Study On The Mechanism Of Acupuncture On P2X7R Of TRN Microglia For Attention Deficit Of Insomnia Rats

Objective: To explore the mechanism of acupuncture on P2X7 R of TRN microglia for attention deficit of insomnia rats.Method: After SD(Sprague-Dowley)rats were acclimated for 1 week according to the normal light and dark cycle(L: D = 12: 12).Thirty rats synchronized with light and dark cycle were chose.According to the amount of spontaneous activity in the light period,they were randomly divided into a control group(10 rats)and a model replication group(20 rats).The insomnia model were established by intraperitoneal injection of para-chlorophenylalanine(PCPA)once a day for 2 days.Then the model replication group were randomly divided into a model group and an acupuncture group,with 10 rats in each group.Rats in the acupuncture groups received acupuncture at Neiguan(PC 6)and Zusanli(ST 36)for continuous 5d at ZT4,once a day every time lasted 15 minutes.The control group and the model group were synchronously bundled and fixed for 15 minutes at ZT4.The Clocklab 2 data acquisition system was used to dynamically monitor the 24-hour spontaneous circadian rhythm of rats.The event-related potential(ERP)P3 and CNV were used to evaluate the attention of rats.After acupuncture,the whole brain structure was harvested.Immunofluorescence was used to detect the microglia fluorescence expression in TRN region.The concentrations of interleukin-1?(IL-1?)and tumor necrosis factor-?(TNF-?)were detected by ELISA.Results:1.During L: D(12:12)light and dark domestication,the circadian rhythm of the rats' sleep activity was stable,which was manifested as rest in light period and activity in dark period.And there was no significant difference in the amount of spontaneous activity during the light period among the groups in this period(P > 0.05).After PCPA modeling,the spontaneous circadian rhythm of the model group and the acupuncture group disappeared,and the spontaneous activity during the light period increased significantly(P<0.01).After treatment,the sleep-wake cycle gradually recovered,and the amount of spontaneous activity in the light period decreasedsignificantly compared with the model group(P<0.01).2.At the baseline,there were no significant differences in the latency and amplitude of P3 among the three groups before the model establishment(P>0.05).After modeling,the P3 latency was significantly longer and the amplitude of P3 was decreased.Compared with the control group,the differences were all statistically significant(P<0.01).After acupuncture treatment,the latency and amplitude of P3 presented significant changes(P<0.05)in the latency and amplitude of P3 compared with the model group.3.Immunofluorescence results showed that the average optical density value of P2X7 R on microglia in the model group was significantly higher than that in the control group(P<0.01).After acupuncture treatment,the average optical density of P2X7 R on microglia in TRN area of acupuncture group was significantly lower than that of model group(P<0.01).The average optical density of P2X7 R on microglia in the acupuncture group was not significantly different from that in the blank group(P>0.05).The microglia cell body in TRN area of the control group was smaller,light density was low,and it was the microglia in the resting state.The microglia cell body in the model group was significantly increased with branch becoming shorter and thicker.And the number of activated microglia was significantly increased.Compared with the control group,its average light density value was significantly increased(P<0.01).The activated morphology of microglia in the acupuncture group was significantly improved compared to that in the model group.The number of activated cells decreased with the reduction of cell body in the acupuncture group.Compared with the model group,the average optical density of microglia in the acupuncture group decreased significantly(P< 0.05).4.Compared with the control group,the concentrations of IL-1? and TNF-? in TRN area of the model group increased significantly(both P < 0.05).Compared with the model group,the concentrations of IL-1? and TNF-? in TRN of the acupuncture group were significantly reduced(both P < 0.05).Conclusions:1.Acupuncture Neiguan(PC 6)and Zusanli(ST 36)could improve attention deficitafter insomnia.2.Acupuncture may reduce the activation of microglia by regulating the expression of P2X7 R in TRN area,and decrease the damage of neurons induced by inflammatory response,so as to achieve the treatment of attention deficit after insomnia.