Ethanol Intake In Alcohol-preferring Rats Research Articles

Effects of orally administered Augmentin on glutamate transporter 1, cystine-glutamate exchanger expression and ethanol intake in alcohol-preferring rats

Alcohol dependence is associated with deficits in glutamate uptake and impairment of glutamate homeostasis in different brain reward regions. Glutamate transporter subtype 1 (GLT-1), cystine-glutamate exchanger (xCT) and glutamate/aspartate transporter (GLAST) are one of the key players in regulating extracellular glutamate concentration in the brain. Parenteral treatment with ceftriaxone, β-lactam antibiotic, has been reported to attenuate ethanol consumption and reinstatement to cocaine-seeking behavior, in part, by restoring the expression of GLT-1 and xCT in mesocorticolimbic brain regions in rats. In this study, we focused to test Augmentin (amoxicillin/clavulanate), which can be administered orally to subjects. Therefore, we examined the effects of orally administered Augmentin on ethanol intake as well as GLT-1, xCT and GLAST expression in male alcohol-preferring (P) rats. We found that orally administered Augmentin significantly attenuated ethanol consumption in P rats as compared to the vehicle-treated group. Importantly, the attenuation in ethanol consumption was associated with a significant upregulation of GLT-1 and xCT expression in nucleus accumbens (NAc) and prefrontal cortex (PFC). There was no effect of orally administered Augmentin on GLAST expression in either NAc or PFC. These findings present strong evidence that oral administration of Augmentin can be used as an alternative to parenteral treatment.

Effects of Amoxicillin and Augmentin on Cystine-Glutamate Exchanger and Glutamate Transporter 1 Isoforms as well as Ethanol Intake in Alcohol-Preferring Rats.

Alcohol dependence is associated with alteration of glutamate transport and glutamate neurotransmission. Glutamate transporter 1 (GLT-1) is a major transporter that regulates the majority of extracellular glutamate concentration, which is also regulated by cystine-glutamate exchanger (xCT). Importantly, we recently reported that amoxicillin and Augmentin (amoxicillin/clavulanate) upreglulated GLT-1 expression in nucleus accumbens (NAc) and prefrontal cortex (PFC) as well as reduced ethanol consumption in male P rats. In this study, we examined the effects of amoxicillin and Augmentin on GLT-1 isoforms (GLT-1a and GLT-1b), xCT, and glutamate/aspartate transporter (GLAST) expression in NAc and PFC as well as ethanol intake in male P rats. We found that both compounds significantly reduced ethanol intake, and increased GLT-1a, GLT-1b, and xCT expression in NAc. However, only Augmentin increased GLT-1a, GLT-1b, and xCT expression in PFC. There were no effects of these compounds on GLAST expression in NAc and PFC. These findings demonstrated that Augmentin and amoxicillin have the potential to upregulate GLT-1 isoforms and xCT expression, and consequently attenuate ethanol dependence.

Effects of ampicillin, cefazolin and cefoperazone treatments on GLT-1 expressions in the mesocorticolimbic system and ethanol intake in alcohol-preferring rats

Chronic ethanol consumption is known to downregulate expression of the major glutamate transporter 1 (GLT-1), which increases extracellular glutamate concentrations in subregions of the mesocorticolimbic reward pathway. While β-lactam antibiotics were initially identified as potent upregulators of GLT-1 expression, only ceftriaxone has been extensively studied in various drug addiction models. Therefore, in this study, adult male alcohol-preferring (P) rats exposed chronically to ethanol were treated with other β-lactam antibiotics, ampicillin, cefazolin or cefoperazone (100mg/kg) once daily for five consecutive days to assess their effects on ethanol consumption. The results demonstrated that each compound significantly reduced ethanol intake compared to the saline-treated control group. Importantly, each compound significantly upregulated both GLT-1 and pAKT expressions in the nucleus accumbens and prefrontal cortex compared to saline-treated control group. In addition, only cefoperazone significantly inhibited hepatic aldehyde dehydrogenase-2 enzyme activity. Moreover, these β-lactams exerted only a transient effect on sucrose drinking, suggesting specificity for chronically inhibiting ethanol reward in adult male P rats. Cerebrospinal fluid concentrations of ampicillin, cefazolin or cefoperazone have been confirmed using high-performance liquid chromatography. These findings demonstrate that multiple β-lactam antibiotics demonstrate efficacy in reducing alcohol consumption and appear to be potential therapeutic compounds for treating alcohol abuse and/or dependence. In addition, these results suggest that pAKT may be an important player in this effect, possibly through increased transcription of GLT-1.

