Attenuated Morphine Tolerance Research Articles

Ferroptosis inhibitor ferrostatin-1 attenuates morphine tolerance development in male rats by inhibiting dorsal root ganglion neuronal ferroptosis.

Ferrostatin-1 and liproxstatin-1, both ferroptosis inhibitors, protect cells. Liproxstatin-1 decreases morphine tolerance. Yet, ferrostatin-1's effect on morphine tolerance remains unexplored. This study aimed to evaluate the influence of ferrostatin-1 on the advancement of morphine tolerance and understand the underlying mechanisms in male rats. This experiment involved 36 adult male Wistar albino rats with an average weight ranging from 220 to 260 g. These rats were categorized into six groups: Control, single dose ferrostatin-1, single dose morphine, single dose ferrostatin-1 + morphine, morphine tolerance (twice daily for five days), and ferrostatin-1 + morphine tolerance (twice daily for five days). The antinociceptive action was evaluated using both the hot plate and tail-flick tests. After completing the analgesic tests, tissue samples were gathered from the dorsal root ganglia (DRG) for subsequent analysis. The levels of glutathione, glutathione peroxidase 4 (GPX4), and nuclear factor erythroid 2-related factor 2 (Nrf2), along with the measurements of total oxidant status (TOS) and total antioxidant status (TAS), were assessed in the tissues of the DRG. After tolerance development, the administration of ferrostatin-1 resulted in a significant decrease in morphine tolerance (P < 0.001). Additionally, ferrostatin-1 treatment led to elevated levels of glutathione, GPX4, Nrf2, and TOS (P < 0.001), while simultaneously causing a decrease in TAS levels (P < 0.001). The study found that ferrostatin-1 can reduce morphine tolerance by suppressing ferroptosis and reducing oxidative stress in DRG neurons, suggesting it as a potential therapy for preventing morphine tolerance.

Blocking Palmitoylation of Apelin Receptor Alleviates Morphine Tolerance in Neuropathic Cancer Pain.

Neuropathic cancer pain (NCP) is an important symptom in patients with cancer. However, significant analgesic tolerance and other side effects critically hamper the administration of morphine. Protein palmitoylation mediated by the DHHC family may be involved in the glial activation and inflammatory responses underlying organ failure. In this study, we investigated the key role of protein palmitoylation in cancer pain and sought to target palmitoylation to suppress morphine tolerance. We found that long-term use of morphine led to the accumulation of the morphine metabolite, morphine-3-glucuronide, in vivo and activated ERK1/2 and microglia to release inflammatory factors through the apelin receptor APLNR. Palmitoyltransferase ZDHHC9 was upregulated in NCP, and APLNR was palmitylated to protect it from lysosomal degradation and to maintain its stability. We also designed competitive inhibitors of APLNR palmitoylation to inhibit the development of NCP, release of inflammatory factors, and attenuation of morphine tolerance. Therefore, targeting APLNR palmitoylation in combination with morphine is a potent method for cancer pain treatment. Our data provide a basis for the future clinical use of related drugs combined with morphine for the treatment of cancer-related pain.

Ondansetron and AS19 attenuate morphine tolerance by modulating serotonin 5-HT3 and 5-HT7 receptor expressions in rat dorsal root ganglia

PurposeOpioid drugs are frequently preferred drugs for severe pain, but their long-term use causes tolerance to their analgesic effects. However, the factors that contribute to the development of tolerance to opioids are diverse and not fully understood. In this study, our aim was to investigate the effects of 5-HT3 receptor antagonist ondansetron (ODN) and 5-HT7 receptor agonist AS19 on morphine analgesia and tolerance in rats. Material and methodsEighty-four randomly selected Wistar Albino (230–260 g) male rats were included in the study. To induce morphine tolerance in rats, morphine (10 mg/kg) was administered subcutaneously twice daily for 7 days, and tolerance was assessed by thermal analgesia tests on day 8 for 120 min. After thermal analgesia tests, dorsal root ganglia (DRGs) at spinal L4-S5 level were isolated. 5-HT3 and 5-HT7 receptor expressions in DRGs were measured by immunohistochemical method. ResultsThe results show that co-administration of ODN or AS19 with morphine indicated a significant increase in analgesic effect compared to morphine alone In addition, the difference between the mean of ODN or AS19 combined with morphine and the morphine-tolerant group was statistically significant. H-score analysis significantly increased 5-HT3 receptor expressions in the morphine-tolerant rats, and administration of ODN prevented this. Similarly, 5-HT7 receptor expressions were significantly decreased in the morphine tolerant group, and the administration of AS19 increased this expression. ConclusionThese study results suggest that both ODN and AS19 alter the 5-HT3 and 5-HT7 receptor expressions in the DRG and possibly attenuate morphine tolerance in this way.

