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 3 days 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.

► NOX2 holoenzyme is critical for the development of antinociceptive tolerance.
► NOX2 is a major source of superoxide-derived PN during late stages of tolerance.
► NOX2 activity drives proinflammatory cytokine production.
► The loss of NOX2 activity attenuates tolerance and reduces nitroxidative stress.
► The loss of NOX2 activity favors antiinflammatory cytokine production.