Role of Acetaldehyde in Ethanol Reversal of Tolerance to Morphine-Induced Respiratory Depression in Mice

Abstract

Background: Opioid users regularly consume other drugs such as alcohol (ethanol). Acute administration of ethanol rapidly reverses tolerance to morphine-induced respiratory depression. However, recent research has suggested that the primary metabolite of ethanol, acetaldehyde, may play a key role in mediating the CNS effects seen after ethanol consumption. This research investigated the role of acetaldehyde in ethanol reversal of tolerance to morphine-induced respiratory depression.

Methods: Tolerance was induced in mice by 6-days implantation of a 75 mg morphine pellet with control mice implanted with a placebo pellet. Tolerance was assessed by acute morphine administration on day 6 and respiration measured by plethysmography. Levels of acetaldehyde were inhibited or enhanced by pre-treatments with the acetaldehyde chelator D-penicillamine and the inhibitor of acetaldehyde dehydrogenase disulfiram respectively.

Results: Morphine pellet implanted mice displayed tolerance to an acute dose of morphine compared to placebo pellet implanted controls. Acute acetaldehyde administration dose-dependently reversed tolerance to morphine respiratory depression. As previously demonstrated, ethanol reversed morphine tolerance, and this was inhibited by D-penicillamine pre-treatment. An acute, low dose of ethanol that did not significantly reverse morphine tolerance was able to do so following disulfiram pre-treatment.

Conclusion: These data suggest that acetaldehyde, the primary metabolite of ethanol, is responsible for the reversal of morphine tolerance observed following ethanol administration.

Keywords: acetaldehyde; ethanol; morphine; opioid; polydrug; polypharmacology; respiration; tolerance.

FIGURE 1

Induction of morphine tolerance. (A) Acute injection of morphine (10 mg/kg i.p.), significantly depressed minute volume (MV) in 6 days PP- but not 6 days 75 mg MP-implanted, mice. (B) Data in (A) replotted as percentage of baseline MV. (C) Area under the curve (AUC) calculated from (B) (as difference from 100%). All data are presented as mean ± s.e.m. Statistical comparison made by 2-way ANOVA with Bonferroni’s comparison (A) (F (DFn, DFd—F (50, 300) = 5.034), (B) (F (DFn, DFd—F (7, 70) = 11.96) or unpaired t-test (C). * indicates p < 0.05 compared to placebo. n = 6 for each group.

FIGURE 2

Acetaldehyde modulates morphine tolerance. (A,B) Acute administration of acetaldehyde 50 mg/kg i.p (open symbols) or 100 mg/kg i.p. (closed symbols) did not depress minute volume (MV) in mice implanted for 6 days with a 75 mg MP. (C,D) Acute administration of morphine (10 mg/kg i.p.) did not depress respiration in MP- implanted mice indicating that tolerance had developed. Co-administration of acetaldehyde (Acet) and morphine depressed respiration in an acetaldehyde dose-dependent manner. (E) Area under the curve (AUC) analysis of data in (B,D) shows that when acetaldehyde 50 mg/kg or 100 mg/kg was administered along with acute morphine respiratory depression was restored i.e. morphine tolerance had been reversed. MP, morphine pellet; PP, placebo pellet. All data are presented as mean ± s.e.m. Statistical comparison made by One-way ANOVA with Bonferroni’s comparison (F (DFn, DFd—F (5, 29) = 9.971) in (E). * indicates p < 0.05 as shown. n = 6 for each group.

FIGURE 3

Pre-treatment with D-penicillamine inhibits ethanol reversal of morphine tolerance. (A) Morphine (10 mg/kg i.p.) co-administered with ethanol (0.3 g/kg i.p.) to 6 days placebo pellet implanted mice induced the same level of respiratory depression with and without a 30 min pre-treatment (PT) with D-penicillamine (DP, 50 mg/kg i.p.). (B) Administration of morphine, ethanol or saline (i.p.) to 6 days 75 mg morphine pellet implanted mice did not depress respiration following a 30 min pre-treatment with D-penicillamine. (C) Pre-treatment with D-penicillamine (DP, 50 mg/kg i.p.) significantly reduced the level of respiratory depression induced by morphine and ethanol co-administration in 6 days MP- implanted mice. (D) Area under the curve (AUC) analysis of data in (A,C). MP, morphine pellet; PP, placebo pellet. All data are presented as mean ± s.e.m. Statistical comparison made by Two-way ANOVA with Bonferroni’s comparison (F (DFn, DFd—F (1, 20) = 5.094) in (D). * indicates p < 0.05 as shown. n = 6 for each group.

FIGURE 4

Pre-treatment with disulfiram enhances ethanol reversal of morphine tolerance. (A,B) Morphine (10 mg/kg i.p.) induced no respiratory depression in 6 days 75 mg MP-implanted mice with or without a 30 min disulfiram (DF) (40 mg/kg i.p.) pre-treatment. Similarly, co-administration of morphine with a low dose of ethanol (0.1 g/kg i.p.) did not induce significant respiratory depression in MP- implanted mice. Pre-treatment (PT) with disulfiram (40 mg/kg i.p.) in morphine pellet implanted mice enhanced the degree of respiratory depression to co-administration of morphine and ethanol. (C) Area under the curve (AUC) analysis of data in (B) shows that pre-treatment with disulfiram (40 mg/kg i.p.) in MP- implanted mice prior to acute co-administration of morphine and ethanol significantly enhanced the degree of respiratory depression observed when compared to morphine and ethanol co-administered in vehicle pre-treated mice. MP, morphine pellet; PP, placebo pellet. All data are presented as mean ± s.e.m. Statistical comparison made by One-way ANOVA (C) with Bonferroni’s comparison (F (DFn, DFd—F (3, 20) = 0.570). * indicates p < 0.05 as shown. n = 6 for each group.