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Drugs of abuse induce widespread synaptic adaptations in the mesolimbic dopamine (DA) neurons. Such drug-induced neuroadaptations may constitute an initial cellular mechanism eventually leading to compulsive drug-seeking behavior. To evaluate the impact of GABA B receptors on addiction-related persistent neuroplasticity, we tested the ability of orthosteric agonist baclofen and two positive allosteric modulators (PAMs) of GABA B receptors to suppress neuroadaptations in the ventral tegmental area (VTA) and reward-related behaviors induced by ethanol and cocaine. A novel compound (S)-1-(5-fluoro-2,3-dihydro-1H-inden-2-yl)-4-methyl-6,7,8,9-tetrahydro-[1,2,4]triazolo[4,3-a]quinazolin-5(4H)-one (ORM-27669) was found to be a GABA B PAM of low efficacy as agonist, whereas the reference compound (R,S)-5,7-di-tert-butyl-3-hydroxy-3-trifluoromethyl-3H-benzofuran-2-one (rac-BHFF) had a different allosteric profile being a more potent PAM in the calcium-based assay and an agonist, coupled with potent PAM activity, in the [ 35 S] GTPγS binding assay in rat and human recombinant receptors. Using autoradiography, the high-efficacy rac-BHFF and the low-efficacy ORM-27669 potentiated the effects of baclofen on [ 35 S] GTPγS binding with identical brain regional distribution. Treatment of mice with baclofen, rac-BHFF, or ORM-27669 failed to induce glutamate receptor neuroplasticity in the VTA DA neurons. Pretreatment with rac-BHFF at non-sedative doses effectively reversed both ethanol- and cocaine-induced plasticity and attenuated cocaine i.v. self-administration and ethanol drinking. Pretreatment with ORM-27669 only reversed ethanol-induced neuroplasticity and attenuated ethanol drinking but had no effects on cocaine-induced neuroplasticity or self-administration. These findings encourage further investigation of GABA B receptor PAMs with different efficacies in addiction models to develop novel treatment strategies for drug addiction., (© 2018 Society for the Study of Addiction.)

Opioid antagonist treatments reduce alcohol drinking in rodent models and in alcohol-dependent patients, with variable efficacy across different studies. These treatments may suffer from the development of tolerance and opioid receptor supersensitivity, as suggested by preclinical models showing activation of these processes during and after subchronic high-dose administration of the short-acting opioid antagonist naloxone. In the present study, we compared equipotent low and moderate daily doses of naltrexone and nalmefene, two opioid antagonists in the clinical practice for treatment of alcoholism. The antagonists were given here subcutaneously for 7 days either as daily injections or continuous osmotic minipump-driven infusions to alcohol-preferring AA rats having trained to drink 10% alcohol in a limited access protocol. One day after stopping the antagonist treatment, [ 35 S]GTPγS autoradiography on brain cryostat sections was carried out to examine the coupling of receptors to G protein activation. The results prove the efficacy of repeated injections over infused opioid antagonists in reducing alcohol drinking. Tolerance to the reducing effect on alcohol drinking and to the enhancement of G protein coupling to μ-opioid receptors in various brain regions were consistently detected only after infused antagonists. Supersensitivity of κ-opioid receptors was seen in the ventral and dorsal striatal regions especially by infused nalmefene. Nalmefene showed no clear agonistic activity in rat brain sections or at human recombinant κ-opioid receptors. The findings support the as-needed dosing practice, rather than the standard continual dosing, in the treatment of alcoholism with opioid receptor antagonists., (© 2016 Society for the Study of Addiction.)

