Sheketha R. Hauser

The Orexin-1 Receptor Antagonist SB-334867 Reduces Alcohol Relapse Drinking, but not Alcohol-Seeking, in Alcohol-Preferring (P) Rats.

Principle:The orexin system has been hypothesized to regulate drug-seeking and drug self-administration behaviors, including ethanol (EtOH) seeking and consumption. However, studies on the effects of orexin receptor antagonists have not been conducted on robust alcohol-relapse behavior. Objectives:This study assessed the effects of the orexin-1 receptor antagonist, SB-334867, on alcohol-seeking behavior and responding for alcohol under relapse conditions. Methods:Adult alcohol-preferring (P) rats self-trained in 2-lever operant chambers to administer 15% EtOH (vol/vol) on a fixed-ratio-5 and water on a fixed-ratio-1 schedule of reinforcement. After 10 weeks, rats underwent extinction training for 7 sessions. Animals were then maintained in their home cages for 2 weeks before being tested for Pavlovian Spontaneous Recovery (PSR; a measure of alcohol seeking) for 4 sessions. Rats were then allowed a week in their home cages before being returned to the operant chamber with access to EtOH and water (relapse). Thirty minutes before the PSR and relapse test sessions, rats received 0, 10, or 20 mg/kg SB-334867. Results:Responses on the EtOH lever during the first PSR test session were ∼70 presses/session (3-fold higher than baseline); SB-334867 did not alter responses on the EtOH lever. Under relapse conditions, P rats increased responding on the EtOH lever from 250 (at baseline) to 350 responses/session; both doses of SD-334867 prevented this increase. Conclusions:The results of this study suggest that activation of orexin-1 receptors is not involved in intrinsically initiated EtOH seeking, but may regulate the consummatory behavior of EtOH consumption.

Nicotine Modulates Alcohol-Seeking and Relapse by Alcohol-Preferring (P) Rats in a Time-Dependent Manner

BACKGROUND Alcohol is frequently co-abused with smoking. In humans, nicotine use can increase alcohol craving and consumption. The objectives of the current study were to assess the acute effects of nicotine on alcohol seeking and relapse at 2 different time points. METHODS Adult female alcohol-preferring (P) rats were trained in 2-lever operant chambers to self-administer 15% ethanol (EtOH) (v/v) and water on a concurrent fixed-ratio 5-fixed-ratio 1 (FR5-FR1) schedule of reinforcement in daily 1-hour sessions. Following 10 weeks of daily 1-hour sessions, rats underwent 7 extinction sessions, followed by 2 weeks in their home cages. Rats were then returned to the operant chambers without EtOH or water being present for 4 sessions (Pavlovian Spontaneous Recovery [PSR]). Rats were then given a week in their home cage before being returned to the operant chambers with access to EtOH and water (relapse). Nicotine (0, 0.1, 0.3, or 1.0 mg/kg) was injected subcutaneously immediately or 4 hours prior to PSR or relapse testing. RESULTS Injections of nicotine immediately prior to testing reduced (5 to 10 responses PSR; 50 to 60 responses relapse), whereas injections of nicotine 4 hours prior to testing increased (up to 150 responses for PSR; up to 400 responses for relapse with 1.0 mg/kg dose) responses on the EtOH lever during PSR and relapse tests. CONCLUSIONS The results of this study demonstrate that acute effects of nicotine on EtOH-seeking and relapse behaviors may be time dependent, with the immediate effects being a result of nicotine possibly acting as a substitute for EtOH, whereas with a delay of 4 hours, priming effects of nicotine alterations in nicotinic receptors, and/or the effects of nicotines metabolites (i.e., cotinine and nornicotine) may enhance the expression of EtOH-seeking and relapse behaviors.

The posterior ventral tegmental area mediates alcohol-seeking behavior in alcohol-preferring rats.

