The Effect Of Minocycline On Thermal Hyperalgesia,Anxiety,Depression-like Behaviors And Its Mechanism In CCI Rats

Neuropathic pain is a chronic pain which resulted from peripheral nerve injury.Chronic pain and comorbid depression are frequently encountered clinically.The hippocampus is a key region for learning and memory.Recently,it is noted that the hippocampus is a part of the brain that plays a key role in painful feeling and mood regulation.In addition,the prefrontal cortex plays an important role in the executive functions and behavior-making that is associated with pain and depression.Chronic restraint stress or LPS adiministration can induce long-term depressive-like behaviors and the increased expression levels of NLRP3,IL-1β,IL-18 and Iba-1 in hippocampus.In CCI rats,it is not known whether the expression of NLRP3 inflammasome in the hippocampus and prefrontal cortex can be influenced after intra-hippocampus CA1 injection of minocycline.Therefore,we observed: 1)the effect of intra-hippocampus injection of minocycline on the thermal hyperalgesia and anxiety-or depression-like behaviors in CCI rats;2)the effect of intra-hippocampus injection of minocycline on the expression of Iba-1 in hippocampus and prefrontal cortex of CCI rats;3)the effect of intra-hippocampus injection of minocycline on the expression of NLRP3,Caspase-1,IL-1β and L-18 in hippocampus and prefrontal cortex of CCI rats.Objective: 1.The aim of this study is to explore whether intra-hippocampus injection of minocycline can alleviate pain,anxiety and depression-like behavior by affecting microglia activation and NLRP3 signaling pathway in hippocampus and prefrontal cortex of CCI rats.Methods: We randomly selected SD male rats with 5~6 weeks old.All rats should be implantation of intra-hippocampus catheter first.1.All rats were randomized into three groups(n=8):(1)Sham +PBS group,(2)CCI group(CCI+intra-hippocampus in 0.01 M PBS,0.5μL),(3)Minocycline-treated group(CCI+intra-hippocampus1,2,5,10 and 15μg/μL Minocycline).For the rats after ligation,minocycline or PBS were intra-hippocampus administered twice a day for 7 and 14 consecutively days.Then,the TWL was evaluated before the surgery and 1,3,5,7,10 and 14 days after nerve surgery.2.Several different tests were utilized to investigate the effects of minocycline on anxietyor depressive-like behaviors(SPT,OFT and EPM)in CCI rats.3.The expression of NLRP3,Caspase-1,IL-1β and IL-18 in hippocampus and prefrontal cortex were assessed by using RTFQ-PCR on the 7 and 14 post-operative days.The expression of IL-1β and IL-18 in hippocampus and prefrontal cortex was also examined.The expression of Iba-1 in hippocampus and prefrontal cortex were examined by immunohistochemistry at the 7 post-operative days.The expression of NLRP3 and Iba-1 in hippocampus and prefrontal cortex were examined by western blot on the 7 and 14 post-operative days,respectively.Results: 1.The results of thermal pain threshold show that,compared with sham group,CCI rats displayed significantly decreased TWL on day 1 after nerve injury(P<0.05),and the allodynia was sustained for 14 days.Compared with CCI group,minocycline exerted anti-hyperalgesic effects as early as 1 day after CCI(P<0.05),and the effects lasted for 14 days(P<0.05).2.The results of anxiety and depression-like behaviors test show that,compared with the sham group,the percentage of sucrose preference was evaluated 7,10 and 14 days after surgery in the CCI group decreased significantly(P<0.05).Compared with the CCI group,the percentage of sucrose preference was evaluated 7 and 14 days after surgery in rats with intra-CA1 injection of minocycline group increased significantly(P<0.05).Compared with the sham group,the CCl group spent significantly less time and ran shorter distances in the center area(P<0.05),Compared to the CCI group,intra-CA1 injection of minocycline treatment group as expected increased the spent time and ran longer distances in the center area(P<0.05).Compared with the sham group,the CCI group showed significantly decreased number of open arm entries and time spent in open arms.Compared with CCI rats,intra-CA1 injection of minocycline treatment group inecreased number of open arm entries and time spent in open arms(P<0.05).3.The PCR results indicated that the expression of NLRP3,Caspase-1,IL-1β and IL-18 were markedly enhanced in the hippocampus and prefrontal cortex on 7 and 14 days after nerve injury(P<0.05).Compared with CCI group,intra-hippocampus administration of minocycline significantly reduced CCI-induced the increase in NLRP3,Caspase-1,IL-1β and IL-18 m RNA on days 7 and 14(P<0.05).4.The ELISA results indicated that,compared with sham group,the expression of IL-1β and IL-18 was significantly increased in the hippocampus and prefrontal cortex on 7 and 14 days after nerve injury(P<0.05).Compared with CCI group,intra-hippocampus administration of minocycline suppressed the increased content of IL-1β and IL-18 in the hippocampus and prefrontal cortex at the days 7 and 14(P<0.05).5.The immunoreactivity results indicated that,compared with sham group,the expression of Iba-1 was significantly increased in the hippocampus and prefrontal cortex at the 7 days after nerve injury(P<0.05).Compared with CCI group,intra-hippocampus administration of minocycline suppressed the increased expression of Iba-1 in the hippocampus and prefrontal cortex at the 7 days after nerve injury(P<0.05).6.The WB results showed that compared with sham group,the expression of Iba-1 and NLRP3 was significantly increased in the hippocampus and prefrontal cortex at the 7 and 14 days after nerve injury(P<0.05).Compared with CCI group,intra-hippocampus administration of minocycline suppressed the increased expression of Iba-1 and NLRP3 in the hippocampus and prefrontal cortex at the 7 and 14 days after nerve injury(P<0.05).Conclusions: 1.Intra-hippocampus CA1 injection of minocycline can alleviate pain,anxiety and depression-like behavior in CCI rats.2.Intra-hippocampal CA1 injection of minocycline may inhibition of hippocampus and prefrontal cortex microglia activation and the decreased expression of NLRP3 and Caspase-1 m RNA and results in the decreased production of IL-1β and IL-18.