Rewarding Effect In Mice Research Articles

Structure-Activity Relationship of N-Ethyl-Hexedrone Analogues: Role of the α-Carbon Side-Chain Length in the Mechanism of Action, Cytotoxicity, and Behavioral Effects in Mice.

Synthetic cathinones are β-keto amphetamine derivatives whose appearance has increased dramatically in the past decades. N-Ethyl substituted cathinones have been proven to potently inhibit dopamine (DA) uptake and induce psychostimulant and rewarding effects in mice. However, little is known about the influence of the alpha-carbon side-chain length of N-ethyl cathinones on their pharmacological and toxicological effects. Thus, the aim of this study was to synthesize and investigate the in vitro and in vivo effects of five N-ethyl substituted cathinones: N-ethyl-cathinone (NEC), N-ethyl-buphedrone (NEB), N-ethyl-pentedrone, N-ethyl-hexedrone (NEH), and N-ethyl-heptedrone. HEK293 cells expressing the human DA or serotonin transporter (hDAT and hSERT) were used for uptake inhibition and binding assays. PC12 cells were used for the cytotoxicity assays. Swiss CD-1 mice were used to study the in vivo psychostimulant, anxiogenic, and rewarding properties. Our results show that all tested cathinones are able to inhibit DA uptake and are DAT-selective. The potency of DA uptake inhibitors increases with the elongation of the aliphatic side chain from methyl to propyl and decreases when increasing from butyl to pentyl, which correlates with an inverted U-shape psychostimulant response in mice at the medium dose tested. On the other hand, an increase in the α-carbon side-chain length correlates with an increase in the cytotoxic properties in PC12 cells, probably due to better membrane penetration. Moreover, all the cathinones tested have shown higher cytotoxicity than methamphetamine. Finally, our study not only demonstrated the rewarding properties of NEC and NEB but also the anxiety-like behavior induced at high doses by all the cathinones tested.

Role of amino terminal substitutions in the pharmacological, rewarding and psychostimulant profiles of novel synthetic cathinones

The emergence of new synthetic cathinones continues to be a matter of public health concern. In fact, they are quickly replaced by new structurally related alternatives. The main goal of the present study was to characterize the pharmacological profile, the psychostimulant and rewarding properties of novel cathinones (pentedrone, N-ethyl-pentedrone, α-PVP, N,N-diethyl-pentedrone and α-PpVP) which only differs in their amino terminal substitution.Rat synaptosomes were used for [3H]dopamine uptake experiments. HEK293 transfected cells (hDAT, hSERT, hOCT; human dopamine, serotonin and organic cation transporter) were also used for [3H]monoamine uptake and transporter binding assays. Molecular docking was used to investigate the effect of the amino substitutions on the biological activity. Hyperlocomotion and conditioned place preference paradigm were used in order to study the psychostimulant and rewarding effects in mice.All compounds tested are potent inhibitors of DAT with very low affinity for SERT, hOCT-2 and -3, and their potency for inhibiting DAT increased when the amino-substituent expanded from a methyl to either an ethyl-, a pyrrolidine- or a piperidine-ring. Regarding the in vivo results, all the compounds induced an increase in locomotor activity and possess rewarding properties. Results also showed a significant correlation between predicted binding affinities by molecular docking and affinity constants (Ki) for hDAT as well as the cLogP of their amino-substituent with their hDAT/hSERT ratios.Our study demonstrates the role of the amino-substituent in the pharmacological profile of novel synthetic cathinones as well as their potency inhibiting DA uptake and ability to induce psychostimulant and rewarding effects in mice.

The antibiotic minocycline prevents methamphetamine-induced rewarding effects in mice

Repeated use of methamphetamine (METH) causes dependence in humans, and to date, there are no effective medication treatments for METH addiction. We previously reported that the antibiotic minocycline attenuated behavioral abnormalities (hyperactivity and behavioral sensitization) and dopaminergic neurotoxicity in mice and monkeys, after the administration of METH. In this study, we examined the effect of minocycline on METH-induced rewarding effects in mice using the conditioned place preference (CPP) paradigm. Minocycline (40mg/kg, IP) significantly attenuated METH (1.0mg/kg, SC)-induced place preference in mice. In vivo microdialysis experiments using free-moving mice, showed that minocycline (40mg/kg, IP) significantly attenuated the increased extracellular dopamine (DA) levels within the nucleus accumbens, typically seen after the administration of METH (1.0mg/kg, SC). These findings suggest that minocycline may block METH-induced rewarding effects by down regulating extracellular DA levels in the nucleus accumbens of mice. This would make minocycline a potential therapeutic drug for the treatment of METH induced disorders.

