| Objective: Through establishing neuropathic pain(NP)rat model of spared nerve injury(SNI),1.To observe the neuralgia and depression response induced by SNI.2.To observe the therapeutic effect of electroacupuncture(EA)on neuralgia and depression induced by SNI.3.To observe the effect on synaptic plasticity in spinal cord and hippocampal neurons after EA.4.To observe the effect on microglia proliferation and activation in spinal dorsal horn and hippocampus after EA.5.To observe the effects on inflammatory mediators CXCL1,IL-1β,IL-6,TNF-α and TGF-β in serum,spinal cord and hippocampus after EA.6.The effects on CaMKII/Kalirin-7/Rac1 signaling pathway,p-NR2 B and PSD-95 in spinal cord and hippocampus by EA were also detected.Methods: 180 SD male rats were randomly divided into 5 groups,each with 36 rats,namely sham operation group(Sham),sham operation + EA group(Sham+EA),model group(SNI),model + sham EA group(SNI+Mock EA),model + EA group(SNI+EA).To establish a spared nerve injury(SNI)rat model,the specific method was as followes: under pentobarbital anesthesia,the rats were placed in the left decubitus position to expose the right sciatic nerve,the sural nerve was retained,the tibial nerve and common peroneal nerve were ligated,and then excised.In the sham group,only the nerves were exposed without injured.The EA group received daily EA treatment(bilateral Zusanli(ST36)and Yanglingquan(GB34)acupoints,2Hz,1-2m A,30 minutes,consecutive 7 days)from the first day after surgery,while the Mock EA group only inserted acupuncture needles without current.Mechanical withdrawal threshold(MWT)and thermal withdrawal latency(TWL)were used as indicators to measure NP on one day before surgery,day 3,day 7 and day 14 after surgery.Depressive behaviors were evaluated using forced swimming test(FST)and sucrose preference test(SPT)on day 7 and day 14 after surgery.After completion of the behaviors measurement on the 14 th day after surgery,the samples were collected and the density of dendritic spines in the spinal cord and hippocampal neurons was detected by Golgi staining.Immunofluorescence and Western blot were conducted to observe the proliferation and activation of microglia in spinal dorsal horn and hippocampal DG area.Serum,hippocampal and spinal cord tissues were collected on days 3,7 and 14 after behavioral tests,respectively.The expression of inflammatory mediators CXCL1,IL-1β,IL-6,TNF-α and TGF-β in serum,spinal cord and hippocampus were analyzed by Enzyme-linked immunosorbent assay(ELISA).Reverse transcription-polymerase chain reaction(RT-PCR)was used to detect the expression of Kalirin-7 m RNA in the spinal cord and hippocampus.The expression of CaMKII/Kalirin-7/Rac1 signaling pathway,p-NR2 B and PSD-95 protein in the spinal cord and hippocampus were detected by Western blot.Results: 1.There was no statistically significant difference in MWT and TWL among all groups before surgery(P>0.05).Compared with Sham group,MWT and TWL in SNI group were decreased on days 3,7 and 14 after surgery(P<0.05).MWT and TWL in SNI+EA group were increased on days 3,7 and14 after EA(P<0.05),while there was no significant change in the SNI+Mock EA group after sham EA(P>0.05).2.On days 7 and 14 after surgery,the immobility time of forced swimming test in SNI group were increased comparing with Sham group(P<0.05),while the climbing time and sucrose preference value were decreased(P<0.05).Compared with SNI group,the immobility time in SNI+EA group on days 7 and 14 after surgery were reduced(P<0.05),while the climbing time and sucrose preference value were increased(P<0.05),and there was no significant change in SNI+ MOCK EA group(P>0.05).3.Compared with Sham group,the dendritic spine density in spinal dorsal horn of rats in SNI group was significantly increased(P<0.05),while significantly reduced in hippocampus(P<0.05).After EA,the density of dendritic spines of spinal dorsal horn neurons in SNI+EA group was significantly decreased(P<0.05),while significantly increased in hippocampus(P<0.05),and there was no significant change in SNI+ MOCK EA group(P>0.05).4.Compared with Sham group,the expression of microglia marker Iba-1 in the spinal cord and hippocampus DG area of the SNI group was increased(P<0.05).After EA,the expression of Iba-1 in the spinal cord and hippocampus of SNI+EA group was decreased(P<0.05),while there was no significant change in SNI+ MOCK EA group(P>0.05).5.Compared with Sham group,the levels of pro-inflammatory mediators CXCL1,IL-1β,IL-6 and TNF-α in serum,spinal cord and hippocampus of rats in SNI group were all increased on days 3,7 and 14 after operation(P<0.05),while the levels of anti-inflammatory cytokine TGF-β were reduced(P<0.05).After EA treatment,the levels of CXCL1,IL-1β,IL-6 and TNF-α of serum and hippocampus in SNI+ EA group were decreased on days 3,7 and 14 after surgery(P<0.05),while the level of TGF-β was significantly up-regulated(P<0.05).In spinal cord tissue,the levels of CXCL1,IL-1β and TNF-α were reduced on days 3,7 and14 after surgery(P<0.05),and the level of IL-6 was reduced on days 7 and 14(P<0.05),while the level of TGF-β was increased on days 7 and 14(P<0.05),and SNI+ MOCK EA group had no significant change after sham EA treatment(P<0.05).6.Compared with Sham group,the expression of Kalirin-7 m RNA was increased in the spinal cord on days 3,7 and 14 after SNI operation(P<0.05),while decreased in the hippocampus(P<0.05).After EA treatment,the expression of Kalirin-7 m RNA was decreased in the spinal cord and increased in the hippocampus on days 3,7 and 14 in SNI+EA group(P<0.05),while there was no significant change in SNI+ MOCK EA group(P>0.05).7.Compared with Sham group,the protein expressions of p-CaMKII,Kalirin-7,Rac1,p-NR2 B and PSD-95 in the spinal cord were all increased on days 3,7and 14 after operation in SNI group(P<0.05),while those in the hippocampus were all decreased(P<0.05).After EA treatment,the expressions of p-CaMKII,Kalirin-7,p-NR2 B were reduced on days 3,7 and 14,while Rac1 and PSD-95 reduced on days 7 and 14 in the spinal cord,and hippocampal p-CaMKII,Kalirin-7,Rac1,p-NR2 B and PSD-95 expressions were increased on days 3,7and 14 in SNI+EA group(P<0.05),while there was no significant change in SNI+ MOCK EA group(P>0.05).Conclusion: 1.This study confirmed that neuropathic pain and depression-like behaviors could be induced by SNI,and these effects could be improved by EA treatment.2.Electroacupuncture could regulate synaptic plasticity by regulating the density of dendritic spines in spinal cord and hippocampal neurons.3.Electroacupuncture could inhibit the proliferation and activation of microglia cells and reduce neuroinflammation in spinal cord and hippocampus.4.Electroacupuncture may through regulating CaMKII/Kalirin-7/Rac1 signaling pathway improve synaptic plasticity and inhibit neuroinflammation,thus playing anti-neuropathic pain and anti-depressive role. |