Ceftriaxone Treatment Affects the Levels of GLT1 and ENT1 As Well As Ethanol Intake in Alcohol-Preferring Rats

Studies have demonstrated that deletion of equilibrative nucleoside transporter 1 (ENT1) is associated with reduced glutamate transporter 1 (GLT1) level, and consequently increased ethanol intake. In this study, we measured changes in GLT1 and ENT1 levels in prefrontal cortex (PFC), and nucleus accumbens (NAc) core and shell associated with alcohol drinking in alcohol-preferring (P) rats. We examined, then, whether ceftriaxone (CEF) would affect both GLT1 and ENT1 levels in these brain regions. P rats were given 24-h concurrent access to 15 and 30% ethanol, water, and food for 5 weeks. On Week 6, P rats received 100 mg/kg CEF (i.p.) or a saline vehicle for five consecutive days. Ethanol intake was measured daily for 8 days starting on the first day of injections. We found a significant reduction in daily ethanol intake in CEF-treated group, starting on Day 2 of injections. Western blot for GLT1 and binding assay for ENT1 revealed downregulation of GLT1 level, whereas ENT1 levels were increased in the NAc core and NAc shell, respectively, but not in the PFC in saline vehicle group. Importantly, CEF treatment reversed these effects in both NAc core and shell. These findings provide evidence for potential regulatory effects of CEF on both GLT1 and ENT1 expression in reducing ethanol intake.

Effect of the melanocortin receptor stimulation or inhibition on ethanol intake in alcohol-preferring rats

It has been recently reported that acute intracerebroventricular injection of 1 nmol/rat of the non-selective melanocortin 3 and 4 receptor (MC3/4) agonist MTII reduces ethanol intake in female AA alcohol-preferring rats and alters opioid peptide levels in the ventral tegmental area of rats. To better understand the role of the MC system in the control of ethanol intake, we tested the acute and chronic effects of lateral ventricular (LV) injections of 0.01–1 nmol MTII, of 0.1–1 nmol of the MC3/4R receptor antagonist agouti related peptide (AgRP), and 0.1–0.5 nmol of the MC3/4R receptor antagonist SHU9119 on food, water, and 10% ethanol intake in Marchigian-Sardinian alcohol-preferring (msP) rats, which spontaneously ingest pharmacologically relevant quantities of ethanol both under short and long term access conditions. The data showed that with 2 h/day ethanol access, LV MTII injections reduced intake of food and ethanol intakes. When food, water, and ethanol were available ad libitum and 0.01 nmol MTII was given by daily LV injection, however, ethanol intake was reduced for only the first 2 days, whereas food intake was reduced for all 5 days of treatment. Finally, acute LV injection of neither AgRP nor SHU9119 affected ethanol intake under ad libitum conditions, although both antagonists significantly increased food and water intake. In conclusion, these data fail to support a role for endogenous MC3/4R in the control of spontaneous ethanol intake in the msP rat. MC3/4R agonism, however, reduced ethanol intake in association with reduced food intake, suggesting that MTII might reduce nutrient-related controls of ethanol intake rather than, or in addition to, reward-related controls of ethanol intake.