Lack of Association Between Genetic Polymorphisms of ARRB2 and Alcohol Dependence in a Caucasian Population

Beta-arrestin 2 ( ARRB2 ) is expressed in the brain and it is involved in μ-opiate and dopamine D2 receptor signaling (Lefkowitz and Shenoy, 2005). It has recently been shown that the ARRB2 genotype may contribute to the inter-individual variability in the response to methadone maintenance treatment (Oneda et al ., 2010). In addition, ARRB2 modulates the acute response to ethanol and mediates its rewarding effects in mice (Bjork et al ., 2008). To investigate the significance of this finding in human, genomic DNA was collected from 45 males and 28 females of Caucasian origin, 18 years old or older, who met the criteria for the diagnosis of alcohol dependence (Diagnostic and Statistical Manual of Mental Disorders, DSM-IV, American Psychiatric Association, 1994). Potential study subjects were excluded if they had …

Inhibitory effect of berberine on the motivational effects of ethanol in mice

It is believed that drug-induced rewarding effects play an important role in the development of substance dependence. Recently, berberine was reported to inhibit the rewarding effects of drugs of abuse such as cocaine, morphine, and nicotine. Berberine is also demonstrated to modulate the activity of several neurotransmitter systems like, dopamine, nitric oxide, serotonin, and NMDA, which are implicated in rewarding effects of ethanol. Hence, we hypothesized that berberine may modulate the ethanol-induced rewarding effects. Therefore, we studied the effect of berberine on locomotor sensitization, conditioned place preference (CPP), and ethanol drinking preference in mice. The results revealed that acute administration of berberine (2.5, 5, and 10 mg/kg, i.p.) dose dependently reduced locomotor stimulant effect of acute ethanol and expression of sensitization to locomotor stimulant effect of ethanol. Further, pretreatment with berberine (2.5, 5, and 10 mg/kg, i.p.) prior to each dose of ethanol, blocked the development as well as expression of sensitization to locomotor stimulant effect of ethanol. In another set of experiment, treatment with berberine (5 and 10 mg/kg, i.p.) reduced the induction and expression of ethanol-induced CPP in mice. In addition, berberine in these doses also reduced preference to ethanol drinking over water, but did not alter the general reward. In conclusion, the results of the present study revealed that berberine attenuates ethanol-induced rewarding effects in mice and that could be attributed to its neuro-modulatory action.

A Proteomics Approach for Identification of Single Strand DNA-binding Proteins Involved in Transcriptional Regulation of Mouse μ Opioid Receptor Gene

The pharmacological actions of morphine and morphine-like drugs such as heroin are mediated primarily through the μ opioid receptor. Previously a single strand DNA element of the mouse μ opioid receptor gene (Oprm1) proximal promoter was found to be important for regulating Oprm1 in neuronal cells. To identify proteins binding to the single strand DNA element as potential regulators for Oprm1, affinity column chromatography with the single strand DNA element was performed using neuroblastoma NS20Y cells followed by two-dimensional gel electrophoresis and MALDI-TOF mass spectrometry. We identified five poly(C)-binding proteins: heterogeneous nuclear ribonucleoprotein (hnRNP) K, α-complex proteins (αCP) αCP1, αCP2, αCP2-KL, and αCP3. Binding of these proteins to the single strand DNA element of Oprm1 was sequence-specific as confirmed by supershift assays. In cotransfection studies, hnRNP K, αCP1, αCP2, and αCP2-KL activated the Oprm1 promoter activity, whereas αCP3 acted as a repressor. Ectopic expression of hnRNP K, αCP1, αCP2, and αCP2-KL also led to activation of the endogenous Oprm1 transcripts, and αCP3 repressed endogenous Oprm1 transcripts. We demonstrate novel roles as transcriptional regulators in Oprm1 regulation for hnRNP K and αCP binding to the single strand DNA element.

MGM-9 [( E)-methyl 2-(3-ethyl-7a,12a-(epoxyethanoxy)-9-fluoro-1,2,3,4,6,7,12,12b-octahydro-8-methoxyindolo[2,3- a]quinolizin-2-yl)-3-methoxyacrylate], a derivative of the indole alkaloid mitragynine: A novel dual-acting μ- and κ-opioid agonist with potent antinociceptive and weak rewarding effects in mice