Several studies have revealed that acute ethanol inhibits the function of glutamate receptors. Glutamate receptor-mediated synaptic plasticity, such as N-methyl-D-aspartate-dependent long-term potentiation, is also inhibited by ethanol. However, the inhibition seems to be restricted to certain brain areas such as the hippocampus, amygdala and striatum. Ethanol inhibition of glutamate receptors generally requires relatively high concentrations and may therefore explain consequences of severe ethanol intoxication such as impairment of motor performance and memory. Effects of ethanol on glutamate system of developing nervous system may have a role in causing foetal alcohol syndrome. Newly found regulatory proteins of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid AMPA receptors seem to affect ethanol inhibition thus opening new lines of research., (© 2012 The Authors Basic & Clinical Pharmacology & Toxicology © 2012 Nordic Pharmacological Society.)

Rationale: Neuronal histamine has a prominent role in sleep-wake control and body homeostasis, but a number of studies suggest that histamine has also a role in higher brain functions including drug reward., Objective: The present experiments characterized the involvement of histamine and its H3 receptor in ethanol-related behaviors in mice., Materials and Methods: Male histidine decarboxylase knockout (HDC KO) and control mice were used to study the role of histamine in ethanol-induced stimulation of locomotor activity, impairment of motor coordination, and conditioned place preference (CPP). Male C57BL/6Sca mice were used to study the effects of H3 receptor antagonist in the effects of ethanol on locomotor activity., Results: The HDC KO mice displayed a weaker stimulatory response to acute ethanol than the wild-type (WT) mice. No differences between genotypes were found after ethanol administration on accelerating rotarod. The HDC KO mice showed stronger ethanol-induced CPP than the WT mice. Binding of the GABA(A) receptor ligand [(3)H]Ro15-4513 was not markedly changed in HDC KO mouse brain and thus could not explain altered responses in KO mice. Ethanol increased the activity of C57BL/6Sca mice, and H3 receptor antagonist ciproxifan inhibited this stimulation. In CPP paradigm ciproxifan, an H3 receptor inverse agonist potentiated ethanol reward., Conclusions: Histaminergic neurotransmission seems to be necessary for the stimulatory effect of ethanol to occur, whereas lack of histamine leads to changes that enhance the conditioned reward by ethanol. Our findings also suggest a role for histamine H3 receptor in modulation of the ethanol stimulation and reward.

Glutamate receptors are important target molecules of the acute effect of ethanol. We studied ethanol sensitivity of homomeric GluR-D receptors expressed in human embryonic kidney 293 cells and examined whether recently discovered transmembrane alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor regulatory proteins (TARPs) affect ethanol sensitivity. Coexpression of the TARPs, stargazin, and gamma4 increased the time constant (tau-value) of current decay in the presence of agonist, thus slowing the onset of desensitization and increasing the steady-state current. Ethanol produced less inhibition of the peak current than the steady-state current for all types of the GluR-D receptors. In addition, ethanol concentration-dependently accelerated the rate of desensitization, measured as the tau-value of fast decay of peak current. This effect was enhanced with coexpression of TARPs. The recovery from desensitization was slowed down by coexpression of gamma4 but ethanol did not affect this process in any GluR-D combination. The results support the idea that increased desensitization is an important mechanism in the ethanol inhibition of AMPA receptors and indicate that coexpression of TARPs can alter this effect of ethanol.

