Role of hypothermia in ethanol-induced conditioned taste aversion

Two experiments examined the effect of ambient temperature during ethanol exposure on development of conditioned taste aversion to saccharin. In both studies, male albino rats receiving saccharin-ethanol (1.5 g/kg, IP) pairings followed by 6-h exposure to a 32° C environment developed a weaker saccharin aversion than did rats experiencing ethanol at room temperature. Exposure to the warm environment reduced ethanol-induced hypothermia, but enhanced ethanol's motor-impairing effect. The influence of ambient temperature on ethanol-induced taste aversion may be due to changes in body temperature, neural sensitivity, or elimination rate. Although alternative accounts cannot be entirely dismissed, this outcome suggests that ethanol-induced hypothermia plays a role in determining strength of conditioned taste aversion and thus may be involved in the regulation of oral ethanol intake in rats. Offprint requests to: C.L. Cunningham     

Role of acetaldehyde in ethanol-induced conditioned taste aversion in rats

Rationale. In spite of many recent studies on the effects of acetaldehyde, it is still unclear whether acetaldehyde mediates the reinforcing and/or aversive effects of ethanol. Objectives. The present study reexamined the role of acetaldehyde in ethanol-induced conditioned taste aversion (CTA). A first experiment compared ethanol- and acetaldehyde-induced CTA. In a second experiment, cyanamide, an aldehyde dehydrogenase inhibitor, was administered before conditioning with either ethanol or acetaldehyde to investigate the effects of acetaldehyde accumulation. Methods. A classic CTA protocol was used to associate the taste of a saccharin solution with either ethanol or acetaldehyde injections. In experiment 1, saccharin consumption was followed by injections of either ethanol (0, 0.5, 1.0, 1.5 or 2.0 g/kg) or acetaldehyde (0, 100, 170 or 300 mg/kg). In experiment 2, the rats were pretreated with either saline or cyanamide (25 mg/kg) before conditioning with either ethanol or acetaldehyde. Results. Both ethanol and acetaldehyde induced significant CTA. However, ethanol produced a very strong CTA relative to acetaldehyde that induced only a weak CTA even at toxic doses. Cyanamide pretreatments significantly potentiated ethanol- but not acetaldehyde-induced CTA. Conclusions. The present results indicate that ethanol-induced CTA does not result from brain acetaldehyde effects. In contrast, it is suggested that the reinforcing effects of brain acetaldehyde might actually reduce ethanol-induced CTA. Our results also suggest that the inhibition of brain catalase activity may contribute to the potentiating effects of cyanamide on ethanol-induced CTA. Electronic Publication     

Caffeine promotes an ethanol-induced conditioned taste aversion: a dose-dependent interaction

The present study examined whether caffeine administered within a dose range previously shown to promote ethanol drinking would also alter an ethanol-induced conditioned taste aversion (CTA). The results revealed a dose-dependent interaction between caffeine and ethanol where caffeine (2.5 and 10 mg/kg) promoted an ethanol-induced CTA at a low ethanol dose (1.0 g/kg) but had no effect in blocking CTA at the higher ethanol dose (1.5 g/kg). These results were found to be unrelated to an alteration in ethanol metabolism, as caffeine had no effect in altering blood ethanol levels at the doses tested. In agreement with the reward comparison hypothesis, the present results suggest that rather than attenuate ethanol's "aversive" effects, caffeine may have promoted an ethanol-induced CTA by increasing the reinforcing efficacy of ethanol.     

Genetic differences in naloxone enhancement of ethanol-induced conditioned taste aversion

