Ultra-low dose naloxone restores the antinociceptive effect of morphine in pertussis toxin-treated rats by reversing the coupling of μ-opioid receptors from Gs-protein to coupling to Gi-protein

Pertussis toxin (PTX) treatment results in ADP-ribosylation of Gi-protein and thus in disruption of μ-opioid receptor signal transduction and loss of the antinociceptive effect of morphine. We have previously demonstrated that pretreatment with ultra-low dose naloxone preserves the antinociceptive effect of morphine in PTX-treated rats. The present study further examined the effect of ultra-low dose naloxone on μ-opioid receptor signaling in PTX-treated rats and the underlying mechanism. Male Wistar rats implanted with an intrathecal catheter received an intrathecal injection of saline or PTX (1 μg in 5 μl of saline), then, 4 days later, were pretreated by intrathecal injection with either saline or ultra-low dose naloxone (15 ng in 5 μl of saline), followed, 30 min later, by saline or morphine (10 μg in 5 μl of saline). Four days after PTX injection, thermal hyperalgesia was observed, together with increased coupling of excitatory Gs-protein to μ-opioid receptors in the spinal cord. Ultra-low dose naloxone pretreatment preserved the antinociceptive effect of morphine, and this effect was completely blocked by the μ-opioid receptor antagonist CTOP, but not by the κ-opioid receptor antagonist nor-BNI or the δ-opioid receptor antagonist naltrindole. Moreover, a co-immunoprecipitation study showed that ultra-low dose naloxone restored μ-opioid receptor/Gi-protein coupling and inhibited the PTX-induced μ-opioid receptor/Gs-protein coupling. In addition to the anti-neuroinflammatory effect and glutamate transporter modulation previously observed in PTX-treated rats, the re-establishment of μ-opioid receptor Gi/Go-protein coupling is involved in the restoration of the antinociceptive effect of morphine by ultra-low dose naloxone pretreatment by normalizing the balance between the excitatory and inhibitory signaling pathways. These results show that ultra-low dose naloxone preserves the antinociceptive effect of morphine, suppresses spinal neuroinflammation, and reduces PTX-elevated excitatory Gs-coupled opioid receptors in PTX-treated rats. We suggest that ultra-low dose naloxone might be clinically valuable in pain management.