Development Of Tolerance To Morphine-induced Antinociception Research Articles

Microglial TLR4-induced TAK1 phosphorylation and NLRP3 activation mediates neuroinflammation and contributes to chronic morphine-induced antinociceptive tolerance

Background and purposeThe aim of this work was to investigate the role and signal transduction of toll-like receptor 4 (TLR4), TGF-β-activated kinase 1 (TAK1) and nod-like receptor protein 3 (NLRP3) in microglial in the development of morphine-induced antinociceptive tolerance. MethodsTLR4 and NLRP3 knockout mice and 5Z-7-oxozeaeno (a selective inhibitor against TAK1 activity) were used to observe their effect on the development of morphine tolerance. Intrathecal injections of morphine (0.75 mg/kg once daily for 7 days) were used to establish anti-nociceptive tolerance, which was measured by the tail-flick test. Spinal TLR4, TAK1, and NLRP3 expression levels and phosphorylation of TAK1 were evaluated by Western blotting and immunofluorescence. ResultsRepeated treatment with morphine increased total expression of spinal TLR4, TAK1, and NLRP3 and phosphorylation of TAK1 in wild-type mice. TLR4 knockout attenuated morphine-induced tolerance and inhibited the chronic morphine-induced increase in NLRP3 and phosphorylation of TAK1. Compared with controls, mice that received 5Z-7-oxozeaenol showed decreased development of morphine tolerance and inhibition on repeated morphine-induced increase of NLRP3 but not TLR4. NLRP3 knockout mice showed resistance to morphine-induced analgesic tolerance with no effect on chronic morphine-induced expression of TLR4 and TAK1. TLR4, TAK1, and NLRP3 were collectively co-localized together and with the microglia marker Iba1. ConclusionsMicroglial TLR4 regulates TAK1 expression and phosphorylation and results in NLRP3 activation contributes to the development of morphine tolerance through regulating neuroinflammation. Targeting TLR4-TAK1-NLRP3 signaling to regulate neuro-inflammation will be alternative therapeutics and strategies for chronic morphine-induced antinociceptive tolerance.

Metabotropic glutamate receptor 5 signalling induced NMDA receptor subunits alterations during the development of morphine-induced antinociceptive tolerance in mouse cortex

Background and purposeThe aim of this study was to investigate the interactions between mGluR5 and NMDAR during the development of tolerance to the analgesic effect of chronic morphine following disruption of the link between mGluR5 and NMDAR. MethodsFor these experiments, mGluR5 knockout (KO) mice, Shank3 ΔC mutant mice and their littermate controls were used. Repeated morphine treatment through intrathecal injections (i.t. 10 μg/10 μl twice daily for 5 days) was used to induce antinociceptive tolerance and thermal hyperalgesia, which was determined by tail-flick latency and calculated as a percentage of the maximum possible effect (%MPE). Expression of NMDAR subunits (NR1, NR2A and NR2B) and phosphorylation of NR2B in cortex were evaluated by Western blot. ResultsChronic morphine treatment increased the total expression of mGluR5 and NR2A but not NR1 and NR2B in the cortex and enhanced the phosphorylation of NR2B in wild-type mice after tolerance developed. mGluR5 KOs had attenuated morphine-induced tolerance and thermal hyperalgesia and reduced chronic morphine-induced phosphorylation of NR2B but not total NR2A expression. Shank3 ΔC mice were used to imitate the loss of the physiological connection between mGluR5 and NMDAR, and the mice showed a similar analgesic effect to morphine as their littermate controls, and there were no differential effects on NMDAR subunits compared with controls during the development of morphine-induced tolerance. ConclusionsIt may be mGluR5-mediated PKC signalling that plays the most significant role in the mechanisms underlying morphine-induced antinociceptive tolerance rather than the physiological connection between mGluR5 and NMDAR.

Atorvastatin attenuates the antinociceptive tolerance of morphine via nitric oxide dependent pathway in male mice.

The development of morphine-induced antinociceptive tolerance limits its therapeutic efficacy in pain management. Atorvastatin, or competitive inhibitor of 3-hydroxy-methyl-glutaryl coenzyme A (HMG-CoA) reductase, is mainstay agent in hypercholesterolemia treatment. Beyond the cholesterol-lowering activity, exploration of neuroprotective properties of this statin indicates its potential benefit in central nervous disorders. The aim of the present study was to assess the effects of atorvastatin in development and expression of morphine-induced analgesic tolerance in male mice and probable involvement of nitric oxide. Chronic and acute treatment with atorvastatin 10 and 20mg/kg, respectively, could alleviate morphine tolerance in development and expression phases. Chronic co-administration of nitric oxide synthase (NOS) inhibitors including L-NAME (non selective NOS inhibitor; 2mg/kg), aminoguanidine (selective inducible NOS inhibitor; 50mg/kg) and 7-NI (selective neuronal NOS inhibitor; 15mg/kg) with atorvastatin blocked the protective effect of atorvastatin in tolerance reversal. Moreover, reversing the atorvastatin effect was also observed in acute simultaneous treatment of L-NAME (5mg/kg) and aminoguanidine (100mg/kg) with atorvastatin. Co-treatment of guanylyl cyclase inhibitor, ODQ (chronic dose: 10mg/kg and acute dose: 20mg/kg) was associated with prevention of atorvastatin anti-tolerance properties. Our results revealed that the atorvastatin modulating role in morphine antinociceptive tolerance is mediated at least in part via nitric oxide in animal pain models of hot plate and tail flick.