Mitragynine is a major indole alkaloid isolated from the Thai medicinal plant Mitragyna speciosa that has opium-like properties, although its chemical structure is quite different from that of morphine. We attempted to develop novel analgesics derived from mitragynine, and thus synthesized the ethylene glycol-bridged and C10-fluorinated derivative of mitragynine, MGM-9 [( E)-methyl 2-(3-ethyl-7a,12a-(epoxyethanoxy)-9-fluoro-1,2,3,4,6,7,12,12b-octahydro-8-methoxyindolo[2,3- a]quinolizin-2-yl)-3-methoxyacrylate]. We hypothesized that a dual-acting μ- and κ-opioid agonist could produce potent antinociceptive effects with fewer rewarding effects compared with μ agonists. In this study, MGM-9 exhibited high affinity for μ- and κ-opioid receptors with Ki values of 7.3 and 18 nM, respectively. MGM-9 showed a potent opioid agonistic effect, and its effects were meditated by μ- and κ-opioid receptor mechanisms in in vitro assays. Subcutaneous and oral administration of MGM-9 produced potent antinociceptive effects in mouse tail-flick, hot-plate, and writhing tests. When administered orally, the antinociceptive effect of MGM-9 was seven to 22 times more potent than that of morphine. The antinociceptive effects of MGM-9 were mediated by both μ- and κ-opioid receptors. Subcutaneous administration of MGM-9 twice daily for 5 days led to antinociceptive tolerance. In the gastrointestinal transit study, MGM-9 inhibited gastrointestinal transit, but its effect was weaker than that of morphine at equi-antinociceptive doses. Furthermore, MGM-9 induced less hyperlocomotion and fewer rewarding effects than morphine. The rewarding effect of MGM-9 was blocked by a μ antagonist and enhanced by a κ antagonist. Taken together, the results suggest that MGM-9 is a promising novel analgesic that has a stronger antinociceptive effect and weaker adverse effects than morphine.

Implication of endogenous β-endorphin in the inhibition of the morphine-induced rewarding effect by the direct activation of spinal protein kinase C in mice

It has often been proposed that opioid addiction does not arise as a consequence of opioid treatment for pain. Recently, we demonstrated that activated protein kinase C (PKC) in the spinal cord associated with chronic pain-like hyperalgesia suppressed the morphine-induced rewarding effect in mice. In the present study, we investigated whether a gene deletion for an endogenous mu-opioid peptide beta-endorphin could affect pain-like behavior and the suppression of the morphine-induced rewarding effect by the direct activation of PKC in the spinal cord. We found that activation of spinal PKC by intrathecal (i.t.) treatment with phorbol 12,13-dibutyrate (PDBu), a specific PKC activator, caused thermal hyperalgesia, pain-like behaviors and suppression of the morphine-induced rewarding effect. This suppression of morphine reward was eliminated in mice that lacked beta-endorphin. In contrast, thermal hyperalgesia and pain-like behaviors were not affected in beta-endorphin knockout mice. These results suggest that the activation of PKC in the spinal cord may play an essential role in the suppression of the morphine-induced rewarding effect in mice with neuropathic pain through the constant release of beta-endorphin.

Methamphetamine Reward in Mice as Assessed by Conditioned Place Preference Test with Supermex® Sensors: Effect of Subchronic Clorgyline Pretreatment

Recent studies in our laboratory have shown that methamphetamine (METH)-induced hyperlocomotion and behavioral sensitization in mice were inhibited by clorgyline, an irreversible monoamine oxidase inhibitor. In this study, the effect of clorgyline pretreatment on METH-induced rewarding effect was assessed by a conditioned place preference (CPP) test, using an apparatus developed with Supermex sensors (infrared pyroelectric sensors). Although intact male ICR mice showed significant CPP for METH (0.5 mg/kg, i.p.), pretreatment with subchronic clorgyline (0.1 and 10 mg/kg, s.c.) did not affect the magnitude of CPP. At a dose of 1 mg/kg, pretreatment of the mice with clorgyline showed a similar CPP index in both saline/saline and METH/saline pairing groups. During the conditioning session, the mice did not express behavioral sensitization to METH. Pretreatment with clorgyline (0.1, 1, and 10 mg/kg) decreased striatal apparent monoamine turnover in a dose-dependent manner. These results indicated that clorgyline pretreatment (0.1 and 10 mg/kg) did not influence the METH-induced rewarding effect in mice, although pretreatment of the mice with clorgyline at a dose of 1 mg/kg appeared to influence the CPP for METH.

Role of extracellular signal-regulated kinase in the ventral tegmental area in the suppression of the morphine-induced rewarding effect in mice with sciatic nerve ligation.