Lymphocyte DNA alteration by sub-chronic ethanol intake in alcohol-preferring rats

Background: Excessive alcohol consumption has been correlated with a higher susceptibility to infections among humans. Chromosome aberrations and other parameters have been suggested as useful biomarkers in assessing genetic damage due to ethanol intake. Methods: Genetically selected alcohol-preferring rats were given water, 10% ethanol and water or 10% ethanol alone for 3 months as fluid to drink. Food was available ad libitum for the entire period. At the end of the sub-chronic treatment their blood and liver were collected. All blood cells were counted and both lymphocytes and hepatocytes of all three groups were tested with the Comet assay to determine whether any DNA damage had occurred. Results: Only lymphocytes showed DNA damage, with differences among groups. The group that had only ethanol to drink showed greater lymphocyte DNA damage than the ethanol/water and water alone groups. On the other hand, hepatocyte DNA did not show any signs of damage. Conclusions: Ten weeks of sub-chronic ethanol treatment produces small but significant damage to lymphocytes but not to hepatocytes, a result which confirms the observations of previous authors, and extends them even to a strain of rats genetically selected for high ethanol intake.

Blockade of gamma-aminobutyric acid receptors does not modify the inhibiton of ethanol intake induced by Hypericum perforatum in rats.

Recent studies have shown that Hypericum perforatum extracts (HPE) inhibit ethanol intake in alcohol-preferring rats, but their mechanism of action is still unknown. HPE have been shown to bind at gamma-aminobutyric acid (GABA)(A) and GABA(B) receptors, to inhibit GABA reuptake, to evoke GABA release from synaptosomes and to exert an anxiolytic effect that is blocked by the benzodiazepine antagonist flumazenil. Since GABA-ergic mechanisms are known to influence ethanol intake, the present study was aimed at investigating whether they might mediate the effect of a CO2 Hypericum extract (HPCO2) on ethanol intake in genetically selected Marchigian Sardinian alcohol-preferring (msP) rats. The GABA(A) receptor antagonist bicuculline and the GABA(B) receptor antagonists CGP-36742 and phaclofen were tested versus the effect of HPCO2 on ethanol intake. The results of the present study confirm that HPCO2, given by intragastric injection, markedly reduces ethanol intake in msP rats and its effect is behaviourally selective, since the same doses which inhibited ethanol intake did not modify the simultaneous intake of food or water. The GABA(A) receptor antagonist bicuculline, given by intraperitoneal (i.p.) injection at a dose of 2 mg/kg, which effectively antagonizes the effects of GABA(A) receptor agonists, did not modify the effect of HPCO2, 15 or 125 mg/kg. The GABA(B) receptor antagonists CGP-36742, given by i.p. injection at a dose of 100 mg/kg, and phaclofen, given by intracerebroventricular injection at a dose of 25 micro g/rat, did not modify the inhibitory effect on alcohol intake induced by HPCO2, 15 or 125 mg/kg. The same doses of the two GABA(B) receptor antagonists induced a pronounced reduction of the effect of the GABA(B) receptor agonist bacoflen, given by i.p. injection at a dose of 5 mg/kg. These findings suggest that the inhibitory effects of HPE on ethanol intake are not mediated by GABA agonist actions.

Pharmacological characterization of the nociceptin receptor which mediates reduction of alcohol drinking in rats

Chronic intracerebroventricular (ICV) treatment with nociceptin/orphanin FQ (NC), the endogenous ligand for the opioid receptor-like 1 (ORL1) receptor, reduces ethanol intake in alcohol-preferring rats and abolishes the rewarding properties of ethanol in the place conditioning paradigm. To pharmacologically characterize the receptor involved, the present study evaluated the effect on ethanol drinking in genetically selected Marchigian Sardinian alcohol-preferring (msP) rats of ICV injections for 8 days of NC or of the NC analogs NC(1–17)NH 2, NC(1–13)NH 2, NC(1–12)NH 2 and [Nphe 1]NC(1–13)NH 2. In vitro studies indicate that NC, NC(1–17)NH 2, NC(1–13)NH 2 and NC(1–12)NH 2 are agonists, while [Nphe 1]NC(1–13)NH 2 is a selective antagonist at the ORL1 receptor. Freely feeding and drinking rats were offered 10% ethanol 30 min/day at the beginning of the dark phase of the light cycle. NC significantly attenuated ethanol intake at 500 or 1000 ng/rat (210 or 420 pmol/rat). NC(1–17)NH 2, markedly reduced ethanol intake, but its effect was statistically significant at 1000 (420 pmol/rat), not at 500 ng/rat (210 pmol/rat). After the end of treatment ethanol drinking promptly came back to baseline level. Ethanol consumption was also reduced by NC(1–13)NH 2; however, its effect was less potent and pronounced. NC(1–12)NH 2 did not modify ethanol intake at doses up to 4000 ng/rat (2339 pmol/rat). Water and food consumption were not modified. Treatment with [Nphe 1]NC(1–13)NH 2, 66 or 99 μg/rat, did not modify ethanol intake; however, [Nphe 1]NC(1–13)NH 2, 66 μg/rat, given just before 1000 ng/rat of NC(1–17)NH 2, abolished the effect of the agonist. The present results show that the 13 aminoacid N-terminal sequence of NC is essential for the effect on ethanol intake and indicate that [Nphe 1]NC(1–13)NH 2 acts as an antagonist to block the effect of NC. These findings provide further evidence that selective agonists at the ORL-1 receptor attenuate ethanol intake in alcohol-preferring rats and suggest that the NC/ORL1 system may represent an interesting target for treatment of alcohol abuse.