Monoclonal antibody targeting mu-opioid receptor attenuates morphine tolerance via enhancing morphine-induced receptor endocytosis

Morphine is a frequently used analgesic that activates the mu-opioid receptor (MOR), which has prominent side effects of tolerance. Although the inefficiency of morphine in inducing the endocytosis of MOR underlies the development of morphine tolerance, currently, there is no effective therapy to treat morphine tolerance. In the current study, we aimed to develop a monoclonal antibody (mAb) precisely targeting MOR and to determine its therapeutic efficacy on morphine tolerance and the underlying molecular mechanisms. We successfully prepared a mAb targeting MOR, named 3A5C7, by hybridoma technique using a strategy of deoxyribonucleic acid immunization combined with cell immunization, and identified it as an immunoglobulin G mAb with high specificity and affinity for MOR and binding ability to antigens with spatial conformation. Treatment of two cell lines, HEK293T and SH-SY5Y, with 3A5C7 enhanced morphine-induced MOR endocytosis via a G protein-coupled receptor kinase 2 (GRK2)/β-arrestin2-dependent mechanism, as demonstrated by immunofluorescence staining, flow cytometry, Western blotting, coimmunoprecipitation, and small interfering ribonucleic acid (siRNA)-based knockdown. This mAb also allowed MOR recycling from cytoplasm to plasma membrane and attenuated morphine-induced phosphorylation of MOR. We established an in vitro morphine tolerance model using differentiated SH-SY5Y cells induced by retinoic acid. Western blot, enzyme-linked immunosorbent assays, and siRNA-based knockdown revealed that 3A5C7 mAb diminished hyperactivation of adenylate cyclase, the in vitro biomarker of morphine tolerance, via the GRK2/β-arrestin2 pathway. Furthermore, in vivo hotplate test demonstrated that chronic intrathecal administration of 3A5C7 significantly alleviated morphine tolerance in mice, and withdrawal jumping test revealed that both chronic and acute 3A5C7 intrathecal administration attenuated morphine dependence. Finally, intrathecal electroporation of silencing short hairpin RNA illustrated that the in vivo anti-tolerance and anti-dependence efficacy of 3A5C7 was mediated by enhanced morphine-induced MOR endocytosis via GRK2/β-arrestin2 pathway. Collectively, our study provided a therapeutic mAb, 3A5C7, targeting MOR to treat morphine tolerance, mediated by enhancing morphine-induced MOR endocytosis. The mAb 3A5C7 demonstrates promising translational value to treat clinical morphine tolerance.

Intra-incisional injection of magnesium sulfate for post cesarean pain management

Background and objectives: cesarean section is estimated for about 30% of child births worldwide and it is one of the most common surgical procedures. Pain relief after cesarean delivery is especially important as the consequences of inadequate pain relief are expressed not only by the mother but by the newborn as well. Magnesium has been reported to produce important analgesic effects including the potentiation of morphine analgesia, attenuation of morphine tolerance and the suppression of neuropathic pain. The objectives of this study were to determine the efficacy of intra-incisional injection of magnesium sulfate for post cesarean pain management as in patient underwent elective cesarean section and the reduced need for extra analgesic use postoperatively. Methods: a randomized, placebo controlled, single-blinded study with a total of 200 pregnant participants scheduled for cesarean section in Sulaimani maternity teaching hospital, from 1st of May 2016 to 15th of August 2016. Patients were randomly allocated to two groups, those with odd numbers were assigned to case group (100) received 750mg of magnesium sulfate and patients with even numbers assigned to the placebo group (100) received 20mg of normal saline. Results: The mean Visual Analogue Score after 4 hours, 8 hours, 12 hours and 24hours was significantly less for the intervention (Magnesium sulfate) group when compared with the control (Normal Saline). Conclusions: Subcutaneous administration of magnesium sulfate in post cesarean section pain management can be used as a successful modality or method for pain management.

Blockade of orexin receptor type-1 by SB-334867 and activation of orexin receptor type-2 attenuate morphine tolerance in rats.