The mesolimbic dopamine (DA) system is involved in the rewarding process of drugs of abuse and is activated during the anticipation of drug availability. However, the neurocircuitry that regulates ethanol (EtOH)-seeking has not been adequately investigated. The objectives of the present study were to determine 1) whether the posterior ventral tegmental area (p-VTA) mediates EtOH-seeking, 2) whether microinjections of EtOH into the p-VTA could stimulate EtOH-seeking, and (3) the involvement of p-VTA DA neurons in EtOH-seeking. Alcohol-preferring rats were trained to self-administer 15% EtOH and water. After 10 weeks, rats underwent extinction training, followed by 2 weeks in their home cages. During the home-cage period, rats were then bilaterally implanted with guide cannulae aimed at the p-VTA or anterior ventral tegmental area (a-VTA). EtOH-seeking was assessed by the Pavlovian spontaneous recovery model. Separate experiments examined the effects of: 1) microinjection of quinpirole into the p-VTA, 2) EtOH microinjected into the p-VTA, 3) coadministration of EtOH and quinpirole into the p-VTA, 4) microinjection of quinpirole into the a-VTA, and 5) microinjection of EtOH into the a-VTA. Quinpirole microinjected into the p-VTA reduced EtOH-seeking. Microinjections of EtOH into the p-VTA increased EtOH-seeking. Pretreatment with both quinpirole and EtOH into the p-VTA reduced EtOH-seeking. Microinjections of quinpirole or EtOH into the a-VTA did not alter EtOH-seeking. Overall, the results suggest that the p-VTA is a neuroanatomical substrate mediating alcohol-seeking behavior and that activation of local DA neurons is involved.

Reduced ethanol consumption by alcohol-preferring (P) rats following pharmacological silencing and deep brain stimulation of the nucleus accumbens shell

OBJECT There is increasing interest in deep brain stimulation (DBS) for the treatment of addiction. Initial testing must be conducted in animals, and the alcohol-preferring (P) rat meets the criteria for an animal model of alcoholism. This study is composed of 2 experiments designed to examine the effects of 1) pharmacological inactivation and 2) DBS of the nucleus accumbens shell (AcbSh) on the consumption of alcohol by P rats. METHODS In the first experiment, the effects of reversible inactivation of the AcbSh were investigated by administering intracranial injections of γ-aminobutyric acid (GABA) agonists. Bilateral microinjections of drug were administered to the AcbSh in P rats (8-10 rats/group), after which the animals were placed in operant chambers containing 2 levers--one used to administer water and the other to administer 15% EtOH--to examine the acquisition and maintenance of oral EtOH self-administration. In the second experiment, a DBS electrode was placed in each P rats left AcbSh. The animals then received 100 or 200 μA (3-4 rats/group) of DBS to examine the effect on daily consumption of oral EtOH in a free-access paradigm. RESULTS In the first experiment, pharmacological silencing of the AcbSh with GABA agonists did not decrease the acquisition of EtOH drinking behavior but did reduce EtOH consumption by 55% in chronically drinking rats. Similarly, in the second experiment, 200 μA of DBS consistently reduced EtOH intake by 47% in chronically drinking rats. The amount of EtOH consumption returned to baseline levels following termination of therapy in both experiments. CONCLUSIONS Pharmacological silencing and DBS of the AcbSh reduced EtOH intake after chronic EtOH use had been established in rodents. The AcbSh is a neuroanatomical substrate for the reinforcing effects of alcohol and may be a target for surgical intervention in cases of alcoholism.

Alcohol-preferring (P) rats are more sensitive than Wistar rats to the reinforcing effects of cocaine self-administered directly into the nucleus accumbens shell

Wistar rats will self-administer cocaine directly into the nucleus accumbens shell (AcbSh), but not into the nucleus accumbens core. In human and animal literature, there is a genetic association between alcoholism and cocaine dependency. The current experiment examined whether selective breeding for high alcohol preference is also associated with greater sensitivity of the AcbSh to the reinforcing properties of cocaine. P and Wistar rats were given cocaine (0, 100, 200, 400, or 800 pmol/100 nl) to self-infuse into the AcbSh. Rats were given cocaine for the first 4 sessions (acquisition), artificial CSF for sessions 5 and 6 (extinction), and cocaine again in session 7 (reinstatement). During acquisition, P rats self-infused 200-800 pmol cocaine (59 infusions/session), whereas Wistar rats only reliably self-infused 800 pmol cocaine (38 infusions/session). Furthermore, P rats received a greater number of cocaine infusions in the 200, 400 and 800 pmol cocaine groups compared to respective Wistar groups during acquisition. Both P and Wistar rats reduced responding on the active lever when aCSF was substituted for cocaine, and reinstated responding in session 7 when cocaine was restored. However, P rats had significantly greater infusions during session 7 compared to session 4 at all concentrations of cocaine tested, whereas Wistar rats only displayed greater infusions during session 7 compared to session 4 at the 400 and 800 pmol cocaine concentrations. The present results suggest that, compared to Wistar rats, the AcbSh of P rats was more sensitive to the reinforcing effects of cocaine. The reinstatement data suggest that the AcbSh of P rats may have become sensitized to the reinforcing effects of cocaine. Overall, the findings from this study support a genetic association between high alcohol preference and greater sensitivity to the reinforcing effects of cocaine.

