Switching from morphine to fentanyl attenuates the decline of µ-opioid receptor expression in periaqueductal gray of rats with morphine tolerance.

BACKGROUND: Opioid switching is a therapeutic maneuver to improve analgesic response and/or reduce adverse side effects although the underlying mechanisms remain unknown. The µ-opioid receptor (MOR) has an important role in mediating the actions of morphine and other analgesic agents. This study is aimed at exploring the changes of MOR in the periaqueductal gray (PAG) in rats when morphine is substituted for equianalgesic fentanyl. METHODS: Forty rats were randomly assigned to five treatment groups: 7 days normal saline group (N group), 7 days fentanyl group (F group), 7 days morphine group (M group), 7 days morphine and 7 days fentanyl-switching group (MF group), and 14 days morphine group (MM group). Rats repeatedly received subcutaneous injections of morphine sulfate (10 mg/kg) or equianalgesic fentanyl sulfate (0.1 mg/kg) twice daily. Rats' antinociceptive response to thermal pain was evaluated by the tail flick latency assay. MOR mRNA and protein expression in the PAG were measured using RT-PCR and Western blotting analyses respectively. RESULTS: This study showed that after morphine was substituted with fentanyl on day 8, the tail flick latency (TFL) increased from (3.9 ± 0.4) seconds to (11.4 ± 0.4) seconds. The results also demonstrated that both MOR mRNA and protein expression in the PAG of rats in the MF group were less than that in the M group (P < 0.05) but more than that in MM group (P < 0.05). CONCLUSIONS: Equianalgesic fentanyl was still antinociceptive effective in rats with morphine tolerance, which may be due to the switching from morphine to fentanyl attenuating the decline of MOR expression in the PAG of rats.

The kappa-opioid receptor is upregulated in the spinal cord and locus ceruleus but downregulated in the dorsal root ganglia of morphine tolerant rats.

As a non-selective agonist of opioid receptors, morphine can also act on the kappa-opioid receptor (KOR) when activating the mu-opioid receptor (MOR) and delta-opioid receptor (DOR). Although previous findings indicate that KOR plays an important role in morphine analgesia and antinociceptive tolerance, the reasons for the paradoxical functions of KOR in analgesia and anti-analgesia responses are still unclear. The aim of this study was to explore the role of the KOR in morphine analgesia and antinociceptive tolerance. As such, the changes in KOR expression in different regions of the nervous system in morphine tolerant rats were examined. We were able to attain morphine tolerance in rats via subcutaneous injection of morphine (10 mg/kg) twice daily for 7-consecutive days. Competitive real-time PCR, immunohistochemistry, and Western blot analyses were used to assess KOR expression in related regions of the nervous system, including the thalamus, hypothalamus, hippocampus, locus ceruleus (LC), periaqueductal gray (PAG), lumber-sacral spinal cord, and dorsal root ganglia (DRG). The expression of KOR increased in the locus ceruleus and spinal cord, but was significantly decreased in the DRG of morphine tolerant rats (P<0.05). No other significant changes in KOR expression were observed in the other regions. Consequently, we propose that the locus ceruleus and spinal cord are likely the dominant CNS regions and the DRG is the main peripheral site in which chronic morphine exerts its effect on KOR. Prolonged morphine administration induces inconsistent changes of KOR in the central and peripheral nervous system.

[Effect of fentanyl on expression of mu-receptor and beta-arrestin 2 in periaqueductal gray of rats tolerant to morphine].