In rodent models, gamma-aminobutyric acid A (GABAA) receptors with the alpha6 and delta subunits, expressed in the cerebellar and cochlear nucleus granule cells, have been linked to ethanol sensitivity and voluntary ethanol drinking. Here, we review the findings. When considering both in vivo contributions and data on cloned receptors, the evidence for direct participation of the alpha6-containing receptors to increased ethanol sensitivity is poor. The alpha6 subunit-knockout mouse lines do not have any changed sensitivity to ethanol, although these mice do display increased benzodiazepine sensitivity. However, in general the compensations occurring in knockout mice (regardless of which particular gene is knocked out) tend to fog interpretations of drug actions at the systems level. For example, the alpha6 knockout mice have increased TASK-1 channel expression in their cerebellar granule cells, which could influence sensitivity to ethanol in the opposite direction to that obtained with the alpha6 knockouts. Indeed, TASK-1 knockout mice are more impaired than wild types in motor skills when given ethanol; this might explain why GABAA receptor alpha6 knockout mice have unchanged ethanol sensitivities. As an alternative to studying knockout mice, we examined the claimed delta subunit-dependent/gamma2 subunit-independent ethanol/[3H]Ro 15-4513 binding sites on GABAA receptors. We looked at [3H]Ro 15-4513 binding in HEK 293 cell membrane homogenates containing rat recombinant alpha6/4beta3delta receptors and in mouse brain sections. Specific high-affinity [3H]Ro 15-4513 binding could not be detected under any conditions to the recombinant receptors or to the cerebellar sections of gamma2(F77I) knockin mice, nor was this binding to brain sections of wild-type C57BL/6 inhibited by 1-100 mM ethanol. Since ethanol may act on many receptor and channel protein targets in neuronal membranes, we consider the alpha6 (and alpha4) subunit-containing GABAA receptors unlikely to be directly responsible for any major part of ethanol's actions. Therefore, we finish the review by discussing more generally alcohol and GABAA receptors and by suggesting potential future directions for this research.

Brain regional gamma-aminobutyric acid type A (GABAA) receptor subunit mRNA expression was studied in ethanol-preferring AA (Alko, Alcohol) rats after moderate ethanol drinking for up to 2 years of age. In situ hybridization with oligonucleotide probes specific for 13 different subunits was used with coronal cryostat sections of the brains. Selective alterations were observed by ethanol exposure and/or aging in signals for several subunits. Most interestingly, the putative highly ethanol-sensitive alpha4 and beta3 subunit mRNAs were significantly decreased in several brain regions. The age-related alterations in alpha4 subunit expression were parallel to those caused by lifelong ethanol drinking, whereas aging had no significant effect on beta3 subunit expression. The results suggest that prolonged ethanol consumption leading to blood concentrations of about 10 mM may downregulate the mRNA expression of selected GABAA receptor subunits and that aging might have partly similar effects.

Ethanol actions on alpha-amino-3-hydyroxy-5-methyl-4-isoxazolepropionic acid (AMPA)-type glutamate receptors were studied using voltage-clamp recordings from mouse cortical and hippocampal neurons. During whole-cell recordings ethanol (EtOH) inhibited AMPA receptor-mediated currents in a dose-dependent manner at concentrations from 10 to 500 mM. The steady-state component of AMPA-activated current was more sensitive to EtOH than the peak component. To examine the effect of EtOH on a well resolved peak current component, patches were excised from cultured cortical neurons, to which AMPA and EtOH were applied using a piezoelectric solution application system. Under this condition, the peak current was not inhibited significantly by EtOH. To further study possible mechanisms of EtOH inhibition, kainate and AMPA were used to evoke currents in the absence and presence of cyclothiazide. Ethanol inhibition was stronger when receptors were activated by low than high kainate concentrations. Cyclothiazide reduced inhibition by EtOH regardless of the agonist used to activate the receptor. Finally, EtOH inhibition was reduced in a point mutated (L497Y) GluRAi receptor that lacks desensitization. These findings suggest that EtOH inhibits AMPA receptors by stabilizing the desensitized state. Our results can explain some of the variation observed in EtOH inhibition in previous studies, and support the idea that physiologically relevant concentrations of EtOH can have a strong effect on AMPA receptor function.

We studied the effect of a novel delta-opioid receptor antagonist N,N(CH(3))(2)Dmt-Tic-OH (Me(2)-Dmt-Tic-OH) on voluntary ethanol intake in an alcohol-preferring AA (Alko, Alcohol) rat line using a 4-hour limited access paradigm. Acute injections of Me(2)-Dmt-Tic-OH (10 and 30 mg/kg, i.p.) did not reduce 1-hour or 4-hour ethanol intake. Subtype non-selective opioid receptor antagonist naltrexone [0.1 and 0.3 mg/kg, subcutaneously (s.c.)] significantly reduced 1-hour ethanol drinking but had no effect on 4-hour ethanol consumption. Locomotor stimulation induced by the delta-opioid receptor agonist Tyr-D-Pen-Gly-Phe-D-Pen (DPDPE; 15 microg, intracerebroventricularly) was significantly attenuated by Me(2)-Dmt-Tic-OH (10 and 30 mg/kg, i.p.), which confirmed its efficacy as a delta-opioid receptor antagonist in rat brain. Our results support the idea that delta-opioid receptors do not mediate alcohol reward in AA rats.

