Kappa antagonist to prevent drug relapse: does stage in addiction cycle matter?

Introduction: An upregulation of the kappa opioid receptor (KOR) system during preoccupation stage in addiction cycle will cause dysphoria among addicts which can lead to relapse. Therefore, KOR antagonism might hold the key to prevent relapse. In this study, we aim to identify the exact addiction's stage in which KOR antagonist can be given. Methods: Using a conditioned place preference (CPP) model, adult Swiss albino mice were divided into two major groups. The first group received treatment at the initial stage of morphine withdrawal (7.5 mg/kg. i.p) while the second group received treatment after complete abstinence was achieved. Each major groups were further divided into two treatment groups (n=8-12), either received a functional KOR antagonist (0.3 mg/kg buprenorphine/ 1mg/kg naltrexone combination, i.p) or a selective KOR antagonist (10 mg/kg nor-BNI, i.p) prior to morphine priming (2.5 mg/kg, i.p). All data were analyzed using paired sample t-test. Results: The results showed that relapse was successfully attenuated in the groups that received KOR antagonists only after complete abstinence was successfully achieved (not significantly different from their baseline). However, the mice developed unusual sign of behavior sensitization (intermittent freezing, licking) when buprenorphine/naltrexone combination was given at initial stage of withdrawal. Conclusion: Our initial findings suggest that KOR antagonism might be beneficial only after the addicts achieved complete abstinence to prevent future drug-induced relapse. Brain study should be conducted to explain the unusual behavior seen when the drug intervention is given at an earlier stage of withdrawal

The influence of duration of confinement towards morphine dependence intensity in conditioned place preference (CPP) model

Introduction: Conditioned Place Preference (CPP) continues to be one of the most popular models to study the dependence effects of substance abuse. This study is divided into three main phases which are pre-conditioning, conditioning and post-conditioning phases. It can be conducted using different protocols which can be used to investigate the effects of drug dependence. Since there is not much study done to investigate the effects of duration of confinement, this study is conducted to figure out the effects of different duration of confinement towards intensity of morphine dependence. Methods: Using adult male Swiss albino mice, 12 mice were used for each group with different duration of confinement times, 40 to 60 minutes respectively. During conditioning phase, the mice received 7.5 mg/kg morphine and saline through intraperitoneal route on alternate day for 6 days. Result: Using unbiased method, it was found that both groups had significantly developed dependence (P<0.05, P<0.01). Later, a comparison between these groups had shown that the group of mice that underwent 60 minutes confinement time showed greater intensity of dependence than the other group, although the difference was not statistically significant. Interestingly, it was also noted that the number of mice that was excluded after post-conditioning test was higher in the group that underwent 40 minutes confinement cpmpared to the counterpart (n=5, n=2). Conclusion: These results proved that increasing in confinement time caused an increase in the intensity of morphine dependence in mice. Therefore, it is suggested that 60 minutes confinement time is more suitable for morphine to develop dependence in mice and to minimise subject elimination throughout a long cycle of CPP procedure, provided that other variables are maintained

The effect of dose on the intensity of dependence in morphine-dependent mice

Introduction: Animal behaviour research is an important tool to study the addiction properties of addictive substances. The Conditioned Place Preference (CPP)paradigm is among the model used to achieve this purpose. It can be used to investigate a wide range of addiction related behaviour including intoxication and relapse potential. Since it involves a long cycle experiment, it is important to optimise the experimental conditions, but minimising any unnecessity. Therefore, in this study we aim to investigate the effects of different morphine dose (3.75 - 10 mg/kg) towards intensity of morphine dependence in adult Swiss albino mice. Methods: The mice were divided into four group (n=10-12 for each group), where each group received different doses of morphine and saline on alternate day for 6 days during conditioning phase. The intensity of dependence was measured during post-conditioning phase where the preference at the drug-paired compartment was calculated during a 15 minutes observation. Results: The result has shown that three out of four groups significantly developed dependence (P < 0.05). However, the group of mice that received highest morphine dose (10 mg/kg) did not develop dependence. When baseline was compared, it was found that the mice that received the lowest morphine treatment (3.75 mg/kg) shows the highest increment from their baseline (164.7±33.3 seconds), while the group that received 10 mg/kg morphine shows the lowest increment (119.0±31.6 seconds). The trend of increment at the drug-paired compartment shows a declining trend when the dose of morphine is increased. Conclusion: Therefore, it is concluded that the optimum dose of morphine that should be administered to the adult Swiss albino mice is around 3.75 mg/kg, provided that all other experimental conditions and protocols are maintained

Anti-stress properties of buprenorphine: a better choice for stress-induced relapse to drug addiction

