the protection of learning and memory damage caused by microwave radiation.Methods: The primary hippocampal neurons of rats were irradiated by microwaves with average power density of 30 m W/cm2 and 50 m W/cm2.Cells were radiated 3 times,6 min/time,with interval of 6mins.NMDAR subunits(NR1,NR2 A,and NR2B),BDNF,and Trk B m RNA were detected by Realtime-PCR at 1h,6h,12 h and 24 h after irradiation.NMDAR and the related post-synaptic molecules(PSD-95 and Cortactin),BDNF and Trk B proteins,the synaptic vesicle-related molecules(Synaptobrevin,Synaptophysin,Synapsin I)were detected by Western Blot at 6h,24 h and 48 h after irradiation.The signal kinases ERK,PKC and PI3 K immediately after irradiation were detected by spectrophotometry.Then BDNF or TrkB inhibitor(K252a),NMDA or NMDAR inhibitor D-AP5 and PKC inhibitor Go6976 were used to explore the mechanism.Results:(1)The effect of BDNF on NMDAR in hippocampal neurons after microwave radiation and its role in abnormal post-synaptic information transmission.(1)The changes of NMDAR m RNA: At 1h after irradiation,the expression of NR1,NR2 A and NR2 B m RNA in the 30 m W/cm2 and 50 m W/cm2 groups decreased;At 6-12 h after irradiation,the expression of NR1,NR2 A and NR2 B m RNA increased in the 30 m W/cm2 group and decreased in the 50 m W/cm2 group(P<0.05 or P<0.01).(2)Effects of BDNF or K252 a on NMDAR m RNA: After 1-6h of microwave exposure in the BDNF intervention group,the expression of NR1,NR2 A,and NR2 B m RNA increased in the 30 m W/cm2 and 50 m W/cm2 groups;the expression decreased in the K252 a intervention group(P<0.05 or P<0.01).(3)Effect of BDNF or K252 a on the expression of NMDAR protein and post-synaptic related molecules: BDNF intervention group,after 6-24 h of microwave exposure,the expression of NR1,NR2 A and NR2 B increased in the 30 m W/cm2 and 50 m W/cm2 groups,After 24-48 h,the expression of Cortactin and PSD-95 increased;In the K252 a intervention group,their expressions decreased(P<0.05 or P<0.01).(4)Effect of BDNF or K252 a on kinase activity: Immediately after irradiation,the PKC kinase activity increased in the 30 m W/cm2 group and decreased in the 50 m W/cm2 group(P<0.05 or P<0.01).In the BDNF intervention group,PKC kinase activity increased;In the K252 a intervention group,PKC kinase activity decreased(P<0.05 or P<0.01).(5)The effect of Go6976 on NMDAR and post-synaptic related molecules: In the Go6976 intervention group,after 6h of microwave exposure,the expression of NR1,NR2 A and NR2 B protein in the 30 m W/cm2 and 50 m W/cm2 groups decreased;After 24 h of microwave exposure,PSD-95 and Cortactin protein expression decreased;after 48 h of microwave exposure,PSD-95 protein expression still decreased(P<0.05).(2)Effect of NMDAR on BDNF in hippocampal neurons after microwave irradiation and its role in the abnormal release of amino acid transmitters:(1)The changes of BDNF and Trk B m RNA: after 1 h of 30 m W/cm2 irradiation,BDNF m RNA decreased,and after 6-12 h of irradiation,BDNF and Trk B m RNA increased;after 50 m W/cm2 irradiation of 1-12 h,BDNF and Trk B m RNA expression decreased(P<0.05 or P<0.01).(2)Effect of NMDA or D-AP5 on BDNF m RNA: NMDA intervention group,BDNF m RNA increased after microwave exposure for 1-6h;D-AP5 intervention group,BDNF m RNA decreased(P<0.05 or P<0.01).(3)Effect of NMDA or DAP5 on BDNF and Trk B protein expression: In the NMDA intervention group,after 6h of microwave exposure,the expression of BDNF protein in 30 m W/cm2 and 50 m W/cm2 groups increased;In the D-AP5 intervention group,the expression of BDNF protein decreased(P<0.05 or P <0.01).(4)Effect of NMDA or D-AP5 on PKC kinase activity: NMDA intervention group,the expression of PKC kinase activity increased in the 30 m W/cm2 and 50 m W/cm2 groups immediately after microwave exposure;PKC kinase activity expression decreased in the D-AP5 intervention group(P<0.05).(5)Effect of NMDA or D-AP5 on presynaptic vesicle-related proteins: At 6h after irradiation,the expression of Synaptophysin in the 30 m W/cm2 group decreased,and the expression of Synaptobrevin,Synaptophysin,and Synapsin I in the 50 m W/cm2 group decreased;In the NMDA intervention group,the expression of Synaptobrevin increased in the 50 m W/cm2 group(P<0.05 or P<0.01).(6)The effect of NMDA or DAP5 on the release of amino acid transmitters: The release of GABA reduced at 6h and 24 h after 30 m W/cm2 irradiation and the release of Glu and GABA was promoted with NMDA intervention.The release of Glu reduced and that of Gly increased after 50 m W/cm2 irradiation.After NMDA administration the release of Glu was promoted and the release of Gly was inhibitd with NMDA intervention,but the release of GABA and Gly was inhibitd with D-AP5 intervention(P<0.05 or P <0.01).Conclusions:(1)Different conditions of microwave radiation can cause abnormal expression of presynaptic NMDAR,BDNF and related signaling molecules in primary hippocampal neurons of rats,and there are dose differences.(2)NMDAR and BDNF play different roles in the synaptic transmission abnormality of rat hippocampal neurons caused by microwave radiation under different conditions through mutual regulation: after 30 m W/cm2 microwave radiation,the expression of NMDAR in hippocampal neurons is up-regulated,which promotes the transcription and expression of BDNF,not only causes GABA release disorder and activate the PKC signaling pathway,further up-regulate the expression of NMDAR subunits and related molecules,promote the recovery of synaptic transmission damage,and thus play a protective role in learning and memory damage.After 50 m W/cm2 microwave irradiation,the expression of NMDAR in hippocampal neurons was down-regulated,which inhibited the transcription and expression of BDNF;in turn,down-regulated the expression of vesicle-associated protein Synaptobrevin and reduced the release of Glu.At the same time,by inhibiting the PKC signaling pathway,it further down-regulated the NMDAR subunit and related molecules Expression leads to abnormal synaptic transmission and exacerbates learning and memory impairment.