Repeated Use of Morphine Induces Anxiety by Affecting a Proinflammatory Cytokine Signaling Pathway in the Prefrontal Cortex in Rats.

We examined the role of toll-like receptors (TLRs) and proinflammatory cytokine signaling pathways in the prefrontal cortex (PFC) in anxiety-like behaviors after repeated use of morphine. Morphine (10 mg/kg) was used twice daily for 8 days to induce morphine dependence in male Wistar rats. On day 8, opioid dependence was confirmed by measuring naloxone-precipitated withdrawal signs. On days 1 and 8, anxiety-like behaviors were evaluated using a light/dark box test. Expression of TLR1 and 4, proinflammatory cytokines, and some of the downstream signaling molecules was also evaluated in the bilateral PFC at mRNA and protein levels following morphine dependence. The results revealed that morphine caused anxiolytic-like effects on day 1 while induced anxiety following 8 days of repeated injection. On day 8, a significant decrease in TLR1 expression was detected in the PFC in morphine-dependent rats, but TLR4 remained unaffected. Repeated morphine injection significantly increased IL1-ß, TNFα, and IL6 expression, but decreased IL1R and TNFR at mRNA and protein levels except for IL6R at the protein level in the PFC. The p38α mitogen-activated protein (MAP) kinase expression significantly increased but the JNK3 expression decreased in the PFC in morphine-dependent rats. Repeated injection of morphine also significantly increased the NF-κB expression in the PFC. Further, significant increases in Let-7c, mir-133b, and mir-365 were detected in the PFC in morphine-dependent rats. We conclude that TLR1 and proinflammatory cytokines signaling pathways in the PFC are associated with the anxiogenic-like effects of morphine following its chronic use in rats via a MAP kinase/NF-κB pathway.

Differential expression of H19, BC1, MIAT1, and MALAT1 long non-coding RNAs within key brain reward regions after repeated morphine treatment.

Morphine-induced analgesic tolerance and dependence are significant limits of pain control; however, the exact molecular mechanisms underlying morphine tolerance and dependence have remained unclear. The role of long non-coding RNAs (lncRNAs) in morphine tolerance and dependence is yet to be determined. We aimed to explore the association of specific lncRNAs expression in key brain reward regions after repeated injection of morphine. Male Wistar rats received subcutaneous injections of twice-daily morphine (10 mg/kg) or saline (1 mL/kg) for eight days. On day 8 of the repeated injections, induction of morphine analgesic tolerance and dependence was confirmed through a hotplate test and a naloxone-precipitated withdrawal analysis, respectively. Expression of H19, BC1, MIAT1, and MALAT1 lncRNAs was determined from the midbrain, striatum, hypothalamus, prefrontal cortex (PFC), and hippocampus by real-time PCR on day 8 of the repeated injections. The H19 expression was significantly different between morphine-treated and control saline-treated rats in all investigated areas except for the hippocampus. The BC1 expression significantly altered in the midbrain, hypothalamus, and hippocampus, but not in the striatum and PFC after repeated morphine treatment. The MIAT1 and MALAT1 expression site-specifically altered in the midbrain, hypothalamus, and striatum; however, no significant changes were detected in their expression in the PFC and hippocampus after repeated morphine treatment. We conclude that alterations in the expression of these lncRNAs in the brain reward regions especially in the midbrain, striatum and hypothalamus may have critical roles in the development of morphine dependence and tolerance, which need to be considered in future researches.