Effects of a methanolic extract and a hyperforin-enriched CO2 extract of Hypericum perforatum on alcohol intake in rats

Hypericum perforatum extracts (HPE) inhibit ethanol intake in rats. Hypericin and hyperforin have been proposed as major active principles of HPE. The present study compared the effect on ethanol intake in alcohol-preferring rats of two Hypericum perforatum extracts: a methanolic extract containing 0.3% hypericin and 3.8% hyperforin (HPE1) and a CO2 extract (HPE2) with 24.33% hyperforin and very low hypericin content. Freely feeding and drinking rats were offered 10% ethanol 2 h/day and HPE were given intragastrically 1 h before access to ethanol. Both extracts dose-dependently reduced ethanol intake, HPE2 being about eight times more potent than HPE1. Food and water intakes were not affected by doses that reduced ethanol intake. HPE2, unlike HPE1, reduced blood-alcohol levels (BAL) at doses of > or = 31.2 mg/kg, whereas the dose of 15.6 mg/kg, which reduced ethanol intake, did not significantly modify BAL; blood-acetaldehyde levels were never increased. As previously observed for HPE1, intracerebroventricular pretreatment with 5,7-dihydroxytryptamine (150 microg/rat) did not affect attenuation of ethanol intake induced by HPE2, but reduced its effect in the forced swimming test (FST). Intraperitoneal pretreatment with the sigma-1 receptor antagonist NE-100 (0.25 mg/kg) did not affect inhibition of ethanol intake induced by HPE1 (250 mg/kg) or HPE2 (125 mg/kg), but abolished the effect of both extracts in the FST. In conclusion, the present results indicate that HPE2 inhibits ethanol intake more potently than HPE1; the higher potency of HPE2 parallels the hyperforin content, suggesting that hyperforin may have an important role in reducing ethanol intake. Moreover, different neurochemical mechanisms are apparently responsible for the reduction of ethanol intake and for the antidepressant-like effect of HPE.

Studies of 5-HT3 functioning in alcohol-dependence, by combined use of behavioral animal models and microdialysis

The development of a new animal model for alcoholism is described. Objective, biologic criteria for an animal model were established which did not conflict with either the criteria for the alcohol dependence syndrome of ICD-10 or the criteria for the alcohol dependence in DSM-IV. The 5-hydroxytryptamine (5-HT) system has long been thought to play a role at several steps in the cycle of alcohol abuse and dependence. Agents that increase serotonergic tone have shown promising results in assisting with recovery from alcoholism, whereas the 5-HT3 antagonist ondansetron was found effective in a clinical trial. Dopamine in the nucleus accumbens is thought to be involved in alcoholism via a ‘‘common reward pathway’’, perhaps as the final common denominater. The role of serotonin as a main regulator of dopamine in the nucleus accumbens is emphasized. Activation of 5-HT3 receptors depolarizes the membrane, causing neurotransmitter release, primarily dopamine in the nucleus accumbens. 5-HT3 antagonists can counteract some of the events in the common reward pathway and ‘‘break’’ some of the effects of ethanol in both initial drinking and during abstinence. Tropisetron reduces ethanol consumption in alcoholpreferring rats and increases it in abstainers. Tropisetron significantly reduced ethanol intake in Alcohol-Preferring Rats, whereas it increased ethanol intake in Low-Alcohol-Preferring Rats. These findings draw attention to the possibility of tropisetron having a more complex action than merely 5-HT3 receptor blockade. The effect of tropisetron applied by re6erse microdialysis into the6entral tegmental area on ethanol consumption in rats. This study examined the effect of tropisetron applied directly into the ventral tegmental area, by reverse microdialysis, using rats selected on the basis of a free-choice model for natural alcohol preference. Although not significant in this study, there was a tendency for tropisetron to attenuate the rats’ preference for alcohol when applied by this route. The effect of alcohol and tropisetron on dopamine le6els in the nucleus accumbens. This study examined the effect of a single dose of ethanol on the dopamine level in the nucleus accumbens and also the combined effect of single doses of tropisetron and