The effect of ethanol on motor performance in humans is well established but how neural mechanisms are affected by ethanol action remains largely unknown. To investigate whether the brain histaminergic system is important in it, we used a genetic model consisting of rat lines selectively outbred for differential ethanol sensitivity. Ethanol-sensitive rats had lower levels of brain histamine and lower densities of histamine-immunoreactive fibers than ethanol-insensitive rats, although both rat lines showed no changes in histamine synthesizing neurons. Lowering the high brain histamine content of the ethanol-insensitive rats with alpha-fluoromethylhistidine before ethanol administration increased their ethanol sensitivity in a behavioral motor function test. Higher H3 receptor ligand binding and histamine-induced G-protein activation was detected in several brain regions of ethanol-naive ethanol-sensitive rats. Brain histamine levels and possibly signaling via H3 receptors may thus correlate with genetic differences in ethanol-induced motor impairment.

Background: Psychedelics, like lysergic acid diethylamide (LSD), are again being studied as potential therapies for many neuropsychiatric disorders, including addictions. At the same time, the acute effects of psychedelics on rewarding behaviours have been scarcely studied. Aims: The current study aimed to clarify if LSD decreases binge-like ethanol drinking in mice, and whether the observed acute effects on ethanol consumption are generalizable to a natural reinforcer, sucrose, and if the effects resulted from aversive or reward-attenuating effects caused by LSD. Methods: The effects of acute LSD were examined using 2-bottle choice intermittent ethanol (20%) and sucrose drinking (10%), discrete-trial current-intensity threshold method of intracranial self-stimulation and short-term feeding behaviour assay in C57BL/6 male mice. Results: The results showed that acute 0.1 mg/kg, but not 0.05 mg/kg, dose (i.p.) of LSD reduced 2-h intermittent ethanol drinking transiently without any prolonged effects. No effects were seen in intermittent 2-h sucrose drinking. The tested LSD doses had neither effect on the intracranial self-stimulation current-intensity thresholds, nor did LSD affect the threshold-lowering, or rewarding, effects of simultaneous amphetamine treatment. Furthermore, LSD had small, acute diminishing effects on 2-h food and water intake. Conclusions: Based on these results, LSD decreases binge-like ethanol drinking in mice, but only acutely. This effect is not likely to stem from reward-attenuating effects but could be in part due to reduced consummatory behaviour. [ABSTRACT FROM AUTHOR]

In the brain, extrasynaptically expressed ionotropic, δ subunit-containing γ-aminobutyric acid A-type receptors (δ-GABAARs) have been implicated in drug effects at both neuronal and behavioral levels. These alterations are supposed to be caused via drug-induced modulation of receptor ionophores affecting chloride ion-mediated inhibitory tonic currents. Often, a transgenic mouse model genetically lacking the δ-GABAARs (δ-KO) has been used to study the roles of δ-GABAARs in brain functions, because a specific antagonist of the δ-GABAARs is still lacking. We have previously observed with these δ-KO mice that activation of δ-GABAARs is needed for morphine-induced conditioning of place preference, and others have suggested that δ-GABAARs act as targets selectively for low doses of ethanol. Furthermore, activation of these receptors via drug-mediated agonism induces a robust increase in the slow-wave frequency bands of electroencephalography (EEG). Here, we tested δ-KO mice (compared to littermate wild-type controls) for the pharmaco-EEG responses of a broad spectrum of pharmacologically different drug classes, including alcohol, opioids, stimulants, and psychedelics. Gaboxadol (THIP), a known superagonist of δ-GABAARs, was included as the positive control, and as expected, δ-KO mice produced a blunted pharmaco-EEG response to 6 mg/kg THIP. Pharmaco-EEGs showed notable differences between treatments but also differences between δ-KO mice and their wild-type littermates. Interestingly mephedrone (4-MMC, 5 mg/kg), an amphetamine-like stimulant, had reduced effects in the δ-KO mice. The responses to ethanol (1 g/kg), LSD (0.2 mg/kg), and morphine (20 mg/kg) were similar in δ-KO and wild-type mice. Since stimulants are not known to act on δ-GABAARs, our findings on pharmaco-EEG effects of 4-MMC suggest that δ-GABAARs are involved in the secondary indirect regulation of the brain rhythms after 4-MMC. [ABSTRACT FROM AUTHOR]