The interaction of orexinergic neurons with the opioidergic system and their effects on morphine analgesia and tolerance have not been fully elucidated. The purpose of the study was to evaluate the effects of the orexin-1 and orexin-2 receptor (OX1R and OX2R) agonist and antagonist on morphine analgesia and tolerance in rats. A total of 90 Wistar albino male rats weighing 180-220 g wereused in the experiments. To induce morphine tolerance, rats were injected with a single dose of morphine (50 mg kg-1, s.c.) for 3 days. Morphine tolerance was assessed on day 4 in randomly selected rats by analgesia tests. In order to evaluate morphine tolerance situation, orexin-A, SB-334867, orexin-B andTCSOX2 29 were administered together with morphine for 3 days. The analgesic effects of orexin-A (10 μg kg-1), OXR1 antagonist SB-334867 (10 mg kg-1), OXR2 agonist orexin-B (15 μg kg-1), OXR2 antagonist TCS OX2 29 (0.5 mg kg-1) and morphine (5 mg kg-1) were measured at 15 or 30-min intervals by tail-flick and hot-plate antinociceptive tests. The results suggested that the combination of orexin-1 receptor antagonist SB-334867 and orexin-B with morphine significantly increased the analgesic effect compared to morphine-tolerant rats. In addition, administration of orexin-A and -B alone showed significant analgesic effects compared to the saline group. However, co-administration of orexin-A and -B with morphine didnot increase the analgesic efficacy of morphine. The results of this study demonstrated that co-administration of SB-334867 and orexin-B with morphine attenuated morphine tolerance. Further studies are needed to elucidate the details of the interaction between orexin receptors and the opioidergic system.

Metformin prevents morphine-induced apoptosis in rats with diabetic neuropathy: a possible mechanism for attenuating morphine tolerance.

Morphine is a drug of choice for the treatment of severe and chronic pain, but tolerance to the antinociceptive effect limits its use. The development of tolerance to morphine has recently been associated with neuronal apoptosis. In this study, our aim was to investigate the effects of metformin on morphine-induced neuronal apoptosis and antinociceptive tolerance in diabetic rats. Three days of cumulative dosing were administered to establish morphine tolerance in rats. The antinociceptive effects of metformin (50mg/kg) and test dose of morphine (5mg/kg) were considered at 30-min intervals by thermal antinociceptive tests. To induce diabetic neuropathy, streptozotocin (STZ, 65mg/kg) was injected intraperitoneally. ELISA kits were used to measure caspase-3, bax, and bcl-2 levels from dorsal root ganglion (DRG) tissue. Semi-quantitative scoring system was used to evaluate apoptotic cells with the the terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) method. The findings suggest that co-administration of metformin with morphine to diabetic rats showed a significant increase in antinociceptive effect compared to morphine alone. The antinociceptive tests indicated that metformin significantly attenuated morphine antinociceptive tolerance in diabetic rats. In addition, metformin decreased the levels of apoptotic proteins caspase 3 and Bax in DRG neurons, while significantly increased the levels of antiapoptotic Bcl-2. Semi-quantitative scoring showed that metformin provided a significant reduction in apoptotic cell counts in diabetic rats. These data revealed that metformin demonstrated antiapoptotic activity in diabetic rat DRG neurons and attenuated morphine tolerance. The antiapoptotic activity of metformin probably plays a significant role in reducing morphine tolerance.

Reduced activity of adenylyl cyclase 1 attenuates morphine induced hyperalgesia and inflammatory pain in mice.

Opioid tolerance, opioid-induced hyperalgesia during repeated opioid administration, and chronic pain are associated with upregulation of adenylyl cyclase activity. The objective of this study was to test the hypothesis that a reduction in adenylyl cyclase 1 (AC1) activity or expression would attenuate morphine tolerance and hypersensitivity, and inflammatory pain using murine models. To investigate opioid tolerance and opioid-induced hyperalgesia, mice were subjected to twice daily treatments of saline or morphine using either a static (15 mg/kg, 5 days) or an escalating tolerance paradigm (10–40 mg/kg, 4 days). Systemic treatment with an AC1 inhibitor, ST03437 (2.5–10 mg/kg, IP), reduced morphine-induced hyperalgesia in mice. Lumbar intrathecal administration of a viral vector incorporating a short-hairpin RNA targeting Adcy1 reduced morphine-induced hypersensitivity compared to control mice. In contrast, acute morphine antinociception, along with thermal paw withdrawal latencies, motor performance, exploration in an open field test, and burrowing behaviors were not affected by intrathecal Adcy1 knockdown. Knockdown of Adcy1 by intrathecal injection also decreased inflammatory mechanical hyperalgesia and increased burrowing and nesting activity after intraplantar administration of Complete Freund’s Adjuvant (CFA) one-week post-injection.

