ABSTRACT
OBJECTIVE To investigate the role of the complement C3/C3aR signaling pathway in the prefrontal cortex and colon neuroglia cell interactions during meth-amphetamine(METH)addiction,to observe the effects of TLR4 inhibitors as well as complement C3 elimination on METH reward and relapse behavior,and to explore the neuroinflammatory mechanisms of complement C3 acti-vation in METH addiction.METHODS ①A 14 d and 28 d rat METH addiction model was established to observe the effects of TLR4 antagonist ibudilast 3 mg·kg-1 and 10 mg·kg-1 on self-administration,reward motivation,relapse,and natural reward behavior in METH-trained 14 d rats and the effects of 0.02 mg·kg-1 complement C3 antago-nist on self-administration behavior in METH-trained 28 d rats.② Differences in the expression of TLR4,NF-κB,GRP94,C3,cathepsin L,CD68,and GFAP in the pre-frontal cortex of each group were examined using West-ern blotting.③ In addition,the expression of ATF6 in the prefrontal cortex of each group and the effects on neuro-nal and microglia/macrophage INOS,CD206 GRP94,and complement C3/C3aR.RESULTS ① Endoplasmic reticulum stress occurred in neurons and microglia after METH exposure depending on GRP94 and unfolded pro-tein responses to the ATF6 pathway.In addition,it acti-vates the TLR4-NF-κB pathway.② Microglia with high complement C3/C3aR expression in the prefrontal cortex were recruited to synaptic pruning and phagocytic responses around neurons with high GRP94,comple-ment C3/C3aR expression and these effects were blocked by complement C3 antagonists.③ In the rec-tum,GRP94 functions as a molecular chaperone for com-plement C3 and cathepsin L.Crosstalk occurs between enteric neurons high in GRP94,complement C3,and macrophages high in C3aR,located in the submucosa,lamina propria,and muscular,respectively,and all of these effects are blocked by complement C3 antago-nists.④ Treatment with the TLR4 antagonist ibudilast inhibits self-administration,reward motivation,and cue-or METH-priming in METH-trained 14 d rats,but fails to affect natural reward behavior.Ibudilast treatment attenu-ates the TLR4-NF-κB inflammatory pathway and comple-ments C3/C3aR pathway in the prefrontal cortex.CON-CLUSION Activation of the complement C3/C3aR signal-ing pathway by TLR4-NF-κB inflammatory signaling in the prefrontal cortex mediates the METH addiction pro-cess,providing an experimental basis for the clinical treatment of METH addiction,and targeting TLR4/NF-κB inflammatory signaling and complement C3/C3aR may be a new way to intervene in METH addiction.
ABSTRACT
OBJECTIVE To investigate the effect of injection of β2-adrenergic receptor agonist clenbuterol into the infralimbic cortex(IL) on drug-seeking behavior triggered by conditioned cues. METHODS Adult male SD rats were trained to self-administer heroin under a FR1 schedule for consecutive 14 d,followed by 2-h extinction training. Cue-induced heroin seeking was measured for 2 h. Clenbuterol was microinjected bilaterally into the IL(8 ng/side)of rats 15 min prior to reinstatement test. Meanwhile,locomotor activity was detected 15 min after clenbuterol or artifial cerebrospinal fluid(mod?el group) was microinjected bilaterally into IL. Western blotting was used to detect the expression of phosphorylated cyclic AMP response element-binding protein(p-CREB)in the prelimbic cortex(PL), IL,nucleus accumbens core (NACc) and shell (NACsh) of rats immediately after reinstatement test. RESULTS After heroin administration training for 14 consecutive days,these animals exhibited reliable heroin self-administration,indicated by the increase in active nose poke responses and infusions. The rats that had received infusion of clenbuterol into the IL had significantly lower active pokes (8 ± 3)than those in model group(45±10)in cue-induced reinstatement(P<0.01),but there was no significant differ?ence between clenbuterol group and vehicle group in the locomotor activity. The expression of p-CREB in either IL or NACsh was significantly decreased in clenbuterol group compared with model group(P<0.01,P<0.05),but significantly increased in NACc(P<0.01). CONCLUSION Microinjection of clenb?uterol into the IL can attenuate the cue-induced reinstatement of heroin-seeking behavior in rats. The underlying mechanism might be related to the regulation of p-CREB expression in the NACc and NACsh.