Drugs of abuse induce widespread synaptic adaptations in the mesolimbic dopamine (DA) neurons. Such drug-induced neuroadaptations may constitute an initial cellular mechanism eventually leading to compulsive drug-seeking behavior. To evaluate the impact of GABAB receptors on addiction-related persistent neuroplasticity, we tested the ability of orthosteric agonist baclofen and two positive allosteric modulators (PAMs) of GABAB receptors to suppress neuroadaptations in the ventral tegmental area (VTA) and reward-related behaviors induced by ethanol and cocaine. A novel compound (S)-1-(5-fluoro-2,3-dihydro-1H-inden-2-yl)-4-methyl-6,7,8,9-tetrahydro-[1,2,4]triazolo[4,3-a]quinazolin-5(4H)-one (ORM-27669) was found to be a GABAB PAM of low efficacy as agonist, whereas the reference compound (R,S)-5,7-di-tert-butyl-3-hydroxy-3-trifluoromethyl-3H-benzofuran-2-one (rac-BHFF) had a different allosteric profile being a more potent PAM in the calcium-based assay and an agonist, coupled with potent PAM activity, in the [35 S] GTPγS binding assay in rat and human recombinant receptors. Using autoradiography, the high-efficacy rac-BHFF and the low-efficacy ORM-27669 potentiated the effects of baclofen on [35 S] GTPγS binding with identical brain regional distribution. Treatment of mice with baclofen, rac-BHFF, or ORM-27669 failed to induce glutamate receptor neuroplasticity in the VTA DA neurons. Pretreatment with rac-BHFF at non-sedative doses effectively reversed both ethanol- and cocaine-induced plasticity and attenuated cocaine i.v. self-administration and ethanol drinking. Pretreatment with ORM-27669 only reversed ethanol-induced neuroplasticity and attenuated ethanol drinking but had no effects on cocaine-induced neuroplasticity or self-administration. These findings encourage further investigation of GABAB receptor PAMs with different efficacies in addiction models to develop novel treatment strategies for drug addiction. [ABSTRACT FROM AUTHOR]

Glutamate is the main excitatory transmitter in the human brain. Drugs that affect the glutamatergic signaling will alter neuronal excitability. Ethanol inhibits glutamate receptors. We examined the expression level of glutamate receptor subunit mRNAs in human post-mortem samples from alcoholics and compared the results to brain samples from control subjects. RNA from hippocampal dentate gyrus (HP-DG), orbitofrontal cortex (OFC), and dorso-lateral prefrontal cortex (DL-PFC) samples from 21 controls and 19 individuals with chronic alcohol dependence were included in the study. Total RNA was assayed using quantitative RT-PCR. Out of the 16 glutamate receptor subunits, mRNAs encoding two AMPA [2-amino-3-(3-hydroxy-5-methyl-isoxazol-4-yl)propanoic acid] receptor subunits GluA2 and GluA3; three kainate receptor subunits GluK2, GluK3 and GluK5 and five NMDA (N-methyl-D-aspartate) receptor subunits GluN1, GluN2A, GluN2C, GluN2D, and GluN3A were significantly increased in the HP-DG region in alcoholics. In the OFC, mRNA encoding the NMDA receptor subunit GluN3A was increased, whereas in the DL-PFC, no differences in mRNA levels were observed. Our laboratory has previously shown that the expression of genes encoding inhibitory GABA-A receptors is altered in the HP-DG and OFC of alcoholics (Jin et al., 2011). Whether the changes in one neurotransmitter system drives changes in the other or if they change independently is currently not known. The results demonstrate that excessive long-term alcohol consumption is associated with altered expression of genes encoding glutamate receptors in a brain region-specific manner. It is an intriguing possibility that genetic predisposition to alcoholism may contribute to these gene expression changes. [ABSTRACT FROM AUTHOR]