OBJECTIVE: To investigate the effect of fentanyl upon the expression of mu-receptor and beta-arrestin 2 in peri-aqueductal gray of morphine-tolerant rats. METHODS: Forty male SD rats weighing (230 +/- 20) g were randomly divided into 5 groups of eight animals each: group NS, group M, group MF1, group MF2 and group MF3. Rats in group NS received only subcutaneous normal saline 1 ml/kg twice a day for 9 consecutive days; group M received subcutaneous morphine 10 mg/kg followed by NS 1 ml/kg twice a day for 9 consecutive days; In groups MF1, MF2 and MF3, morphine 10 mg x kg(-1) was injected subcutaneously followed by fentanyl 3, 6, 12 microg/kg respectively. All animals were sacrificed at Day 9 after measurement of pain threshold. Periaqueductal gray was removed for determination of the expression of mRNA (RT-PCR) and protein (Western-blot) of mu-receptor and beta-arrestin 2. RESULTS: Compared with group NS, TFL of group M was significantly elevated after the first morphine injection (P < 0.01). But TFL of group M returned to the baseline value after chronic morphine treatment. Compared with group M, TFL increased in groups MF2 and MF3 at Days 7 and 9 (P < 0.05 or 0.01). However, TFL of group MF1 was negative (P > 0.05). The expression of mu-receptor mRNA and protein was significantly lower in group M than in group NS (P < 0.01). Compared with group M, the expressions of mu-receptor mRNA and protein were significantly elevated in group MF2 and MF3 (P < 0.05 or 0.01) but there was no significant change in group MF1 (P > 0.05). The expression of beta-arrestin 2 mRNA and protein significantly decreased in group M as compared with group NS (P < 0.01). Compared with group M, the expressions of beta-arrestin 2 mRNA and protein were significantly elevated in group MF2 and MF3 (P < 0.05 or 0.01), but there was no significant change in group MF1 (P > 0.05). CONCLUSION: Fentanyl at 6 and 12 microg/kg can partly inhibit morphine tolerance through an increased expression of mu-receptor and beta-arrestin 2 in periaqueductal gray of morphine-tolerant rats.

Cyclooxygenase-2 promotes angiogenesis by increasing vascular endothelial growth factor and predicts prognosis in gallbladder carcinoma.

AIM: To investigate the relationships between the expression of cyclooxygenase-2 (COX-2), vascular endothelial growth factor (VEGF) and the degree of vascularization, clinicopathologic feature, survival time of patients with gallbladder carcinomas. METHODS: Sixty-four gallbladder carcinoma specimens were evaluated for COX-2, VEGF expression by immunohistochemical methods. Microvessel counts (MVC) were determined using CD(34). The relationships between COX-2, VEGF expression, CD(34)-stained MVC, clinicopathologic features and survival time were analyzed. The correlations between COX-2 and VEGF expression, CD(34)-stained MVC were also investigated. RESULTS: COX-2, VEGF immunoreactivity were observed in 71.9% (46/64) and 54.7% (35/64) specimens, respectively. The average MVC in 64 cases of gallbladder carcinoma was 57+/-14 per high power vision field. The status of MVC was closely correlated with Nevin staging, tumor differentiation and lymph node metastasis (P<0.01, 0.002, and 0.003, 0.000, respectively). Increased VEGF expression was significantly correlated with tumor differentiation (poorly and moderately>well differentiated, P<0.05, P = 0.016). Clinical stages had no relation with the expression of VEGF (P>0.05, P = 0.612). There was a positive correlation between COX-2 expression and clinical stages. The positive rate of COX-2 was higher in cases of Nevin stages S(4)-S(5) (81.8%) than in those of Nevin stages S(1)-S(3) (50.0%) with a statistical significance (P<0.01, P = 0.009). The expression of COX-2 did not vary with differentiation (P>0.05, P = 0.067). Statistically significant differences were also observed according to lymph node metastasis, COX-2 expression and VEGF expression (P<0.01, 0.000, and 0.001, respectively). There was no relation between VEGF, COX-2 expression, MVC and the age and sex of patients. MVC and VEGF positive rate in the COX-2 positive gallbladder carcinoma tissue was higher than that in the COX-2 negative tissue (P<0.05, 0.000, and 0.032, respectively). Patients with VEGF, COX-2 positive tumors had a significantly shorter survival time than those with negative tumors (P<0.05, 0.004, 0.01, respectively). CONCLUSION: Augmented tumor neovascularization induced by VEGF may be one of the several effects of COX-2 responsible for poor prognosis of human gallbladder carcinoma. COX-2 inhibitor, either in combination therapy with other agents, or for chemoprevention, may be effective via suppression of angiogenesis in this fatal disease.