Daniel E. Womer

Intrathecal pertussis toxin produces hyperalgesia and allodynia in mice.

Abstract Pertussis toxin (PTX), which causes the ADP‐ribosylation and thereby inactivation of Gi‐proteins, has been employed in analgesia testing to elucidate receptors that are coupled to inhibitory G‐proteins, such as the mu‐opioid receptor. Consistent with previous findings, the antinociceptive effects of morphine (1–10 &mgr;g) as measured by tail‐flick latency using a 55°C water bath, were blocked by a single intrathecal injection of 0.5 &mgr;g PTX 6 days prior to intrathecal morphine administration. In addition, mice treated intrathecally with 0.5 &mgr;g of PTX had significantly shorter baseline tail‐flick latencies compared with vehicle treated mice using a 55°C water bath when tested 6 days after PTX or vehicle administration. Morphine‐induced antinociception was blocked in a dose‐dependent manner by PTX with complete blockade of morphine following a 0.3‐&mgr;g dose of PTX. Further, mice administered 0.1 &mgr;g or 0.3 &mgr;g PTX intrathecally had significantly shorter tail‐flick latencies compared with vehicle injected mice using a 40, 45 or 50°C water bath when tested 7 days after intrathecal injection. Shorter tail‐flick latencies were observed at 45°C as early as 48 h after intrathecal administration of 0.03, 0.1 or 0.3 &mgr;g PTX and these shorter tail‐flick latencies were observed up to 90 days after intrathecal PTX administration. The intrathecal administration of PTX caused hyperalgesia and allodynia that appears similar to the symptoms reported by patients suffering from neuropathic pain, and suggests that deficiencies in inhibitory systems, as compared with increases in excitatory systems, may play a role in the pathophysiology of at least some central or neuropathic pain states.

In vivo pharmacology of butylthio[2.2.2] (LY297802 / NNC11-1053), an orally acting antinociceptive muscarinic agonist

Butylthio[2.2.2] (LY297802 / NNC11-1053) is a mixed muscarinic cholinergic receptor agonist/antagonist that produces antinociception in mice and rats. As such, butylthio[2.2.2] may have therapeutic utility in the treatment of pain. Butylthio[2.2.2] was fully efficacious in the mouse grid shock, writhing, tail-flick and hot plate tests with ED50 values ranging from 1.5 to 12.2 mg/kg after oral administration. In contrast, the ED50 values for morphine ranged from 7.3 to 72 mg/kg after oral administration. Scopolamine was a competitive antagonist of the antinociceptive effects of butylthio[2.2.2]. Butylthio[2.2.2] did not produce either salivation or tremor at therapeutic doses; rather, there was a 50- to >100-fold separation between therapeutic doses and doses which produced side-effects. Butylthio[2.2.2] had high affinity for muscarinic receptors, but little if any affinity for other neurotransmitter receptors or uptake sites. In isolated tissues, butylthio[2.2.2] was an agonist with high affinity at M1 receptors in rabbit vas deferens, an antagonist at M2 receptors in guinea pig atria as well as an antagonist at M3 receptors in guinea pig urinary bladder. Although it has been suggested that M1 receptors mediate the antinociceptive effects of muscarinic agonists, M1 efficacy is not a requirement for antinociception, and, in vivo, the antinociceptive effects of muscarinic agonists are blocked by the intrathecal administration of pertussis toxin, indicating the involvement of m2 or m4 receptors. Since butylthio[2.2.2] is an M2 antagonist, antinociception is therefore most likely mediated by m4 receptors. Butylthio[2.2.2] is currently undergoing clinical development as a novel analgesic.

Pharmacologic reversal of pertussis toxin-induced thermal allodynia in mice.

We have previously demonstrated that the intrathecal administration of pertussis toxin produces a long-lasting thermal allodynia in mice. The purpose of the present studies was to compare the antinociceptive and the antiallodynic effects of drugs that are commonly used in treating neuropathic allodynia in untreated mice and in mice which had been administered vehicle or pertussis toxin intrathecally 7 days previously. In untreated mice, morphine, fentanyl, clonidine, oxymetazoline, desipramine and lidocaine, but not MK801, produced dose-related antinociception when tested using a 55 degrees C water tail-flick test. However, 7 days after the intrathecal injection of pertussis toxin, which induced a condition of thermal allodynia when tested using a 45 degrees C water bath, the full opioid and the full alpha(2)-adrenergic receptor agonists fentanyl and clonidine, but not the partial opioid nor the partial alpha(2)-adrenergic receptor agonists morphine and oxymetazoline, reversed the pertussis toxin-induced thermal allodynia. Moreover, lidocaine, desipramine, carbamazepine and MK801 failed to reverse the pertussis toxin-induced thermal allodynia. The present results suggest that decrements in G(i)/G(o)-protein function may be involved in initiating and/or maintaining some neuropathic pain states. Moreover, the results of the present study suggest that the use of full, but not partial, opioid or alpha(2)-agonists may be useful in the treatment of thermal allodynic pain states which may be due at least in part to inhibitory second messenger system dysfunction. Further, the underlying biochemistry of the apparent allodynic pain state induced by intrathecal administration of pertussis toxin warrants further investigation.