Introduction: Emerging evidence indicates a critical role for non-coding regulatory RNAs in coordinating many aspects of cellular function that are directly involved in experience-dependent neural plasticity. Non-coding RNAs such as miRNAs alter the expression level of mRNAs while circular RNAs that are another group of non-coding RNAs with covalently closed ends, can sponge miRNAs. An emerging role for the involvement of non-coding RNAs in morphine tolerance and dependence has been proposed. However, very little is known about their potential involvement in epigenetic regulation of morphine tolerance and addiction. The striatum is a part of the reward pathway and it can be affected by morphine. The aim of this study was to investigate the change in Cav2.2 subtype of voltage-gated calcium channel, circular RNA circ-Cacna1b, and some relevant miRNAs including miR-383-3p, miR-133b-3p, miR-182 in the striatum after induction of morphine tolerance and withdrawal in rats. Method: Thirty-two male Wistar rats (n = 8 in each group) were used. Morphine tolerance was induced after 10 days of injections of morphine (10 mg/kg, s.c.) twice a day. A control group received saline (1 ml/kg) twice daily for 10 consecutive days instead of morphine. Two other groups of animals underwent the same treatments of saline and morphine as the two previous groups but were maintained for additional 30 days of withdrawal. For gene expression study, each rat was sacrificed, its whole brain was removed, and the striatum was dissected either on day 10 of the repeated injections or on day 30 of the withdrawal. The gene expression was examined using a quantitative RT-PCR method. The real time-PCR data was analyzed using the Livak method (2-ΔΔCT) and an independent t-test was used for pairwise comparisons. P<0.05 was set as a statistically significant level. Result: The RT-PCR results indicated no significant changes in the Cav2.2 expression, miR-383-3p, and miR-182 but revealed a significant increase in circ-Cacna1b and a significant decrease in miR-133b-3p expression in the striatum of morphine-tolerant rats compared with the saline-treated control group. The gene expression results in the striatum after withdrawal also indicated significant decreases in the expression of Cacna1b, miR-133b-3p, miR-182, and miR-383-3p but a significant increase in the circ-Cacna1b expression in morphine withdrawal group compared to the respective saline-treated animals. Conclusion: It can be concluded that changes in Cav2.2 voltage-dependent calcium channel and its biologically relevant circular RNA circ-Cacna1b and miRNAs in the striatum may underlie, at least partly, neuroadaptations involved in morphine-induced analgesic tolerance and withdrawal.
