Hydromorphone efficacy and treatment protocol impact on tolerance and mu-opioid receptor regulation

Abstract

This study examined the antinociceptive (analgesic) efficacy of hydromorphone and hydromorphone-induced tolerance and regulation of mu-opioid receptor density. Initially s.c. hydromorphone's time of peak analgesic (tail-flick) effect (45 min) and ED50 using standard and cumulative dosing protocols (0.22 mg/kg, 0.37 mg/kg, respectively) were determined. The apparent analgesic efficacy (tau) of hydromorphone was then estimated using the operational model of agonism and the irreversible mu-opioid receptor antagonist clocinnamox. Mice were injected with clocinnamox (0.32-25.6 mg/kg, i.p.) and 24 h later, the analgesic potency of hydromorphone was determined. The tau value for hydromorphone was 35, which suggested that hydromorphone is a lower analgesic efficacy opioid agonist. To examine hydromorphone-induced tolerance, mice were continuously infused s.c. with hydromorphone (2.1-31.5 mg/kg/day) for 7 days and then morphine cumulative dose response studies were performed. Other groups of mice were injected with hydromorphone (2.2-22 mg/kg/day) once, or intermittently every 24 h for 7 days. Twenty-four hours after the last injection, mice were tested using morphine cumulative dosing studies. There was more tolerance with infusion treatments compared to intermittent treatment. When compared to higher analgesic efficacy opioids, hydromorphone infusions induced substantially more tolerance. Finally, the effect of chronic infusion (31.5 mg/kg/day) and 7 day intermittent (22 mg/kg/day) hydromorphone treatment on spinal cord mu-opioid receptor density was determined. Hydromorphone did not produce any change in mu-opioid receptor density following either treatment. These results support suggestions that analgesic efficacy is correlated with tolerance magnitude and regulation of mu-opioid receptors when opioid agonists are continuously administered. Taken together, these studies indicate that analgesic efficacy and treatment protocol are important in determining tolerance and regulation of mu-opioid receptors.

Fig. 1

Mice (N=5–9/dose) were injected with hydromorphone (0.2–1.25 mg/kg s.c.) and tested for antinociception (tail flick) at various time points (15–240 min). Each time action profile was determined once except for the 0.3125mg/kg dose which was determined twice and the combined data (mean + S.E.M.) are presented. Hydromorphone’s time of peak analgesic effect was estimated as 45 min.

Fig. 2

Dose response studies for hydromorphone were performed using standard (left panel) and cumulative dose response protocols (right panel) as described in the methods. For the standard dose response protocol, individual groups of mice (N=5–8/dose) were injected with a single dose of hydromorphone s.c. and tested for antinociception 45 min later. For the cumulative dose response protocol, mice (N=8) were injected with a starting dose of hydromorphone s.c. and tested for antinociception 45 min later. Mice that were not analgesic (i.e., tail flick latency< 10 s), were injected with another dose (see Methods) and retested. This cumulative dosing was continued until all mice were analgesic. The data presented are the combined results of five independent experiments for both cumulative and standard dosing protocols .The mean ED₅₀ (95% CL) for the standard dosing experiments was 0.22 mg/kg (0.20–0.24 mg/kg). The mean ED₅₀ (95% CL) for the cumulative dosing experiments was 0.37 mg/kg (0.31–0.43 mg/kg).

Fig. 3

The effect of clocinnamox (CCAM) treatment on the analgesic potency of hydromorphone. Mice (N=6–10/treatment) were injected i.p. with CCAM or saline (control) and 24h later hydromorphone cumulative dose response studies (tail flick) were conducted. The data for control represent the mean of three experiments, while the CCAM treatment results are from one experiment for each dose. The data plotted are the mean tail flick latency as a function of cumulative dose. EC₅₀ estimates were calculated for each treatment (see Table 1).

Fig. 4

The effect of hydromorphone treatment on morphine analgesic potency. For infusion studies, mice (N=5–8/dose) were infused for 7 days with hydromorphone (2.1, 10.5, 21, 31.5mg/kg/day; which is equivalent to ≈10 – 150 times the ED₅₀ for hydromorphone). Controls (C) were implanted with placebo pellets. Pumps and pellets were removed at the end of treatment and 16hr later, a morphine cumulative dose response (tail flick) study was conducted. All infusion results are from one determination, except the data for 21 mg/kg/day which are the combined data from two experiments. For acute (one s.c. injection) and intermittent (7 daily s.c. injections) studies, mice (N=8–10/dose) were injected with hydromorphone (2.2–22mg/kg/day; equivalent to ≈10–100 times the ED₅₀). Controls were injected with saline. Morphine cumulative dose response (tail flick) studies were conducted 24 hr after the last treatment. The data for acute and intermittent experiments represent the mean of three experiments. The shift in the ED₅₀ relative to each individual control is presented as a function of the multiple of ED₅₀ for hydromorphone (0.22mg/kg) as determined using the standard dosing protocol. (see Table 3). * significantly different from control (P<0.05).

Fig. 5

The effect of hydromorphone treatment on μ-opioid receptor density in mouse spinal cord. Panel A. Mice (N = 10) were injected s.c. for 7 days with hydromorphone, (22mg/kg/day; equivalent to ≈100 times the standard dose ED₅₀). Controls (N = 10) were injected with saline. Twenty-four h after the last injection mice were sacrificed and spinal cords collected for saturation binding studies ([³H] DAMGO). The B_max (95% CL) was 171 fmol/mg protein (163–179) and 161 (156–166) for control and hydromorphone groups, respectively. K_D ‘s (95% CL) for control and treated groups were 1.0nM (0.8–1.2) and 1.0 (0.8–1.0), respectively. Similar results were found in 2 other experiments. Panel B. Mice (N=10) were infused s.c. with hydromorphone (31.5mg/kg/day; equivalent to ≈150 times the standard dose ED₅₀) for 7 days. Controls (N=10) were implanted with placebo pellets. Pumps and pellets were removed at the end of treatment, and 16 h later, mice were sacrificed and spinal cords collected for ([³H]DAMGO) saturation binding studies. The B_max (95% CL) was 191 fmol/mg protein (182–200) and 193 (181–206), for control and hydromorphone groups, respectively. K_D’s (95% CL) for control and treated groups were 1.2nM (1.1–1.4) and 1.5 (1.2–1.7), respectively. Similar results were found in 2 other experiments. There were no significant differences (P > 0.05) between the groups for B_max or K_D for either treatment.

Fig. 6

The effect of chronic (7 day) infusion with hydromorphone, oxycodone and etorphine on morphine analgesic potency. Oxycodone and etorphine data are from Pawar et al., 2007 using a treatment protocol identical to that used for hydromorphone. The shift in ED₅₀ relative to control was calculated as: ED_{50 treated} / ED_{50 control}.