The influence of the opioid system on acquisition of an ethanol-induced conditioned taste aversion was examined in alcohol-preferring and avoiding inbred strains of mice (C57BL/6J and DBA/2J). Fluid-deprived mice from each strain received either ethanol alone, naloxone alone, or both ethanol and naloxone immediately after access to a novel tasting fluid. Naloxone alone (1 or 3 mg/kg) did not induce a conditioned taste aversion in either strain of mice. Administration of ethanol (1.5 g/kg) to DBA/2J mice produced a moderate taste aversion that was not affected by co-administration of naloxone. Although ethanol administered alone (3 g/kg) did not cause a taste aversion in C57BL/6J mice, the combination of ethanol and the higher dose of naloxone produced a significant taste aversion that increased across trials. A second experiment addressed the possibility that naloxone failed to enhance the ethanol-induced condition taste aversion in DBA/2J mice due to a floor effect on consumption. A lower ethanol dose (1 g/kg) was given alone or in combination with naloxone (1 or 3 mg/kg). Again, ethanol produced a moderate conditioned taste aversion that was not potentiated by naloxone. Subsequent conditioning with a high ethanol dose produced further suppression of intake, confirming that naloxone's failure to enhance aversion on earlier trials was not due to a floor effect. These data demonstrate a strain specific interaction between the aversive effect of ethanol and naloxone. More specifically, the results indicate that blockade of opioid receptors enhances the aversive effect of ethanol in C57BL/6J but not DBA/2J mice, suggesting that genetically determined differences in the endogenous opioid system of alcohol-preferring mice may mitigate ethanol's aversive effect.     

Effect of 3-amino-1,2,4-triazole on ethanol-induced narcosis, lethality and hypothermia in rats

It has been proposed that ethanol can be oxidized in brain via the peroxidatic activity of catalase and that centrally formed acetaldehyde may mediate several of the psychopharmacological actions of ethanol. The present study was designed to investigate the role of brain catalase in the mediation of ethanol-induced narcosis, hypothermia and lethality in rats. Rats were pretreated with the catalase inhibitor 3-amino-1,2,4-triazole (AT) or saline. Five hours later, animals in each pretreatment group received IP injections of ethanol (3 or 4 g/kg). Ethanol-induced narcosis was significantly attenuated in AT-pretreated rats compared to the saline control group. As well, AT pretreatments reduced significantly the lethal effect of these ethanol doses. However, AT-pretreated ethanol-injected animals significantly reduce their body temperature as compared to the saline-ethanol animals. Blood ethanol determinations revealed that AT did interfere with ethanol metabolism. AT inhibits significantly brain catalase activity at all doses used in this study. The results indicate a role for brain catalase in ethanol effects. Furthermore, they suggest that catalase may be involved in the oxidation of ethanol in brain and that centrally formed acetaldehyde may play a role in ethanol-induced narcosis and lethality, but not hypothermia.     

GABAA receptors modulate ethanol-induced conditioned place preference and taste aversion in mice

  Rationale: GABA_A receptor antagonists have been shown to reduce ethanol self-administration and ethanol-induced conditioned taste aversion (CTA) in rats, suggesting a role for the GABA_A receptor in modulating ethanol’s motivational effects. Objectives: The present experiments examined the effects of the GABA_A receptor antagonists, bicuculline and picrotoxin, on the acquisition of ethanol-induced conditioned place preference (CPP) and CTA in male DBA/2J mice. Methods: Mice in the CPP experiments received four pairings of ethanol (2 g/kg) with a distinctive floor stimulus for a 5-min conditioning session (CS+ sessions). During CS+ sessions, mice also received bicuculline (0, 1.0, 3.0, or 5.0 mg/kg) or picrotoxin (2.0 mg/kg) before an injection of ethanol. On intervening days (CS– sessions), the pretreatment injection was always vehicle followed by saline injections that were paired with a different floor type. For the preference test, all mice received saline injections and were placed on a half grid and half hole floor for a 60-min session. For the CTA experiments, mice were adapted to a 2-h per day water restriction regimen followed by five conditioning trials every 48 h. During conditioning trials, subjects received an injection of vehicle, bicuculline (0.5 and 2.0 mg/kg), or picrotoxin (0.75 and 2.5 mg/kg) before injection of 2 g/kg ethanol or saline following 1-h access to a saccharin solution. Results: Both picrotoxin and the lowest dose of bicuculline (1.0 mg/kg) significantly increased the magnitude of CPP relative to vehicle-treated controls. Picrotoxin alone did not produce place conditioning. Ethanol-stimulated locomotor activity was significantly reduced during conditioning trials with picrotoxin and the higher doses of bicuculline (3.0 and 5.0 mg/kg). Bicuculline did not alter ethanol-induced CTA; however, picrotoxin dose-dependently increased the magnitude of ethanol-induced CTA. Bicuculline and picrotoxin did not produce CTA when administered alone. Conclusions: Overall, these results suggest that blockade of GABA_A receptors with bicuculline and picrotoxin enhances ethanol’s motivational effects in the CPP paradigm; however, only picrotoxin enhances ethanol’s motivational effects in the CTA paradigm. Received: 12 September 1998 / Final version: 21 December 1998     

