The superoxide anion donor, potassium superoxide, induces pain and inflammation in mice through production of reactive oxygen species and cyclooxygenase-2

Abstract

It is currently accepted that superoxide anion (O2 •−) is an important mediator in pain and inflammation. The role of superoxide anion in pain and inflammation has been mainly determined indirectly by modulating its production and inactivation. Direct evidence using potassium superoxide (KO2), a superoxide anion donor, demonstrated that it induced thermal hyperalgesia, as assessed by the Hargreaves method. However, it remains to be determined whether KO2 is capable of inducing other inflammatory and nociceptive responses attributed to superoxide anion. Therefore, in the present study, we investigated the nociceptive and inflammatory effects of KO2. The KO2-induced inflammatory responses evaluated in mice were: mechanical hyperalgesia (electronic version of von Frey filaments), thermal hyperalgesia (hot plate), edema (caliper rule), myeloperoxidase activity (colorimetric assay), overt pain-like behaviors (flinches, time spent licking and writhing score), leukocyte recruitment, oxidative stress, and cyclooxygenase-2 mRNA expression (quantitative PCR). Administration of KO2 induced mechanical hyperalgesia, thermal hyperalgesia, paw edema, leukocyte recruitment, the writhing response, paw flinching, and paw licking in a dose-dependent manner. KO2 also induced time-dependent cyclooxygenase-2 mRNA expression in the paw skin. The nociceptive, inflammatory, and oxidative stress components of KO2-induced responses were responsive to morphine (analgesic opioid), quercetin (antioxidant flavonoid), and/or celecoxib (anti-inflammatory cyclooxygenase-2 inhibitor) treatment. In conclusion, the well-established superoxide anion donor KO2 is a valuable tool for studying the mechanisms and pharmacological susceptibilities of superoxide anion-triggered nociceptive and inflammatory responses ranging from mechanical and thermal hyperalgesia to overt pain-like behaviors, edema, and leukocyte recruitment.

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“…6). The doses of KO 2 and time points for sample collection were determined as previously described [33]. KO 2 was diluted in saline immediately before application.…”

“…injection of 1 mg of KO 2 [33]. Immediately after stimulus injection, each mouse was placed individually in a large glass cylinder, and the intensity of nociceptive behavior was quantified by counting the total number of writhings occurring between 0 and 20 min after stimulus injection.…”

“…Each mouse was placed in a large glass cylinder immediately after stimulus injection. The intensity of nociceptive behavior was quantified by counting the total number of paw flinches and the time (seconds) spent licking ipsilateral paw [33].…”

“…The animals were tested before and after treatments. The results are expressed by delta (D) withdrawal threshold (in grams) calculated by subtracting the mean measurements 0.5, 1, 3, 5 and 7 h after stimulus from the zero-time mean measurements [33].…”

“…Heat thermal hyperalgesia was performed using a hot plate at 55 ± 1°C [33]. The end-point was characterized by the removal of the paw followed by clear paw flinching or licking movements.…”