GPR171 activation regulates morphine tolerance but not withdrawal in a test-dependent manner in mice.

A newly deorphanized G protein-coupled receptor, GPR171, is found to be highly expressed within the periaqueductal gray, a pain-modulating region in the brain. Our recent research has shown that a GPR171 agonist increases morphine antinociception in male mice and opioid signaling in vitro. The objective of this study was to evaluate the effects of combination treatment in females as well as whether chronic treatment can be used without exacerbating morphine-induced tolerance and withdrawal in female and male mice. Our results demonstrate that activation of GPR171 with an agonist attenuates morphine tolerance in both female and male mice on the tail-flick test, but not the hotplate test. Importantly, the GPR171 agonist in combination with morphine does not exacerbate morphine-induced tolerance and withdrawal during long-term morphine treatment. Taken together, these data suggest that the GPR171 agonist may be combined with morphine to maintain antinociception while reducing the dose of morphine and therefore reducing side effects and abuse liability. The outcome of this study is clearly an important step toward understanding the functional interactions between opioid receptors and GPR171 and developing safer therapeutics for long-term pain management.

Cannabinoid Receptor Type 2 Agonist Reduces Morphine Tolerance via Mitogen Activated Protein Kinase Phosphatase Induction and Mitogen Activated Protein Kinase Dephosphorylation

Morphine is an opioid drug often used in treating moderate to severe pain. However, morphine tolerance in patients limits its used in clinical settings. Our previous study showed that a cannabinoid type 2 (CB2) receptor agonist attenuated morphine tolerance. However, the exact mechanism by which CB2 agonists reduce morphine tolerance remains unclear. In this study, we investigated the effect of mitogen activated protein kinase (MAPK) and mitogen activated protein kinase phosphatases 1 and 3 (MKP-1 and MKP-3) on the regulation of morphine tolerance by CB2 receptor agonist. Chronic morphine treatments for 7 days reduced the protein expression of MKP-1 and MKP-3 in the spinal cord and increased the phosphorylation of p38, ERK1/2 and the level of proinflammatory mediator, such as IL-1β, IL-6 and TNF-α. Coadministration of CB2 receptor agonist AM1241 alleviated the inhibition of MKP-1 and MKP-3 by chronic morphine administration and reduced the expression of phosphorylated MAPK and proinflammatory factors. The effect of the CB2 receptor agonist on morphine-induced downregulation of MKP-1 and MKP-3 was reversed by the MKP-1 and MKP-3 antagonist triptolide. Our findings suggested that CB2 receptor agonist may induce the expression of MKP-1 and MKP-3 to promote MAPK dephosphorylation and reduce the production of downstream cytokine, thereby reducing morphine tolerance. This finding suggested that MKPs may serve as a new target for alleviating morphine tolerance.

Effects of dezocine on morphine tolerance and opioid receptor expression in a rat model of bone cancer pain

BackgroundClinically, the coadministration of opioids to enhance antinociception and decrease tolerance has attracted increasing research attention. We investigated the effects of dezocine, a mu- and kappa-opioid receptor agonist/antagonist, on morphine tolerance and explored the involvement of opioid receptor expression in a rat model of bone cancer pain.MethodsThermal nociceptive thresholds were measured after the subcutaneous injection of morphine (10 mg/kg) alone or combined with dezocine (10 or 1 mg/kg) for 7 consecutive days. Real-time PCR and western blot analysis were used to examine opioid receptor expression in the periaqueductal gray (PAG) and spinal cord.ResultsThe analgesic effect was significantly decreased after 4 days of morphine administration. We observed that low-dose dezocine significantly attenuated morphine tolerance without reducing the analgesic effect of morphine. Low-dose dezocine coadministration significantly reversed the downregulated expression of mu (MOR) and delta (DOR) opioid receptors in the PAG and the upregulated expression of kappa (KOR) and DOR in the spinal cord induced by morphine. Moreover, low-dose dezocine coadministered with morphine significantly inhibited KOR expression in both the PAG and spinal cord.ConclusionsThe combination of low-dose dezocine with morphine may prevent or delay the development of morphine tolerance in a rat model of bone cancer pain. The regulation of opioid receptor expression in the PAG and spinal cord may be part of the mechanism.