Modulation of ethanol-induced conditioned place preference in mice by 3-amino-1,2,4-triazole and D-penicillamine depends on ethanol dose and number of conditioning trials

Previous studies have shown that both 3-amino-1,2,4-triazole (AT), which inhibits metabolism of ethanol (EtOH) to acetaldehyde by inhibiting catalase, and D-penicillamine (D-P), an acetaldehyde-sequestering agent, modulate EtOH-conditioned place preference (CPP) in male albino Swiss (IOPS Orl) mice. These studies followed a reference-dose-like procedure, which involves comparing cues that have both been paired with EtOH. However, the role of EtOH-derived acetaldehyde has not been examined using a standard CPP method, and efficacy of these treatments could be different under the two circumstances. In the present investigation, we manipulated the strength of CPP across five separate studies and evaluated the effect of D-P and AT on EtOH-induced CPP following a standard unbiased CPP procedure. Mice received pairings with vehicle-saline injections with one cue and, alternatively, with AT- and D-P-EtOH with another cue. Our studies indicate that AT and D-P only disrupt CPP induced by EtOH in mice when the number of conditioning sessions and the dose of EtOH are low. These findings suggest that acquisition of EtOH-induced CPP may depend on the levels of acetaldehyde available during memory acquisition and the strength of the memory. Therefore, we propose that, at least when the memory processes are labile, brain acetaldehyde could participate in the formation of Pavlovian learning elicited by EtOH.     

Suicide inactivation of lactoperoxidase by 3-amino-1,2,4-triazole

Amitrole (3-amino-1,2,4-triazole) meets the criteria for a suicide (mechanism-based) inhibitor of lactoperoxidase. Amitrole causes rapid inactivation of lactoperoxidase only in the presence of hydrogen peroxide, and the kinetics are consistent with a suicide mechanism. Approximately 7 mol of radiolabeled amitrole binds covalently per equivalent of lactoperoxidase activity lost. The visible spectrum of lactoperoxidase inactivated by amitrole is unchanged, suggesting that covalent modification of the heme prosthetic group does not occur. The 13C NMR spectrum of lactoperoxidase inactivated by [13C]amitrole shows unique resonances which support the hypothesis that covalent binding occurs on the protein moiety. The similarities between lactoperoxidase and thyroid peroxidase suggest a similar mechanism for inhibition of thyroid hormone synthesis by amitrole.     

Dose- and time-dependent effect of an acute 3-amino-1,2,4-triazole injection on rat brain catalase activity

The results presented in this study demonstrate a progressive inhibition of rat brain catalase activity by AT in vivo. Furthermore, the inhibition of brain catalase by AT demonstrates the presence of hydrogen peroxide in brain, since AT inhibits catalase in the presence of this compound. The rate of inhibition of catalase seems to be dependent upon the rate by which H2O2 is generated. A time course study showed slower onset of the inhibition of brain as compared to liver catalase, possibly reflecting tissue hydrogen peroxide levels or, alternatively, a rate-limiting penetration of AT into brain and into the catalase compartment. The presence of AT in brain was confirmed over the time period of the observed inhibition of brain catalase. Catalase inhibitors are of particular interest in the study of the physiological role of catalase. This study further supports the use of AT in investigations designed to further understand the role of brain catalase.     

