Backgrounds: Molecular mechanisms involved in adverse effects of morphine, including tolerance and dependence, have remained elusive. We examined possible alterations in the gene expression of proenkephalin (Penk), prodynorphin (Pdyn), and mu-opioid receptor (Oprm1) in reward brain areas following frequent morphine treatment. Methods: Two groups of male Wistar rats were used. The groups received either saline (1 mL/kg) or morphine (10 mg/kg) twice daily for eight days. On day 8, rats were decapitated, brain areas involved in addiction were dissected, including the midbrain, striatum, prefrontal cortex (PFC), hippocampus, and hypothalamus, and gene expression was evaluated with real-time PCR. Results: Prolonged morphine treatment decreased Penk, Pdyn, and Oprm1 gene expressions in the midbrain but upregulated them in the striatum compared to the control group treated with saline. Significant increases in Pdyn and Oprm1 gene expressions were detected in the PFC, but there was no significant difference in Penk gene expression between the two groups. Besides, Pdyn gene expression was decreased in the hippocampus and hypothalamus; however, no significant differences in Penk and Oprm1 gene expressions were detected between the groups in these areas. Conclusions: The expression of endogenous opioid peptides and receptors after frequent use of morphine follows a region specificity in brain areas involved in addiction. These alterations may result in new physiological setpoints outside the normal range, which need to be considered when using morphine in medicine.
We aim to examine anxiety-like behaviors and expression of specific genes complicated in neuroinflammation in the prefrontal cortex (PFC) after repeated use of morphine. A group of male Wistar rats received injections of morphine (10 mg/kg) twice a day for eight days while a control group received saline (1 ml/kg) instead of morphine. On days 1 and 8, anxiety-like behaviors were evaluated using a light/dark box test. On day 8, opioid dependence was confirmed by measuring the behavioral expression of morphine withdrawal precipitated with naloxone. Expression of neuroinflammation genes were also evaluated at mRNA levels in the PFC on day 8. The results revealed that morphine induced anxiolytic-like effects on day 1, which significantly decreased after the repeated injection of the drug on day 8. The results also revealed that repeated morphine injection significantly increased the mRNA level of Il1, Tnfα, and Il6 but decreased Il1r and Tnfr while increased Il6r in the PFC. The gene expression results also revealed a significant decrease in Tlr1 but not in Tlr4 in the PFC of morphine-dependent rats. Although Erk1 expression had no significant alteration but p38 increased and Jnk3 decreased significantly in the PFC in morphine-dependent rats. Creb and Nfkb significantly increased but Fos expression decreased. Let-7c, mir-133b, and mir-365 also significantly increased in the PFC in morphine-dependent rats. We conclude that the alteration in neuroinflammatory pathways at gene expression level in the PFC may party underlie neuroadaptive changes leading to the decrease in anxiolytic effect of morphine in dependent rats.
We examined the involvement 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 dependence in male Wistar rats was induced via twice-daily morphine injection (10 mg/kg) for eight days. On day 8, opioid dependence was confirmed by measuring morphine withdrawal signs precipitated with naloxone. On days 1 and 8, anxiety-like behaviors were evaluated using a light/dark box test. On day 8, TLR1 and 4, proinflammatory cytokines, and some of the downstream signaling molecules were evaluated at mRNA and protein levels in the PFC. The results revealed that morphine caused anxiolytic-like effects on day 1, which significantly decreased after eight days of the repeated injection. On day 8, a significant decrease in TLR1 expression in the PFC was detected in morphine-dependent rats but TLR4 remained unaffected. Repeated morphine injection significantly increased theIL1-β, 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 of morphine-dependent rats. The NF-κB expression also increased significantly in the PFC after repeated injection of morphine. Significant increases in Let-7c, mir-133b, and mir-365 were also detected in the PFC in morphine-dependent rats. We conclude that TLR1 and proinflammatory cytokines in the PFC via a MAP kinase/NF-κB pathway may party underlie the decrease in the anxiolytic-like effect of morphine in dependent rats.
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