A20-Binding Inhibitor of Nuclear Factor-κB Targets β-Arrestin2 to Attenuate Opioid Tolerance.

Opioids play an important role in pain relief, but repeated exposure results in tolerance and dependence. To make opioids more effective and useful, research in the field has focused on reducing the tolerance and dependence for chronic pain relief. Here, we showed the effect of A20-binding inhibitor of nuclear factor-κB (ABIN-1) in modulating morphine function. We used hot-plate tests and conditioned place preference (CPP) tests to show that overexpression of ABIN-1 in the mouse brain attenuated morphine dependence. These effects of ABIN-1 are most likely mediated through the formation of ABIN-1-β-arrestin2 complexes, which accelerate β-arrestin2 degradation by ubiquitination. With the degradation of β-arrestin2, ABIN-1 overexpression also decreased μ opioid receptor (MOR) phosphorylation and internalization after opioid treatment, affecting the β-arrestin2-dependent signaling pathway to regulate morphine tolerance. Importantly, the effect of ABIN-1 on morphine tolerance was abolished in β-arrestin2-knockout mice. Taken together, these results suggest that the interaction between ABIN-1 and β-arrestin2 inhibits MOR internalization to attenuate morphine tolerance, revealing a novel mechanism for MOR regulation. Hence, ABIN-1 may be a therapeutic target to regulate MOR internalization, thus providing a foundation for a novel treatment strategy for alleviating morphine tolerance and dependence. SIGNIFICANCE STATEMENT: A20-binding inhibitor of nuclear factor-κB (ABIN-1) overexpression in the mouse brain attenuated morphine tolerance and dependence. The likely mechanism for this finding is that ABIN-1-β-arrestin2 complex formation facilitated β-arrestin2 degradation by ubiquitination. ABIN-1 targeted β-arrestin2 to regulate morphine tolerance. Therefore, the enhancement of ABIN-1 is an important strategy to prevent morphine tolerance and dependence.

Intrathecal Knockdown of Adenylyl Cyclase 1 Attenuates Morphine Tolerance and Withdrawal in Mice

Opioid tolerance and opioid induced hyperalgesia during repeated opioid administration and chronic pain are associated with upregulation of adenylyl cyclase activity. Therefore, we investigated if expression of components of the cyclic-AMP pathway were increased within in the nervous system tissues of morphine tolerant mice, and if inhibiting any of these changes could alleviate pain and/or tolerance in mouse models. The objective of this study was to test the hypothesis that a reduction in adenylyl cyclase 1 (AC1) activity would attenuate morphine tolerance and hypersensitivity, and inflammatory pain using murine models. Short-hairpin RNA (shRNA) gene knockdown of Adcy1 in the spinal cord and dorsal root ganglia was accomplished using a lumbar injection of an associated adenovirus viral vector (AAV9-GFP-U6-m-Adcy1-shRNA) and negative controls (AAV9-GFP-U6-m-scrambl-shRNA). Behavioral testing such as open field testing, rotarod testing, burrowing, thermal and mechanical paw withdrawal latencies were tested after injection. Morphine tolerance (15mg/kg, sc, 5 days and 10-40mg/kg escalation over 4 days) and opioid-induced hypersenstivity were also assessed after inoculation. Lumbar intrathecal administration of a vector incorporating adeno-associated virus and short-hairpin RNA against Adcy1 did not affect baseline parameters such as open field testing, rotarod testing, and burrowing testing. Morphine tolerance and withdrawal were attenuated in Adcy1 shRNA mice compared to control vector mice. Chronic exposure to morphine leads to changes in mRNA expression of proteins involved in the cAMP signaling pathway in different areas of the nervous system. Gene knockdown of Adcy1 decreases morphine tolerance and opioid-induced hypersensitivity, which could form the basis for novel therapeutics in the future. This work was supported through K01 DA042902 to AHK and the Purdue University College of Pharmacy to VJW.