Effect of 3-amino-1,2,4-triazole treatment on catalase activity and triglyceride level in fatty liver of the rat

The effect of 3-amino-1,2,4-triazole (AT) pretreatment on the triglyceride (TG) level in CCl₄ (2.5 ml/kg, s.c.)-, ethanol (6 g/kg, p.o.)- or ethionine (1 g/kg, i.p.)- induced fatty liver in rats was studied, and the relationship between liver catalase activity and TG metabolism was investigated. The intoxication by each hepatotoxin increased the liver TG level in the rats. However, when the rats were pretreated with AT (1 g/kg, i.p.). the liver TG level decreased about 50 per cent, and the liver catalase activity was inhibited about 85 per cent. When AT and CCl₄ were injected simultaneously. they repressed about 50 per cent of the increase of the TG level caused by the intoxication of CCl₄. When AT alone was injected before the intoxication of ethanol, the liver catalase activity decreased by 50 per cent of that of the ethanol group and the increase in liver TG level was lowered by 40 per cent. On the other hand, when AT was injected after ethanol, the increase in liver TG level was depressed, although a decrease in catalase activity was not found. Although the serum TG level was decreased by CCl₄ treatment and increased by the ethanol treatment. AT did not show any effect on these changes. These results suggest that AT acts independently on the TG level and catalase activity in the liver.     

Inhibition of lipogenesis in isolated hepatocytes by 3-amino-1,2,4-triazole

3-Amino-1,2,4-triazole has been found to be an inhibitor of fatty acid synthesis by isolated rat hepatocytes. Half-maximal inhibition of fatty acid synthesis occurs at approximately 20mM. Acetyl-CoA carboxylase activity in homogenates of hepatocytes is not affected by previous exposure of the intact cells to 3-amino-1,2,4-triazole. The drug opposes the activation of partially purified acetyl-CoA carboxylase by citrate, but does not influence enzyme activity in the absence of citrate. As compared to fatty acid synthesis, cholesterol synthesis by the hepatocytes is more drastically depressed by incubation of the cells with 3-amino-1,2,4-triazole. Half-maximal inhibition of cholesterogenesis occurs at approximately 5 mM 8-amino-1,2,4-triazole.     

Metrazol produces conditioned taste aversion in rats

Employing a two bottle drinking procedure where an animal's preference is measured between plain water and a novel fluid, it was found that the convulsant drug Metrazol produced a conditioned taste aversion to saccharin. This finding is contrary to that of previous reports and highlights the sensitivity of the two bottle method in detecting a taste aversion.     

Non-specific enhancement of ethanol-induced taste aversion by naloxone

The conditioned taste aversion paradigm (CTA) was used to examine the effects of naloxone on ethanol-induced aversion towards a saccharine solution (3 conditioning and 11 extinction trials). Six groups of rats received conditioning trials consisting of two IP injections after saccharine presentation of different combinations of either ethanol (E: 1.75 g/kg), LiCl (L: 12 mEq/kg, 0.1 M), naloxone (N: 10 mg/kg) or saline (S); S-S, S-N, E-S, E-N, L-S and L-N. Naloxone by itself produced no aversion to the saccharin flavor. Based on the onset and extinction of aversion, naloxone significantly enhanced ethanol but also LiCl-induced CTA. The comparative data argues in favor of different mechanisms of action (1) between the aversive central effects of ethanol and morphine and (2) between ethanol's acute behavioral effects and negatively reinforcing properties. Enhancement of ethanol and LiCl-induced CTA by naloxone is compatible with hypernociceptive action of the opiate-antagonist and with the pain-modulating role of opiates in the CNS.     