Mephedrone (4-MMC), despite its illegal status, is still a widely used psychoactive substance. Its effects closely mimic those of the classical stimulant drug methamphetamine (METH). Recent research suggests that unlike METH, 4-MMC is not neurotoxic on its own. However, the neurotoxic effects of 4-MMC may be precipitated under certain circumstances, such as administration at high ambient temperatures. Common use of 4-MMC in conjunction with alcohol raises the question whether this co-consumption could also precipitate neurotoxicity. A total of six groups of adolescent rats were treated twice daily for four consecutive days with vehicle, METH (5 mg/kg) or 4-MMC (30 mg/kg), with or without ethanol (1.5 g/kg). To investigate persistent delayed effects of the administrations at two weeks after the final treatments, manganese-enhanced magnetic resonance imaging brain scans were performed. Following the scans, brains were collected for Golgi staining and spine analysis. 4-MMC alone had only subtle effects on neuronal activity. When administered with ethanol, it produced a widespread pattern of deactivation, similar to what was seen with METH-treated rats. These effects were most profound in brain regions which are known to have high dopamine and serotonin activities including hippocampus, nucleus accumbens and caudate-putamen. In the regions showing the strongest activation changes, no morphological changes were observed in spine analysis. By itself 4-MMC showed few long-term effects. However, when co-administered with ethanol, the apparent functional adaptations were profound and comparable to those of neurotoxic METH. [ABSTRACT FROM AUTHOR]

Graphical abstract Abstract The dopamine D 3 receptor (D 3 R), in the nucleus accumbens (NAc), plays an important role in alcohol reward mechanisms. The major neuronal type within the NAc is the GABAergic medium spiny neuron (MSN), whose activity is regulated by dopaminergic inputs. We previously reported that genetic deletion or pharmacological blockade of D 3 R increases GABA A α6 subunit in the ventral striatum. Here we tested the hypothesis that D 3 R-dependent changes in GABA A α6 subunit in the NAc affect voluntary alcohol intake, by influencing the inhibitory transmission of MSNs. We performed in vivo and ex vivo experiments in D 3 R knockout (D 3 R −/−) mice and wild type littermates (D 3 R +/+). Ro 15-4513, a high affinity α6-GABA A ligand was used to study α6 activity. At baseline, NAc α6 expression was negligible in D 3 R+/+, whereas it was robust in D 3 R−/−; other relevant GABA A subunits were not changed. In situ hybridization and qPCR confirmed α6 subunit mRNA expression especially in the NAc. In the drinking-in-the-dark paradigm, systemic administration of Ro 15-4513 inhibited alcohol intake in D 3 R+/+, but increased it in D 3 R−/−; this was confirmed by intra-NAc administration of Ro 15-4513 and furosemide, a selective α6-GABA A antagonist. Whole-cell patch-clamp showed peak amplitudes of miniature inhibitory postsynaptic currents in NAc medium spiny neurons higher in D 3 R−/− compared to D 3 R+/+; Ro 15-4513 reduced the peak amplitude in the NAc of D 3 R−/−, but not in D 3 R+/+. We conclude that D 3 R-dependent enhanced expression of α6 GABA A subunit inhibits voluntary alcohol intake by increasing GABA inhibition in the NAc. [ABSTRACT FROM AUTHOR]