Hydrogen enriched saline alleviates morphine tolerance via inhibiting neuroinflammation, GLT-1, GS nitration and NMDA receptor trafficking and functioning in the spinal cord of rats

The development and maintenance of morphine tolerance showed association with neuroinflammation and dysfunction of central glutamatergic system (such as nitration of glutamate transporter). Recent evidence indicated that hydrogen could reduce the levels of neuroinflammation and oxidative stress, but its role in morphine tolerance has not been studied. The rats were intrathecally administered with morphine (10 μg/10 μL each time, twice/day for 5 days). Hydrogen enriched saline (HS) or saline was given intraperitoneally at 1, 3 and 10 mL/kg for 10 min before each dose of morphine administration. The tail-flick latency, mechanical threshold and thermal latency were assessed one day (baseline) before and daily for up to 5 days during morphine injection. The pro-inflammatory cytokine expressions [tumor necrosis factor-alpha (TNF-α), interleukin-1β (IL-1β), IL-6)] (by western blotting), astrocyte activation (by immunofluorescence and western blotting), and nitration of glutamate transporter-1 (GLT-1) and glutamine synthetase (GS) (by immunoprecipitation), membrane and total expression of N-methyl-d-aspartic acid (NMDA) receptor NR1 and NR2B subunits were carried out in the spinal dorsal horns. Chronic morphine administration induced antinociceptive tolerance, and together led to increased TNF-α, IL-1β and IL-6 expression, astrocyte activation, GLT-1 and GS nitration, increased membrane and total NR1, NR2B expression. Injection of HS attenuated morphine tolerance in a dose-dependent manner, decreased proinflammatory cytokine expression, inhibited astrocyte activation, decreased GLT-1 and GS nitration, and inhibited membrane trafficking of NMDA receptor. Our result showed that hydrogen pretreatment prevented morphine tolerance by reducing neuroinflammation, GLT-1, GS nitration, NMDA receptor trafficking in the spinal dorsal horn. Pretreatment with hydrogen might be considered as a novel therapeutic strategy for the prevention of morphine tolerance.

15727 KATP channel prodrugs as therapeutics for chronic pain and substance abuse disorders

ABSTRACT IMPACT: Pharmacological activation of KATP channels may provide analgesia and attenuate opioid tolerance and withdrawal OBJECTIVES/GOALS: Our long term goal is to develop therapeutics for the treatment of the overuse of opioids. The objective of this application is to test novel KATP channel-targeting prodrugs in rodent models of neuropathic and inflammatory pain in addition to opioid tolerance after chronic morphine administration. METHODS/STUDY POPULATION: In one study, two different measures for chronic pain were implemented in mice. Male and female mice (n=10) were subjected to spinal nerve ligation (SNL) or intraplantar injection of Complete Freund’s Adjuvant (CFA) to induce neuropathic and inflammatory pain, respectively. Administration of KATP channel prodrugs (60ug, it) attenuated mechanical hypersensitivity after SNL or CFA compared to vehicle (saline). In a separate study, changes in mechanical hypersensitivity were tested while mice undergo chronic morphine treatment (15mg/kg, 2x, 5 days) with administration of the prodrugs. Tolerance was measured as the loss of antinociception, and withdrawal is measured ˜24 hours after the final morphine injection. RESULTS/ANTICIPATED RESULTS: Intrathecal administration of either KATP channel prodrugs significantly attenuated mechanical hypersensitivity after SNL and significantly attenuated mechanical hypersensitivity after CFA in mice. We predict that intrathecal administration of these prodrugs will also attenuate morphine tolerance and withdrawal in mice. This hypothesis is based off our previous data indicating non-water soluble KATP channel agonists produce analgesia and attenuate morphine tolerance in mice. DISCUSSION/SIGNIFICANCE OF FINDINGS: Pharmaceutical strategies to utilize KATP channels for therapeutics have been hindered due to the low solubility and low ability to cross the neurovascular unit. Newly developed, water-soluble KATP channel openers could be useful pharmaceutical strategy to reduce chronic pain, opioid tolerance, and withdrawal in human populations.

