Introduction: Opioid tolerance and the potential for addiction is a significant problem associated with pain management. The precise underlying mechanism of morphine tolerance and prevention remain elusive. Different brain regions including midbrain, striatum and cortical areas are affected by morphine. Molecular studies have shown that during morphine tolerance, the mu-opioid receptors, glutamate receptors and some downstream signaling molecules are affected, which may partly underlie the induction of morphine tolerance. According to recent reports, immune signaling also contributes to the decreased efficacy of opioids. It has been reported that toll-like receptor 4 (TLR4)-mediated neuroinflammation in the midbrain drives tolerance, which is mediated via inflammatory cytokines like tumor necrosis factor (TNF) and interlukine-1 (IL-1). The aim of this study was to examine changes in mRNA level of TNF receptor, IL-1 receptor, TLR1 and 4, as well as downstream mitogen activated protein kinases (MAPKs) p38 and JNK3 in the striatum after induction of morphine tolerance in rats. Method: Male Wistar rats were used in which morphine tolerance was induced with eight days injections of morphine 10 mg/kg (i.p.) twice per day. A control group also received saline (1 ml/kg) twice daily for 8 days. On day 8, morphine-induced analgesic tolerance was assessed using a hotplate test of analgesia. For gene expression study, each rat was sacrificed, the whole brain was removed, and the striatum was dissected in both groups on day 8 of the schedule. The gene expressions of TNFR, IL-1R, TLR1, TLR4, p38 and JNK3 were examined using a quantitative RT-PCR method. The hotplate data was analyzed with a two-way repeated measure ANOVA. The real time-PCR data was analyzed using the 2-ΔΔCT method and an independent t-test was used for the pairwise comparisons. Statistical significant level was set at P<0.05. Result: The results showed that the morphine treatments induced analgesic tolerance on day 8 of the treatments (P<0.001). The results of the qRT-PCR indicated significant increases in TLR1 and TLR4 mRNA levels (P<0.05) but significant decreases in the gene expression of TNFR (P<0.01), IL-1R (P<0.01), p38 and JNK3 MAPKs (P<0.01) in the striatum of morphine-tolerant rats compared to the saline control group. Conclusion: It can be concluded that repeated morphine treatments induce inflammation in the brain, which may alter the gene expression of TLR1, TLR4, TNFR, IL1-R, and p38 and JNK3 MAPKs in the striatum. We propose that inflammation and downstream signaling molecules may underlie, at least partly, morphine tolerance mechanisms, and it must be considered in the future efforts for controlling morphine tolerance and addiction.