Involvement of neuronal TGF‐β activated kinase 1 in the development of tolerance to morphine‐induced antinociception in rat spinal cord

Tolerance induced by morphine and other opiates remains a major unresolved problem in the clinical management of pain. There is now good evidence for the importance of MAPKs in morphine-induced antinociceptive tolerance. A member of the MAPK kinase kinase family, TGF-β activated kinase 1 (TAK1) is the common upstream kinase of MAPKs. Here, we have assessed the involvement of TAK1 in the development of tolerance to morphine-induced analgesia. The effects of an antagonist of TAK1 on morphine tolerance were investigated in vivo using the Randall-Selitto test, and the mechanism was investigated using Western blot and immunohistochemistry. The expression of TAK1 after chronic morphine exposure was also evaluated in vitro by immunohistochemistry. Chronic intrathecal morphine exposure up-regulated protein levels and phosphorylation of spinal TAK1. TAK1 immunoreactivity was co-localized with the neuronal marker NeuN. Intrathecal administration of 5Z-7-oxozeaenol (OZ), a selective TAK1 inhibitor, attenuated the loss of morphine analgesic potency and morphine-induced TAK1 up-regulation. Furthermore, OZ decreased the up-regulated expression of spinal p38 and JNK after repeated morphine exposure. In vitro studies demonstrated that sustained morphine treatment induced TAK1 up-regulation, which was reversed by co-administration of OZ. A bolus injection of OZ showed some reversal of established morphine antinociceptive tolerance. TAK1 played a pivotal role in the development of morphine-induced antinociceptive tolerance. Modulation of TAK1 activation by the selective inhibitor OZ in the lumbar spinal cord may prove to be an attractive adjuvant therapy to attenuate such tolerance.

NADPH-oxidase 2 activation promotes opioid-induced antinociceptive tolerance in mice

The analgesic effectiveness of long-term opioid therapies is compromised by the development of antinociceptive tolerance linked to the overt production of peroxynitrite (ONOO−, PN), the product of the interaction between superoxide (O2−, SO) and nitric oxide (NO), and to neuroinflammatory processes. We have recently reported that in addition to post-translational nitration and inactivation of mitochondrial manganese superoxide dismutase (MnSOD), activation of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase holoenzyme (NOX) in the spinal cord is a major source for the overt production of superoxide-derived PN during the development of morphine-induced antinociceptive tolerance. However, the NOX complex involved in these processes is not known. The objective of these studies is to identify a potential role for the NOX2 complex, an enzyme involved in inflammation. Mice lacking the catalytic subunit of NOX2 (Nox2−/−) or its regulatory subunit, p47phox (p47phox−/−), developed antinociceptive tolerance similar to wildtype (wt) mice after 3days of continuous morphine. However, while wt mice continue to develop tolerance by day six, morphine analgesia was restored in both Nox2−/− and p47phox−/− mice. Moreover, the loss of Nox2 or p47 did not affect acute morphine analgesia in naïve mice. In wt mice, antinociceptive tolerance was associated with increased activation of NOX, nitration of MnSOD, and proinflammatory cytokines production in the spinal cord. These events were markedly attenuated in Nox2−/− and p47phox−/− mice and instead, there was enhanced formation of antiinflammatory cytokine (IL4 and IL10) production. These results suggest that NOX2 activity provides a significant source of superoxide-derived PN to undertake post-translational modifications of mitochondrial MnSOD and to engage neuroinflammatory signaling in the spinal cord associated with opioid-induced antinociceptive tolerance. Thus, NOX2 may provide a potential target for adjuvant therapy to protect opioid analgesia.

Supraspinal inactivation of mitochondrial superoxide dismutase is a source of peroxynitrite in the development of morphine antinociceptive tolerance

Effective treatment of chronic pain with morphine is limited by decreases in the drug's analgesic action with chronic administration (antinociceptive tolerance). Because opioids are mainstays of pain management, restoring their efficacy has great clinical importance. We have recently reported that formation of peroxynitrite (ONOO −, PN) in the dorsal horn of the spinal cord plays a critical role in the development of morphine antinociceptive tolerance and have further documented that nitration and enzymatic inactivation of mitochondrial superoxide dismutase (MnSOD) at that site provides a source for this nitroxidative species. We now report for the first time that antinociceptive tolerance in mice is also associated with the inactivation of MnSOD at supraspinal sites. Inactivation of MnSOD led to nitroxidative stress as evidenced by increased levels of products of oxidative DNA damage and activation of the nuclear factor poly (ADP-ribose) polymerase in whole brain homogenates. Co-administration of morphine with potent Mn porphyrin-based peroxynitrite scavengers, Mn(III) 5,10,15,20-tetrakis( N-ethylpyridinium-2-yl)porphyrin (MnTE-2-PyP 5+) and Mn(III) 5,10,15,20-tetrakis( N-n-hexylpyridinium-2-yl)porphyrin (MnTnHex-2-PyP 5+) (1) restored the enzymatic activity of MnSOD, (2) attenuated PN-derived nitroxidative stress, and (3) blocked the development of morphine-induced antinociceptive tolerance. The more lipophilic analogue, MnTnHex-2-PyP 5+ was able to cross the blood–brain barrier at higher levels than its lipophylic counterpart MnTE-2-PyP 5+ and was about 30–fold more efficacious. Collectively, these data suggest that PN-mediated enzymatic inactivation of supraspinal MnSOD provides a source of nitroxidative stress, which in turn contributes to central sensitization associated with the development of morphine antinociceptive tolerance. These results support our general contention that PN-targeted therapeutics may have potential as adjuncts to opiates in pain management.