Donepezil attenuates the development of morphine tolerance in rats with cancer-induced bone pain: The role of cortical N-methyl-D-aspartate receptors

Cancer-induced bone pain (CIBP), which is associated with poor quality of life, is most commonly treated using opioids. However, long-term use of morphine for analgesia induces tolerance and can diminish the treatment’s effectiveness. The mechanisms that underlie morphine tolerance have been reported to be related to the inflammation of the nervous system and hyperactivation of N-methyl-D-aspartate receptors (NMDARs). Donepezil is an anti-inflammatory and neuroprotective drug that is thought to alleviate morphine tolerance. In this study, we aimed to investigate the effect of three different dosages of donepezil (1, 1.5 and 2 mg/kg) on morphine tolerance in rats with CIBP, and the possible involvement of donepezil-mediated NMDAR subunit 1 (NR1). We found that donepezil can prolong the analgesic efficacy of morphine and delay the development of chronic morphine tolerance. Furthermore, continuous morphine injection increased the expression of NR1, and this was suppressed by co-administration with donepezil using both western blotting and immunofluorescence. Our findings demonstrate that donepezil has the potential to attenuate morphine tolerance, possibly by inhibiting NR1 activity in the cortex.

Electroacupuncture Attenuates Morphine Tolerance in Rats with Bone Cancer Pain by Inhibiting PI3K/Akt/JNK1/2 Signaling Pathway in the Spinal Dorsal Horn.

Purpose:Morphine is often used for the treatment of moderate and severe cancer pain, but long-term use can lead to morphine tolerance. Methods for effectively inhibiting morphine tolerance and the related mechanism of action are of great significance for the treatment of cancer pain. Previous studies have shown that electroacupuncture (EA) can inhibit the occurrence of morphine tolerance, but the mechanism is not yet clear. The aim of the present study was to explore the signaling pathway by which EA attenuates the development of bone cancer pain (BCP)-morphine tolerance (MT).Materials and methods:Changes in the paw withdrawal threshold (PWT) of rats with bone cancer pain-morphine tolerance were observed in a study of EA combined with intrathecal injection of a PI3K inhibitor (LY294002) or agonist (insulin-like growth factor-1 [IGF-1]). We also tested the protein expression of phosphorylated phosphatidylinositol 3-kinase (p-PI3K), phosphorylated protein kinase B (p-Akt), phosphorylated c-Jun NH2-terminal kinase 1/2 (p-JNK1/2), and β-arrestin2 in the L4-6 spinal dorsal horn of rats.Results:The protein expression of p-PI3K, p-Akt, p-JNK1/2, and β-arrestin2 was upregulated in the L4-6 spinal dorsal horn of rats with bone cancer pain and bone cancer pain-morphine tolerance. EA delayed the occurrence of morphine tolerance in rats with bone cancer pain and downregulated the protein expression of p-PI3K, p-Akt, p-JNK1/2, and β-arrestin2 in the L4-6 spinal dorsal horn of rats with bone cancer pain-morphine tolerance. Intrathecal injection of LY294002 attenuated the development of morphine tolerance and downregulated the protein expression of p-Akt, p-JNK1/2, and β-arrestin2 in the spinal dorsal horn of rats with bone cancer pain-morphine tolerance. In addition, the inhibitory effect of EA on morphine tolerance was reversed by IGF-1.Conclusion:The mechanism underlying the ability of EA to attenuate morphine tolerance may be associated with inhibition of the PI3K/Akt/JNK1/2 signaling pathway.

Upregulation of μ-Opioid Receptor in the Rat Spinal Cord Contributes to the α2-Adrenoceptor Agonist Dexmedetomidine-Induced Attenuation of Chronic Morphine Tolerance in Cancer Pain.

BackgroundSustained morphine treatment for cancer pain has been limited due to analgesic tolerance. Opioid receptor internalization and desensitization mediated by downregulation of mu-opioid receptor (MOR) expression have been confirmed as one of the mechanisms of chronic morphine tolerance. In addition to the opiate system, the α2-adrenergic system is involved in the development of morphine tolerance. Several studies reported that co-administration of α2-adrenoceptor agonist dexmedetomidine inhibits morphine tolerance in normal or neuropathic pain animals. However, the effect of dexmedetomidine on morphine tolerance has not been studied in cancer pain. Therefore, we investigated the effect of intrathecal injection of dexmedetomidine on the development of morphine tolerance in cancer pain and on the expression of MOR in the spinal cord of morphine-tolerant cancer pain rats.MethodsThe model was established using a rat’s right hind paw injection of Walker 256 cancer cells. Subcutaneous morphine (10mg/kg) was administrated twice daily for 7 days; meanwhile, the rats received intrathecal α2-adrenoceptor agonist dexmedetomidine (10μ/kg) or antagonist MK-467 (0.25mg/kg) in test groups. Rats receiving drug vehicle served as the control group. Antinociception was detected by von Frey filaments and hot-plate tests. The expression of MOR in the spinal cord was examined through real-time reverse transcription polymerase chain reaction and Western blotting. The data were analyzed via analysis of variance followed by Student t-test with Bonferroni correction.ResultsSeven-day chronic morphine administration elicited notable analgesic tolerance in the rats with cancer pain. Co-administration of α2-adrenoceptor agonist dexmedetomidine enhanced morphine analgesia and attenuated morphine tolerance, which could be blocked by α2-adrenoceptor antagonist MK-467. Furthermore, pre-treatment of dexmedetomidine significantly upregulated MOR protein expression without a notable change in MOR mRNA expression in the spinal cord.ConclusionOur findings suggest that intrathecal injection of dexmedetomidine enhanced morphine analgesia and attenuated morphine tolerance in cancer pain, potentially by upregulating MOR expression in the spinal cord. The α2-adrenoceptor agonist may provide a more versatile analgesia option for morphine treatment for cancer pain.