Stimulus properties of fluvoxamine in a conditioned taste aversion procedure

A conditioned taste aversion (CTA) procedure in mice was used to investigate the stimulus effects of the serotonin reuptake inhibitors (SSRIs) fluvoxamine and fluoxetine. Fluvoxamine elicited a reliable CTA (ED₅₀ = 24 mg/kg, SC) and a number of drugs were tested as pre-exposure drugs. Pre-exposure to the serotonin (5-HT)_1A receptor agonists flesinoxan and ±-8-hydroxy-dipropylaminotetralin (8-OH-DPAT) prevented the CTA induced by fluvoxamine (50 mg/kg, SC). Pre-exposure with the 5-HT_2C receptor agonist MK 212 [6-chloro-2(1-piperazinyl)pyrazine] partially prevented the fluvoxamine-induced CTA, pre-exposure with the 5-HT_2A/2C receptor agonist DOI [1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane HCl] did not prevent the CTA induced by fluvoxamine. Flesinoxan pre-exposure also prevented the taste aversion induced by fluoxetine (10 mg/kg, SC) completely. This contrasts previous results obtained with fluoxetine, where was found that its stimulus is primarily mediated by 5-HT_2C, and to a lesser degree by 5-HT_1A receptors. Therefore, we compared the two SSRIs directly. Pre-exposure to fluvoxamine prevented the fluoxetine-induced CTA, whereas pre-exposure to fluoxetine only partially prevented the fluvoxamine-induced CTA. We conclude that 5-HT_1A receptors are involved in the stimulus properties of both fluvoxamine and fluoxetine, that 5-HT_2C receptors are involved in fluvoxamine and especially fluoxetine, and , based primarily on the cross-comparison tests, that the two SSRIs have somewhat different stimulus properties. Received: 3 March 1998 / Final version: 26 May 1998     

Stimulus properties of venlafaxine in a conditioned taste aversion procedure

Conditioned stimulus properties of venlafaxine are still unknown. In the present study, the discriminative stimulus properties of venlafaxine by using a conditioned taste aversion procedure were investigated. Swiss Webster mice were allowed to reach water from 2 pipettes for 20 min (09:00-11:30 h), plus 30 min (15:30-16:00 h), daily. During the 4 days, the test drugs [fluoxetine, escitalopram, tianeptine, reboxetine, and Nomega-nitro-L-arginine methyl ester (L-NAME)] were injected to mice at least 1 h after they had first water session. On day 5, they consumed glucose solution (5% w/v) and immediately injected with conditioning drug (venlafaxine 32 mg/kg). On day 8, mice were allowed to make a choice between water and glucose solution. The amount of glucose consumption as a percentage of total fluid intakes was calculated for each animal. Significant reduction in glucose choice was defined as conditioned taste aversion. Venlafaxine (32 mg/kg) induced a robust conditioned taste aversion in mice. Pre-exposure to tianeptine (2.5-10 mg/kg), fluoxetine (10 mg/kg), escitalopram (32 mg/kg), and reboxetine (5 mg/kg) substituted for venlafaxine by preventing the conditioned taste aversion induced by venlafaxine. L-NAME did not substitute for venlafaxine. Substitution of venlafaxine by fluoxetine, tianeptine, escitalopram, and reboxetine provides further evidence that both 5-HT and noradrenaline reuptake inhibition may play an important role in the stimulus effect of venlafaxine.     

Effects of atropine on conditioned taste aversion

Atropine sulfate, which has a deleterious effect on various learning tasks, was found to have a similar effect on the acquisition of conditioned taste aversion. Thus, intraperitoneal injection of atropine sulfate shortly before tasting was found to interfere with conditioning of the aversion, but injection of atropine after tasting did not. The interference effect was also obtained with intraventricular administration of atropine sulfate, but not with intraperitoneal injection of the peripherally-acting atropine methylnitrate. These results show that central rather than peripheral mechanisms are involved in this effect, and suggest that conditioned taste aversion, like other kinds of learning, involves cholinergic mediation.     

MK-801 induces a low intensity conditioned taste aversion

N-methyl-d-aspartate (NMDA) receptor antagonists are often used to assess the role of NMDA receptors in learning and memory processes. However, few studies have explored the possibility that the antagonists may induce a conditioned aversion when administered following flavor consumption. We report five experiments with rats intended to evaluate the MK-801 capacity to induce conditioned taste aversion. Our findings suggest that: i) MK-801 produces a low-intensity aversion following repeated pairings with saccharin (Experiments 1 and 2); ii) such aversion was not the result of a non-associative process (Experiment 3); and iii) pre-exposure to MK-801 does not interact with conditioned taste aversion induced by lithium chloride (Experiments 4 and 5). These findings suggest that MK-801 induces a low-intensity aversion, although the underlying mechanisms of this aversion may differ from those of a conditioned aversion produced by lithium chloride.