Ethanol-Associated Changes in Glutamate Reward Neurocircuitry: A Minireview of Clinical and Preclinical Genetic Findings.

Herein, we have reviewed the role of glutamate, the major excitatory neurotransmitter in the brain, in a number of neurochemical, -physiological, and -behavioral processes mediating the development of alcohol dependence. The findings discussed include results from both preclinical as well as neuroimaging and postmortem clinical studies. Expression levels for a number of glutamate-associated genes and/or proteins are modulated by alcohol abuse and dependence. These changes in expression include metabotropic receptors and ionotropic receptor subunits as well as different glutamate transporters. Moreover, these changes in gene expression parallel the pharmacologic manipulation of these same receptors and transporters. Some of these gene expression changes may have predated alcohol abuse and dependence because a number of glutamate-associated polymorphisms are related to a genetic predisposition to develop alcohol dependence. Other glutamate-associated polymorphisms are linked to age at the onset of alcohol-dependence and initial level of response/sensitivity to alcohol. Finally, findings of innate and/or ethanol-induced glutamate-associated gene expression differences/changes observed in a genetic animal model of alcoholism, the P rat, are summarized. Overall, the existing literature indicates that changes in glutamate receptors, transporters, enzymes, and scaffolding proteins are crucial for the development of alcohol dependence and there is a substantial genetic component to these effects. This indicates that continued research into the genetic underpinnings of these glutamate-associated effects will provide important novel molecular targets for treating alcohol abuse and dependence.

Elucidating the biological basis for the reinforcing actions of alcohol in the mesolimbic dopamine system: the role of active metabolites of alcohol

The development of successful pharmacotherapeutics for the treatment of alcoholism is predicated upon understanding the biological action of alcohol. A limitation of the alcohol research field has been examining the effects of alcohol only and ignoring the multiple biological active metabolites of alcohol. The concept that alcohol is a “pro-drug” is not new. Alcohol is readily metabolized to acetaldehyde within the brain. Acetaldehyde is a highly reactive compound that forms a number of condensation products, including salsolinol and iso-salsolinol (acetaldehyde and dopamine). Recent experiments have established that numerous metabolites of alcohol have direct CNS action, and could, in part or whole, mediate the reinforcing actions of alcohol within the mesolimbic dopamine system. The mesolimbic dopamine system originates in the ventral tegmental area (VTA) and projects to forebrain regions that include the nucleus accumbens (Acb) and the medial prefrontal cortex (mPFC) and is thought to be the neurocircuitry governing the rewarding properties of drugs of abuse. Within this neurocircuitry there is convincing evidence that; (1) biologically active metabolites of alcohol can directly or indirectly increase the activity of VTA dopamine neurons, (2) alcohol and alcohol metabolites are reinforcing within the mesolimbic dopamine system, (3) inhibiting the alcohol metabolic pathway inhibits the biological consequences of alcohol exposure, (4) alcohol consumption can be reduced by inhibiting/attenuating the alcohol metabolic pathway in the mesolimbic dopamine system, (5) alcohol metabolites can alter neurochemical levels within the mesolimbic dopamine system, and (6) alcohol interacts with alcohol metabolites to enhance the actions of both compounds. The data indicate that there is a positive relationship between alcohol and alcohol metabolites in regulating the biological consequences of consuming alcohol and the potential of alcohol use escalating to alcoholism.

Synergistic self-administration of ethanol and cocaine directly into the posterior ventral tegmental area: involvement of serotonin-3 receptors.