We recently reported that micro-opioid receptor agonist morphine failed to induce its rewarding effects in rodents with sciatic nerve injury. In the present study, we investigated whether a state of neuropathic pain induced by sciatic nerve ligation could change the activities of the extracellular signal-regulated kinase (ERK) and p38 in the mouse lower midbrain area including the ventral tegmental area (VTA), and these changes could directly affect the development of the morphine-induced rewarding effect in mice. The sciatic nerve ligation caused a long-lasting and profound thermal hyperalgesia. A dose-dependent place preference induced by s.c. administration of morphine was observed in sham-operated mice, but not in sciatic nerve-ligated mice. We found here for the first time that nerve injury produces a sustained and significant reduction in protein levels of phosphorylated-ERK and -p38 in cytosolic preparations of the mouse lower midbrain. The inhibition of ERK activity by i.c.v. pre-treatment with either PD98059 or U0126 impaired the morphine-induced place preference. In contrast, i.c.v. treatment with a specific inhibitor of p38, SB203580, did not interfere with the morphine-induced rewarding effect. Immunohistochemical study showed a drastic reduction in phosphorylated-ERK immunoreactivity within tyrosine hydroxylase-positive cells of the VTA. These results suggest that a sustained reduction in the ERK-dependent signalling pathway in dopamine cells of the VTA may be implicated in the suppression of the morphine-induced rewarding effect under neuropathic pain.

Evaluation of rewarding effect of toluene by the conditioned place preference procedure in mice

Toluene and many toluene-containing products are abused via inhalation. Previous investigations have used the place preference paradigm to evaluate the rewarding effects of commonly abused drugs such as morphine, cocaine, and amphetamine. A conditioning paradigm of toluene inhalation was developed in order to estimate the rewarding effect in mice. Conditioning sessions (five for toluene, five for air) were conducted twice daily for 5 days using a newly developed airtight inhalation shuttlebox (15×30×15 cm: w×l×h), which was divided into two compartments of equal size. One compartment was white with a textured floor, and the other was black with a smooth floor. All conditioning sessions were 20 min in duration, with a minimum of 7 h between sessions. Test sessions were carried out 1 day after the final training session with mice in a drug-free state. The time spent in each compartment during a 20-min session was measured using a digital video camera. Exposure to toluene vapors (700–3200 ppm) produced a significant conditioned place preference in mice. These results suggest that the conditioned place preference procedure using the newly developed airtight inhalation shuttlebox constitutes an important tool for studying the rewarding effect of abused solvents.

Role of Cannabinoid CB<SUB>1</SUB> Receptor in Morphine Rewarding Effects in Mice

It has been reported that, as well as morphine, the cannabinoid CB1 receptor agonist WIN 55,212-2 failed to induce intravenous self- administration in mutant CB1 receptor knockout (CB1(-/-)) mice but not in the corresponding wild type (CB1(+/+)) mice. To verify whether this functional interaction responsible for opioid rewarding effects was specific or could also be extended to other drugs of abuse, we have evaluated the ability of cocaine, amphetamine and nicotine to induce intravenous self- administration in both CB1(-/-) and CB1(+/+) mice. The results showed that, contrary to morphine, the other drugs of abuse were intravenously self- administered to the same extent by both wild type and CB1(-/-) mice. This points to a specific role of the CB1 receptor in the opioid motivational and rewarding properties. In addition, since mesolimbic dopamine transmission is known to have a pivotal role in reward mediation, the effect of morphine on limbic dopamine release in CB1(-/-) and CB1(+/+) mice has been investigated and compared with the effect of cocaine. Morphine did not modify dopamine release in the nucleus accumbens of CB1(-/-) mice whereas it dose- dependently stimulated dopamine release in the corresponding CB1(+/+) mice. In contrast, cocaine increased dopamine release in both strains of mice, showing that its effect on dopamine transmission was not linked to the cannabinoid system. Taken together, our results clearly show that the CB1 receptor is essential for the expression of the behavioural and biochemical effects of morphine. This extends previous observations on a functional specific interaction between endogenous cannabinoid and opiate systems in the central mechanisms of reward.

Absence of opiate rewarding effects in mice lacking dopamine D2 receptors.

Dopamine receptors have been implicated in the behavioural response to drugs of abuse. These responses are mediated particularly by the mesolimbic dopaminergic pathway arising in the ventral tegmental area and projecting to the limbic system. The rewarding properties of opiates and the somatic expression of morphine abstinence have been related to changes in mesolimbic dopaminergic activity that could constitute the neural substrate for opioid addiction. These adaptive responses to repeated morphine administration have been investigated in mice with a genetic disruption of the dopaminergic D2 receptors. Although the behavioural expression of morphine withdrawal was unchanged in these mice, a total suppression of morphine rewarding properties was observed in a place-preference test. This effect is specific to the drug, as mice lacking D2 receptors behaved the same as wild-type mice when food is used as reward. We conclude that the D2 receptor plays a crucial role in the motivational component of drug addiction.