Effect of Pueraria lobata (wild) on ethanol intake of alcohol‐preferring rats

The present study evaluated the effect of intraperitoneal injections of crude extracts of Radix pueraria (RP) on ethanol intake in genetically selected alcohol-preferring rats. Water and food sated rats were offered 10% ethanol intake for 10 h/day, during the dark phase of an inverse light-dark cycle. RP extracts reduced ethanol intake in alcohol-preferring rats. However, the dose of 75 mg/100 g body weight, that produced only minor behavioural alterations and did not modify total fluid intake, elicited only a modest attenuation (about 20%) of ethanol consumption. A more pronounced inhibition of ethanol intake was obtained following 150 and 300 mg/100 g, but these doses induced marked alterations of the gross behaviour in treated rats, which were rather immobile and apparently in a condition of malaise at least during the first hour after drug administration. Moreover, 150 mg/100 g produced a general inhibition of the ingestive behaviour, as shown by the finding that it inhibited water intake in water deprived rats and food deprived rats. Pronounced alterations on the gross behaviour were evident also in deprived rats, which were not offered 10% ethanol, thus indicating that they cannot be accounted for by an effect of RP on ethanol metabolism. © 1998 John Wiley & Sons, Ltd.

Stimulation of tachykinin NK-3 receptors in the nucleus basalis magnocellularis reduces alcohol intake in rats.

Injections in the nucleus basalis magnocellularis (NBM) of the tachykinin (TK) NK-3 receptor agonist [Asp5,6,MePhe8]substance P(5-11), also referred to as amino-senktide (NH2-SENK), markedly reduced alcohol intake in genetically selected alcohol-preferring rats, offered 10% ethanol 2 h/day. The threshold dose in the NBM was 0.5 ng/site, while neither 1 nor 10 ng/rat of NH2-SENK inhibited alcohol intake following administration into the lateral ventricle. Injection of NH2-SENK, 25 ng/site, in the NBM did not modify water or food intake in water deprived rats, providing evidence for the behavioral selectivity of the effect on ethanol intake. The selective TK NK-3 receptor antagonist, R-820, injected in the NBM at the dose of 1000 ng/site 5 min before NH2-SENK 5 ng/site, significantly reduced the effect of NH2-SENK. The selective TK NK-1 receptor agonist [Sar9,Met(O2)11]substance P inhibited alcohol intake following injection in the NBM only at 25 ng/site; but the same dose induced marked grooming and inhibited also water intake in water deprived rats. The present results confirm that TK NK-3, but not NK-1, receptor agonists selectively inhibit ethanol intake in alcohol-preferring rats and suggest that the NBM is a site of action for their effect.

Suppression of alcohol intake after administration of the Chinese herbal medicine, NPI-028, and its derivatives.

