Peripherally administered endomorphin-1 attenuates nociceptive discharge evoked by noxious mechanical stimulation in a rat model of chronic inflammatory joint hypersensitivity

Iain Strickland1, Alison Reeve2, Daniel McQueen1
1The University of Edinburgh, Edinburgh, United Kingdom, 2GlaxoSmithKline, Essex, United Kingdom

Endomorphin-1 (EM-1) is an endogenous tetrapeptide with high selectivity and affinity for the μ-opioid receptor (Zadina et al., 1997). EM-1 has similar functional properties to morphine and has been shown to inhibit nociceptive activity in primary afferents during noxious mechanical hyper-rotation of acutely inflamed knee joints (Li et al., 2005). The aim of this in vivo electrophysiology study was to determine whether EM-1 could attenuate mechanical joint hypersensitivity 14-23 days after induction of an inflammation in the knee joint (Donaldson et al., 1993).

Unilateral knee joint arthritis was induced in the left joint of adult male Wistar rats (n=15, 150-200g) by a 150μl intra-articular injection of Freund’s complete adjuvant (FCA; 1mg.ml-1 Mycobacterium tuberculosis, Sigma, UK). Rats were transiently anaesthetised using 3% isoflurane in oxygen and FCA was injected into the knee joint space. On days 14-23 animals were anaesthetised by an i.p. injection of pentobarbitone sodium (60mg.ml-1 at 10μl.kg-1), maintained throughout the experiments by an i.v. infusion of pentobarbitone (7.8mg.hr-1). Extracellular neuronal recordings were made from the medial articular nerve (MAN) of the ipsilateral knee joint (Gauldie et al., 2001). The MAN was mechanically stimulated using von Frey filaments of 1g, 7g and 21g at a fixed position within the receptive field on the exposed surface of the knee joint. Filaments were tested in ascending order for a period of 10s with a break of 10s in between each stimulation. Applications of the hairs were repeated every 20mins. The effect of each individual filament was measured as the change in the action potential frequency (impulses.s-1) during the test period compared to that of the preceding ten seconds. Five tests were performed before and after dosing with EM-1 (100μg, n=5), naloxone (500μg) and EM-1 (100μg, n=5) and saline vehicle (n=5). Drugs were administered in a volume of 100μl close to the knee via a femoral arterial cannula. The mean (± SEM) normalised frequency of firing was calculated for each individual filament, with the highest frequency within each experiment being designated 100%. Data were analysed using a one-way analysis of variance (ANOVA, Kruskal Wallis) where P<0.05 was considered statistically significant.

Results showed that EM-1 significantly inhibited firing of the MAN evoked by the 7g and 21g von Frey filaments. The mean discharge evoked by the 7g filament fell significantly from 6.7 ± 1.6 impulses.s-1 to 1.3 ± 0.3 impulses.s-1 (P<0.05, 46 ± 5% to 10 ± 2%) and the firing frequency evoked by the 21g filament fell significantly from 9.7 ± 2 impulses.s-1 to 2.2 ± 0.4 impulses.s-1 (P<0.05, 70.8 ± 5% to 24 ± 5%). This significant inhibition was reduced when naloxone was administered before the EM-1. The vehicle had no significant effect on the frequency of firing evoked by any of the filaments tested.

These results suggest that μ-opioid receptors are present and functional in the periphery on the MAN during the advanced stages (days 14-23) of this model of chronic inflammatory joint hypersensitivity. This is further evidence that peripherally acting opioids could play an important role in managing chronic pain in the clinic, without the unwanted side effects of the centrally acting opioid morphine.

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