CCL2/CCR2 In The Nucleus Accumbens Contributes To Pain Hypersensitivity And Depression-like Behaviors

Objective Pain is an unpleasant subjective feeling and emotional experience caused by tissue injury.The present preclinical and clinical data show that chronic pain patients are usually accompanied with depression,anxiety,and anhedonia.However,it is still lack of effective treatments in clinic to treat neuropathic pain and emotional disorders.Thus,developing a new and effective drug for the treatment of pain targets is urgent.Recent evidence showed that dysfunction of nucleus accumnens(NAc)in the mesolimbic reward system plays an important role in chronic pain and other affective disorders.CCL2,a monocyte chemotactic protein,is involved in inflammatory pain and neuropathic pain through its receptor CCR2 in the peripheral tissues and spinal dorsal horn.Nonetheless,the underlying mechanisms are poorly understood.Here we demonstrate a role for chemokine CCL2 and its receptor CCR2 in the regulation of neuropathic pain and depression-like behavior in the NAc shell.Methods Adult ICR mice(8-12 weeks)were used to induce peripheral neuropathic pain via tight ligation of the L5 spinal nerve.The mRNA and protein expression level of CCL2/CCR2 were examined by PCR and Western-Blot,respectively.The expression of pNR2 B,NR2B,pERK and ERK protein was detected by Western Blot.The expression and cellular localization of CCL2/NeuN?CCL2/GFAP?CCL2/IBA-1?CCR2/pNR2B? CCR2/GFAP?CCR2/IBA-1?D1R/CCR2?D2R/CCR2?D1/NeuN?D2/NeuN?D1/pERK and D2/pERK were determined by immunofluorescence staining.The mechanical allodynia,heat hyperalgesia,sucrose preference test(SPT),and forced swimming test(FST)were measured as pain hypersensitivity and depressive behaviors.The effects of local injection of CCR2 shRNA,CCL2 overexpression lentivirus,NR2 B antagonist Ro25-6981,and ERK antagonist PD98059 on the related behaviors were observed.Results 1?The mRNA and protein expression of CCL2 and CCR2 were significantly increased in the NAc after SNL.Immunofluorescence staining showed that both CCL2 and CCR2 were mainly labeled with neuronal marker NeuN.Moreover,they were also expressed in dopamine D1 receptor-and D2 receptor-positive cells.2?Behavioral analysis showed that SNL produced a persistent pain hyperalgesia and depression-like behaviors.The mice showed a significant increase in the immobility time as tested in the forced swimming test(FST),and a significant decrease in sucrose preference(SPT).Intra-NAc shell injection of CCR2 shRNA lentivirus significantly attenuated SNL-induced mechanical allodynia and heat hyperalgesia and improved the depression-like behaviors(SPT/FST).On the contrary,over-expression of CCL2 by lentiviral vectors injection into the NAc shell induced pain hyperalgesia and depression-like behavior in na?ve mice.3?SNL increased the phosphorylation of NR2 B subunits,but the enhancement could be abolished by intra-NAc shell pre-injection of CCR2 shRNA lentivirus.Conversely,over-expression of CCL2 by lentiviral vectors injection into the NAc shell upregulated the phosphorylation of NR2 B.Furthermore,immunofluorescence staining showed that CCR2 was colocalized with pNR2 B.4?NR2B antagonist Ro 25-6981 effectively reduced SNL-induced pain hyperalgesia and depression-like behavior.5?SNL increased the phosphorylation of ERK,p-ERK was colocalized with D1 R and D2 R.Intra-NAc shell pre-injection of CCR2 shRNA lentivirus reduced SNL-induced p ERK upregualtion.Conversely,over-expression of CCL2 by lentiviral vectors injection into the NAc shell upregulated the pERK expression.Furthermore,ERK antagonist PD98059 effectively reduced SNL-induced pain hyperalgesia and depression-like behavior.Conclusions The upregulation of CCL2 and its receptor CCR2 in NAc shell contributes to SNL-induced pain hypersensitivity and depression-like behaviors in mice.CCL2/CCR2 signaling may exert their function via ERK-mediated phosphorylation of NMDA receptor subunit NR2 B in the NAc shell.