Ghrelin receptor agonist hexarelin attenuates antinociceptive tolerance to morphine in rats.

Ghrelin, a peptide hormone released from the gastric endocrine glands, shows analgesic activity apart from its various physiological effects. Nevertheless, the effects of ghrelin receptor (GHS-R) agonists on morphine analgesia and tolerance have not yet been elucidated. The purpose of this study was to evaluate the effects of the ghrelin receptor agonist hexarelin and antagonist [d-Lys3]-GHRP-6 on morphine antinociception and tolerance in rats. A total of 104 Wistar albino male adult rats (weighing approximately 220-240g) were used in the experiments. To induce morphine tolerance, a three-day cumulative dose regimen was used in the rats. Then, randomly selected rats were evaluated for morphine tolerance on day4. The analgesic effects of hexarelin (0.2mg·kg-1), [d-Lys3]-GHRP-6 (10mg·kg-1), and morphine (5mg·kg-1) were measured at 30-min intervals (0, 30, 60, 90, and 120min) by tail-flick and hot-plate analgesia tests. The findings suggest that hexarelin in combination with morphine attenuates analgesic tolerance to morphine. On the other hand, ghrelin receptor antagonist [d-Lys3]-GHRP-6 has no significant analgesic activity on the morphine tolerance in analgesia tests. Furthermore, co-administration of hexarelin and morphine increases the analgesic effect. In conclusion, these data indicate that administration of GHS-R agonist hexarelin with morphine enhances the antinociception and attenuates morphine tolerance.

The Circadian Hormone Melatonin Inhibits Morphine-Induced Tolerance and Inflammation via the Activation of Antioxidative Enzymes.

Opioids are commonly prescribed for clinical pain management; however, dose-escalation, tolerance, dependence, and addiction limit their usability for long-term chronic pain. The associated poor sleep pattern alters the circadian neurobiology, and further compromises the pain management. Here, we aim to determine the correlation between constant light exposure and morphine tolerance and explore the potential of melatonin as an adjuvant of morphine for neuropathic pain treatment. Methods: Wistar rats were preconditioned under constant light (LL) or a regular light/dark (LD) cycle before neuropathic pain induction by chronic constriction injury. An intrathecal (i.t.) osmotic pump was used for continued drug delivery to induce morphine tolerance. Pain assessments, including the plantar test, static weight-bearing symmetry, and tail-flick latency, were used to determine the impact of the light disruption or exogenous melatonin on the morphine tolerance progression. Results: constant light exposure significantly aggravates morphine tolerance in neuropathic rats. Continued infusion of low-dose melatonin (3 μg/h) attenuated morphine tolerance in both neuropathic and naïve rats. This protective effect was independent of melatonin receptors, as shown by the neutral effect of melatonin receptors inhibitors. The transcriptional profiling demonstrated a significant enhancement of proinflammatory and pain-related receptor genes in morphine-tolerant rats. In contrast, this transcriptional pattern was abolished by melatonin coinfusion along with the upregulation of the Kcnip3 gene. Moreover, melatonin increased the antioxidative enzymes SOD2, HO-1, and GPx1 in the spinal cord of morphine-tolerant rats. Conclusion: Dysregulated circadian light exposure significantly compromises the efficacy of morphine’s antinociceptive effect, while the cotreatment with melatonin attenuates morphine tolerance/hyperalgesia development. Our results suggest the potential of melatonin as an adjuvant of morphine in clinical pain management, particularly in patients who need long-term opioid treatment.