Compared to methadone, buprenorphine has a unique pharmacological profile in which it is a partial agonist at the mu- and NOP, but antagonist at the kappa-opioid receptors. This gives advantage to buprenorphine to have lesser abusive tendency and has ability to reduce relapse to drug taking. Previous studies had shown that stress, which has been linked to relapse, increased the release of dynorphin (an endogenous kappa-opioid receptor agonist) and potentiates the rewarding effects of morphine. In this study, we tested if buprenorphine treatment has ability to block the drug seeking behaviour in mice using a conditioned place preference (CPP) paradigm. Initially, a modified forced-swim test (FST) was conducted for two consecutive days to induce stress by testing for immobility in mice. A significant difference between control and treatment groups (n = 8 for each group, P 0.05) in the time spent for the treatment group (n = 8) during post morphine-conditioning session compared to their baseline, which suggested that buprenorphine has the ability to prevent morphine-seeking behaviour as compared to the control group (n = 8, P < 0.05) . These results suggest the advantage of buprenorphine as therapeutic agent to treat stress-induced relapse in drug addiction

The pyrrolidine ring and its relation with the psychoactivity of mitragynine

Mitragynine, the major alkaloid of Mitragyna speciosa has been widely studied for its anti-addictive properties. Similar to morphine, mitragynine mainly acts at mu-opioid receptor which is the main target to treat opioid dependency. Unfortunately, it is believed that the psychoactivity of mitragynine is limiting its use. Compared to morphine, mitragynine contains pyrrolidine ring in its chemical structure, which is suspected as the contributor to the unwanted psychoactive activities seen. Therefore, the aim of this study is to investigate the relationship between pyrrolidine ring and mitragynine’s psychoactive effects. This was done by synthesizing an analogue of mitragynine with epoxy ring to replace pyrrolidine. A tail-flick test (52°C) was conducted in vivo using ICR mice in order to establish the basal analgesic effect for mitragynine mediated by the mu-opioid receptor, with morphine as standard reference. 80 mg of mitragynine was found to be equipotent to 10 mg of morphine (n = 6, P < 0.001). The analogue of mitragynine (that has epoxy ring instead of pyrrolidine ring) was generated by total synthesis using 2-benzofuran-3-yl-ethylamine as the starting material involved three major reaction steps. Each intermediates and reaction products were purified using column chromatography. The analogue has been confirmed using 1H-NMR, 13-C NMR and Mass Spectra. This analogue was then evaluated using the tail-flick test to identify the activity of any mu-opioid receptor of the new analogue. The psychoactivity of this new analogue is currently under evaluation in vivo using an open-field test in ICR mice to observe any behavioural activity related to psychoactive effects. The intensity will be compared to mitragynine

Priming dose of morphine/methamphetamine combination induced stereotyped behavior in morphine/methamphetamine-dependent mice

Concurrent use of methamphetamine and morphine could give a synergistic effect, where it produce greater rewarding effects, and antagonize the side effects of each other, causing it to have a greater potential to be abused. This poly-drug abuse may lead to behavioral sensitization shown by stereotype behavior. The aim of this study is to compare the behavioral effects of priming dose in reinstatement model of morphine- and morphine/methamphetamine-dependent mice. Through conditioned place preference (CPP) test, dependence was successfully established by using 7.5 mg/kg morphine and/or 1 mglkg methamphetamine and then, extinction was achieved by retesting style. The mice were later challenged with priming dose of 2.5 mg/kg morphine and/or 'l mg/kg methamphetamine. Significant preference towards the drug-paired compartment were shown in both morphine- and morphine/methamphetamine-dependent mice (P<0.05 n=12). However, stereotyped behavior was developed only in morphinelmethamphetamine-dependent mice (45-60 mins). This result indicates that concurrent use of morphine and methamphetamine can induce behavioral sensitization in reinstatement modelof morphine/methamphetamine-dependence mice. This finding suggests that stereotype behavior can be one of the parameter to examine drug addiction and craving in animal model and to extend it to human. Thus, this study may lead to potential treatment for poly-drug abuse and the treatment should be designed to inhibit both reinstatement and behavioral sensitization effect of the drug^ Further study on relapse and behavioral sensitization in methamphetamine-dependent mice should be carried out to ascertain any cross-sensitization between morphine and methamphetamine

Distinct role of kappa opioid receptor in attenuating relapse to morphine/methamphetamine (polydrug) dependence

A combination of 0.3 mg/kg buprenorphine and 1.0 mg/kg naltrexone treatment shows a promising result due to its ability to attenuate reinstatement (relapse) in morphine/methamphetamine (polydrug)-dependent mice in a conditioned place preference (CPP) model. This prompted us to identify which opioid receptor that contributes to its anti-relapse activity. Using the same CPP model, 10 mg/kg nor-BNI (a selective kappa opioid receptor [KOR] antagonist) was used to evaluate the involvement of KOR in mediating relapse to polydrug dependence. By applying the immunohistochemistry (IHC) technique, the investigation was extended to the mice brain using KOR antibody (EPR18881), focusing on the brain regions that are abundant in KOR density. The results showed that nor-BNI alone failed to attenuate relapse to polydrug dependence. However, the IHC results proved that the number of KOR significantly increased in the striatum during reinstatement compared to post-conditioning (p <0.05). The KOR was significantly suppressed in the treatment group which strengthens the findings from the previous studies proving that the KOR plays an important role in mediating relapse to polydrug dependence