The Chinese herbal medicine, NPI-028, has been used for centuries in China to counteract alcohol intoxication. The present study used a number of different experimental conditions to determine whether NPI-028 and its derivatives might selectively influence alcohol intake in rodents that naturally exhibit high alcohol intakes. It was determined that intraperitoneal (i.p.) injections of NPI-028 (0.5, 0.75, and 1.0 g/kg) suppressed alcohol intake by up to 30% in both alcohol-preferring P and Fawn-Hooded (FH) rats during a continuous access schedule. These injections did not significantly affect food or water intakes, nor did the highest dose of NPI-028 (1 g/kg) alter blood ethanol levels after an i.p. injection of 2.5 g/kg of ethanol. In P rats, it was found that NPI-028 was orally active with the dose of 1.5 g/kg having a greater effect on ethanol intake than the 1.0 g/kg dose; once again, food and water intakes were not significantly altered. In FH rats maintained on a limited access schedule (1 hr/day), alcohol intake was completely abolished by 1.5 g/kg of NPI-028. Chronic i.p. administration of NPI-028 (0.75 g/kg) for four consecutive days in FH rats maintained on a continuous access schedule did not lead to any diminution of its alcohol-suppressant effects. Thus, NPI-028 has significant effects on alcohol intake without much effect on water and food intake, and tolerance does not readily develop to these effects. The i.p. administration of a partially purified extract (NPI-031) of NPI-028, obtained by countercurrent chromatography, also dose-dependently suppressed ethanol intake in FH rats, but the highest dose 200 mg/kg) also significantly decreased food intake. Finally, the i.p. administration of puerarin (NPI-31G), an isoflavone isolated from NPI-031 by countercurrent chromatography, significantly reduced ethanol intake in FH rats without affecting food or water intake. Therefore, NPI-028 and one of its pure components, NPI-031G, selectively reduced ethanol intake in alcohol-preferring rats.

The 5-HT4 receptor antagonist, GR113808, reduces ethanol intake in alcohol-preferring rats

The present study evaluated the effect of the selective 5-HT4 receptor antagonist, GR113808, on ethanol intake in alcoholpreferring rats. Rats were offered 10% ethanol 2 h/day. In the first experiment, rats had food and water ad lib and 10% ethanol was offered from 1800 to 2000 h. In the second experiment, food was freely available, 10% ethanol was offered 2 h/ day, from 1800 to 2000 h, and water was offered for 4 h, from 1800 to 2200 h. In both experiments GR113808 was subcutaneously injected at doses of 1, 3, or 10 mg/kg for 4 consecutive days, 5 min before access to ethanol. From the first day of administration, GR113808 significantly reduced the volitional ethanol intake in water sated rats at the three doses tested. In water-deprived rats, it reduced ethanol intake at 3 and 10 mg/kg, without modifying total fluid and food intake. In both experiments the effect of GR113808 remained rather stable during the 4 days of administration. The present findings, showing that the 5-HT4 receptor antagonist, GR113808, selectively reduces ethanol intake in alcohol-preferring rats, suggest that 5-HT4 receptors may play a role in alcohol intake control.

Suppression of ethanol intake in alcohol-preferring rats by prior voluntary saccharin consumption

In a situation offering a free choice between 0.1% saccharin solution and tap water, Fawn Hooded (FH) rats consumed 363.0 ± 33.5 ml/kg/day of saccharin solution. Subsequently those animals drank 3.0 ± 0.4 g/kg of ethanol in a free choice between water and 10% ethanol solution. Control FH rats that did not have access to saccharin consumed 5.0 ±0.5 g/kg/day of ethanol in the same situation (difference between groups was significant: p = 0.006). When control rats were exposed to the choice between 10% ethanol solution and 0.1% saccharin solution for 4 days they consumed 383.7 ± 27.5 ml/ kg/day of saccharin solution and their ethanol intake dropped to 1.2 ± 0.3 g/kg/day. When these rats were returned back to alcohol/water choice and exposure to saccharin was discontinued, their alcohol intake was still reduced (3.7 ± 0.3 g/kg/day for at least 10 consecutive days). Exposure of alcohol-experienced alcohol-preferring P rats with high (6.8 ± 0.5 g/kg/day) and stable alcohol intake to saccharin/water choice for 4 days also resulted in a significant attenuation of their ethanol intake for at least 6 days following saccharin cessation. Thus, voluntary consumption of saccharin can suppress subsequent alcohol intake in both alcohol-naive and alcohol-experienced rats.