Ethanol (EtOH) and cocaine are both self-administered into the posterior ventral tegmental area (VTA). Self-administration of either drug is prevented by coadministration of a serotonin (5-HT3) receptor antagonist. Electrophysiological studies indicated that cocaine and EtOH can act synergistically to stimulate VTA dopamine neurons. The current experiment assessed whether cocaine and EtOH would synergistically interact to produce a reinforcing action within the posterior VTA. Adult female Wistar rats were randomly assigned to one of 13 groups. There were three control groups: artificial cerebrospinal fluid (aCSF), a subthreshold EtOH (100 mg%) group, and a subthreshold cocaine (25 pmol/100 nl) group. The other groups self-administered 50 or 75 mg% EtOH containing 6.25, 12.5, or 25 pmol/100 nl cocaine or 100 mg% EtOH containing 3.12, 6.25, 12.5, or 25 pmol/100 nl cocaine. All rats received the assigned infusate for sessions 1 through 4, aCSF alone in sessions 5 and 6, and the original infusate during session 7. The effects of adding a 5-HT3 receptor antagonist [tropisetron, C17H20N2O2 (ICS 205,930) and C17H22N4O.C4H4O4 (LY278-584)] on coadministration of EtOH and cocaine (75 mg% + 12.5 pmol/100 nl) were determined. Rats failed to self-administer aCSF or the subthreshold concentration of EtOH or cocaine. All three concentrations of EtOH (50, 75, and 100 mg%) combined with cocaine (12.5 and 25 pmol/100 nl) supported self-administration. Adding a 5HT3 receptor antagonist attenuated coadministration of EtOH + cocaine. Overall, the data indicate that the reinforcing properties of EtOH and cocaine interacted synergistically within the posterior VTA, and these synergistic effects were mediated, at least in part, by activation of local 5-HT3 receptors.

Repeated exposure of the posterior ventral tegmental area to nicotine increases the sensitivity of local dopamine neurons to the stimulating effects of ethanol

Clinical evidence indicates a frequent co-morbidity of nicotine and alcohol abuse and dependence. The posterior ventral tegmental area (pVTA) appears to support the reinforcing and dopamine-stimulating effects of both drugs. The current study tested the hypothesis that repeated exposure of the pVTA to one drug would increase the sensitivity of local dopamine neurons to the stimulating effects of the other drug. Female Wistar rats received repeated daily microinjections of either 100 μM nicotine or vehicle directly into the pVTA for 7 days. On the 8th day, rats received microinjections of either vehicle or ethanol (100 or 200 mg%) into the pVTA while extracellular dopamine samples were collected from the ipsilateral nucleus accumbens shell (NACsh) with microdialysis. Another experiment tested the effects of challenge microinjections of 200 μM nicotine in the pVTA on extracellular dopamine levels in the NACsh following 7 daily pretreatments with 200 mg% ethanol in the pVTA. Nicotine pretreatments increased the dopamine-stimulating effects of ethanol in the pVTA (100 mg% ethanol: 115% vs 160% of baseline in the vehicle and nicotine groups, respectively, p < 0.05; 200 mg% ethanol: 145% vs 190% of baseline in the vehicle and nicotine groups, respectively, p < 0.05). In contrast, ethanol pretreatments did not alter the stimulating effects of nicotine in the pVTA. The results suggest that repeated exposure of the pVTA to nicotine increased the response of local dopamine neurons to the stimulating effects of ethanol, whereas repeated exposure of the pVTA to ethanol did not alter the responses of pVTA dopamine neurons to nicotine.

CB1 receptors regulate alcohol-seeking behavior and alcohol self-administration of alcohol-preferring (P) rats.

RATIONALE The endogenous cannabinoid (CB) system mediates a number of behaviors associated with drug-seeking and drug self-administration. In this study the effects of CB1 receptor manipulations on operant ethanol (EtOH) responding during EtOH-seeking, EtOH-relapse as well as on-going EtOH self-administration were determined. METHODS Alcohol-preferring (P) rats were trained in 2-lever operant chambers to self-administer 15% EtOH (v/v) and water on a concurrent fixed-ratio 5-fixed-ratio 1 (FR5-FR1) schedule of reinforcement in daily 1-h sessions. After 10 weeks, rats underwent 7 extinction sessions, followed by 2 weeks in their home cages without access to EtOH or operant chambers. Rats were then returned to the operant chambers for testing of EtOH-seeking behavior (no EtOH present) for 4 sessions. After a week in their home cages following the EtOH-seeking test, rats were returned to the operant chambers with access to EtOH and water (relapse). Rats were then maintained in the operant chambers for daily 1-h sessions with access to 15% EtOH and water for several weeks. RESULTS The CB1 receptor antagonist (SR141716A), at doses of 1 and 2 mg/kg, i.p. reduced EtOH-seeking and transiently reduced EtOH self-administration during relapse and maintenance. Conversely, treatment with the CB1 receptor agonist CP 55, 940, at doses of 1 and 10 μg/kg i.p., increased EtOH-seeking and EtOH self-administration during relapse. CONCLUSIONS The results of this study demonstrate that activation of CB1 receptors are involved in regulating EtOH-seeking as well as the reinforcing effects of EtOH under relapse and on-going self-administration conditions.