In vivo activation of a mutant mu-opioid receptor by naltrexone produces a potent analgesic effect but no tolerance: role of mu-receptor activation and delta-receptor blockade in morphine tolerance

Abstract

Opioid analgesics are the standard therapeutic agents for the treatment of pain, but their prolonged use is limited because of the development of tolerance and dependence. Recently, we reported the development of a mu-opioid receptor knock-in (KI) mouse in which the mu-opioid receptor was replaced by a mutant receptor (S196A) using a homologous recombination gene-targeting strategy. In these animals, the opioid antagonist naltrexone elicited antinociceptive effects similar to those of partial agonists acting in wild-type (WT) mice; however, development of tolerance and physical dependence were greatly reduced. In this study, we test the hypothesis that the failure of naltrexone to produce tolerance in these KI mice is attributable to its simultaneous inhibition of delta-opioid receptors and activation of mu-opioid receptors. Simultaneous implantation of a morphine pellet and continuous infusion of the delta-opioid receptor antagonist naltrindole prevented tolerance development to morphine in both WT and KI animals. Moreover, administration of SNC-80 [(+)-4-[(alphaR)-alpha-((2S,5R)-4-allyl-2,5-dimethyl-1-piperazinyl)-3-methoxybenzyl]-N,N-diethylbenzamide], a delta agonist, in the naltrexone-pelleted KI animals resulted in a dose-dependent induction in tolerance development to both morphine- and naltrexone-induced analgesia. We conclude that although simultaneous activation of both mu- and delta-opioid receptors results in tolerance development, mu-opioid receptor activation in conjunction with delta-opioid receptor blockade significantly attenuates the development of tolerance.

Figure 1.

Dose-response effect of morphine- and naltrexone-induced analgesia in WT and KI animals after chronic placebo, morphine, or naltrexone treatment. WT (A) or KI (B, C) animals were implanted with a placebo, morphine, or naltrexone pellet for 72 h, and morphine-induced (A, B) or naltrexone-induced (C) analgesia was measured by the tail-withdrawal test 8 h after pellet withdrawal. There is a significant difference between the placebo-treated and the morphine-treated group in all three experiments. ^*p < 0.05, significant difference from the placebo-treated group (Student's t test). Each dose was tested on at least 8-10 animals. % MPE, Percentage of maximum possible effect. Error bars represent SEM.

Figure 2.

Dose-response effect of morphine-induced analgesia in WT (A) and KI (B, C) animals after chronic naltrindole treatment in WT and KI animals. WT (A) or KI (B, C) animals were implanted with placebo, morphine (A, B), or naltrexone (NLTRX) (C) pellets and infused with either saline or naltrindole (Alzet pump; 10 mg · kg^-1 · 24 h^-1) for 72 h. Morphine-induced analgesia was measured as described above. There was a significant difference between the placebo- plus saline-treated and the morphine- plus saline-treated groups (p < 0.05). There was no significant difference between the placebo plus saline group and the morphine plus NTI group (p > 0.1). ^*p < 0.05, significant difference from the placebo-treated group (Student's t test). No significant difference was observed among the three treatment groups in the naltrexone-pelleted animals (C); p > 0.1 (Student's t test). Each dose was tested on at least 8-10 animals. % MPE, Percentage of maximum possible effect. Error bars represent SEM.

Figure 3.

A, Effect of naltrindole treatment on naltrexone (NLTRX)-induced analgesia in KI animals (as in Fig. 2C, except that naltrexone-induced analgesia was measured). No significant difference was observed among the three treatment groups; p > 0.06 (Student's t test). Each dose was tested on at least 8-10 animals. B, Effect of naltrindole treatment on morphine-induced analgesia after tolerance development. WT animals were implanted with a placebo or morphine pellet for 72 h. At 24 h before pellet removal, animals were implanted with an Alzet pump delivering 10 mg · kg^-1 · 24 h^-1 NTI. Morphine-induced analgesia was measured as described above. ^*p < 0.05, significant difference between the morphine- plus vehicle-treated group and the placebo-treated group; + indicates a significant difference between the morphine plus NTI group and the placebo-treated group (Student's t test). Each dose was tested on at least 8-10 animals. % MPE, Percentage of maximum possible effect. Error bars represent SEM.

Figure 4.

Effect of SNC-80 treatment on morphine-induced (A) and naltrexone-induced (B) analgesia in KI animals. KI animals were implanted with a placebo or a naltrexone (NLTRX) pellet and infused with either vehicle or SNC-80 (Alzet pump; 10 mg · kg^-1 · 24 h^-1) for 72 h; morphine-induced (A) or naltrexone-induced (B) analgesia was then measured. There was a significant difference between the placebo- plus vehicle-treated group and the naltrexone- plus SNC-80-treated group; p < 0.05 (Student's t test). Each asterisk indicates a significant difference from the placebo-treated group. Each dose was tested on at least 8-10 animals. C, Dose-response effect of SNC-80 infusion on morphine-induced analgesia in KI animals. KI animals were implanted with a naltrexone pellet and infused with either vehicle or varying concentrations of SNC-80 (Alzet pump; 0.1, 1, and 10 mg · kg^-1 · 24 h^-1) for 72 h; morphine-induced analgesia was then measured. There was a significant difference between the naltrexone- plus vehicle-treated group and the naltrexone- plus 1 mg/kg SNC-80-treated and naltrexone- plus 10 mg/kg SNC-80-treated groups; p < 0.05 (Student's t test). Each dose was tested on at least 8-10 animals. D, Different durations of SNC-80 exposure on morphine-induced analgesia in KI animals. KI animals were implanted with a naltrexone pellet and infused with either vehicle or SNC-80 (Alzet pump; 10 mg · kg^-1 · 24 h^-1) for 24, 48, and 72 h; morphine-induced analgesia was then measured. There was a significant difference between the naltrexone- plus vehicle-treated group and the naltrexone plus 72 h SNC-80 group; p < 0.05 (Student's t test). Each asterisk indicates a significant difference from the naltrexone- plus vehicle-treated group. Each dose was tested on at least 8-10 animals. %MPE, Percentage of maximum possible effect. Error bars represent SEM.