Effects of mu- and kappa-2 opioid receptor agonists on pain and rearing behaviors

Abstract

Background: Management of pain involves a balance between inhibition of pain and minimization of side effects; therefore, in developing new analgesic compounds, one must consider the effects of treatment on both pain processing and behavior. The purpose of this study was to evaluate the effects of the mu and kappa-2 opioid receptor agonists on general and pain behavioral outcomes.

Methods: As a general behavioral assessment, we modified the cylinder rearing assay and recorded the number and duration of rearing events. Thermal sensitivity was evaluated using either a reflexive measure of hindpaw withdrawal latency to a radiant heat source or using an orofacial operant thermal assay. Acetic acid-induced visceral pain and capsaicin-induced neurogenic inflammatory pain were used as painful stimuli. The mu-opioid receptor agonist, morphine or the kappa-2 receptor agonist GR89696 was administered 30 min prior to testing. A general linear model repeated measures analysis was completed for baseline session comparisons and an analysis of variance was used to evaluate the effects of treatment on each outcome measure (SPSS Inc). When significant differences were found, post-hoc comparisons were made using the Tukey honestly significant difference test. *P < 0.05 was considered significant in all instances.

Results: We found that morphine and GR89,696 dose-dependently decreased the number of reaching events and rearing duration. Rearing behavior was not affected at 0.5 mg/kg for morphine, 1.25 x 10-4 mg/kg for GR89,696. Hindpaw thermal sensitivity was significantly increased only at the highest doses for each drug. At the highest dose that did not significantly influence rearing behavior, we found that visceral and neurogenic inflammatory pain was not affected following GR89,696 administration and morphine was only partially effective for blocking visceral pain.

Conclusion: This study demonstrated that high levels of the opioids produced significant untoward effects and made distinguishing an analgesic versus a more general effect more difficult. Quantification of rearing behavior in conjunction with standard analgesic assays can help in gaining a better appreciation of true analgesic efficacy of experimental drugs.

Figure 1

Illustrated is an animal that is at rest (A) and rearing (B) in the rearing assay testing device. An example of the trace recording is illustrated in panel C with the arrows denoting distinct rearing events. Animals (N = 12) tested on three consecutive days displayed a significant decrease (*P < 0.05) in the number of reaches (D), but not in the total rearing duration (E). The raw number of events or time (s) is denoted in the parentheses.

Figure 2

We evaluated the effects of GR89,696 and morphine on reaching activity and cumulative rearing duration. There was a significant dose-related decrease (*P < 0.05) in the number of reaching events (A) and total time spent rearing (B) following morphine (N = 8–12, right insets) or GR89,696 (N = 6, left insets) administration, compared to baseline values.

Figure 3

The effects of GR89,696 and morphine on normal thermal sensitivity were assessed and demonstrated that hindpaw withdrawal latency to thermal stimulation was significantly increased only at the highest doses tested for GR89,696 (N = 6) and morphine (N = 6).

Figure 4

We found that GR89,696 does not affect operant thermal outcome measures. Animals treated with either GR89,696 (1.25 × 10^-4 mg/kg, subcutaneous) or vehicle (PBS, subcutaneous) prior to facial capsaicin application did not significantly differ from each other when evaluated using the six thermal operant outcome measures (N = 10). Both groups were significantly lower (*P < 0.05) than baseline values in all instances except for GR89696 in Facial Contact Duration (B) and for both GR89696 and vehicle for Facial Duration/Contact (F). Overall, this indicates that the neurogenic inflammatory pain was not inhibited following capsaicin application.

Figure 5

The effects of morphine and GR89,696 on reaching activity and cumulative rearing duration following visceral pain induction was determined. Acetic acid was injected (intraperitoneal) as an acute painful stimulus and produced a significant decrease (*P < 0.05, compared to baseline) in both reaching number (A) and rearing duration (B) during the first 15 min post-injection. Animals were fully recovered by 24 h, as their rearing behavior returned to baseline levels. GR89,696 (N = 8, 1.25 × 10^-4 mg/kg, subcutaneous) was ineffective and morphine (N = 8, 0.5 mg/kg) was only partially effective for blocking this visceral pain induction, as demonstrated by a decrease in reaching number (C) and rearing duration (D). For (C, D), *P < 0.05 as compared to naïve animals.