Weanlings' transfer of conditioned ethanol aversion from olfaction to ingestion depends on the unconditioned stimulus

Weanling (21-day-old) rats were exposed to an alcohol odor paired with either an interoceptive (apomorphine-induced illness) or exteroceptive (footshock-induced distress) reinforcer. Twenty-four hours later, ethanol preferences were measured in a locational olfactory test (ethanol vs lemon odor) or an ingestion test (5.6% v/v ethanol vs 0.25% v/v citric acid solution). Weanling rats expressed substantial olfactory aversions, independent of the reinforcer employed in conditioning. During the drinking test, however, only rats that had experienced the ethanol odor paired with internal malaise showed a significant reduction in the intake of the ethanol solution when compared to unpaired controls. Furthermore, rats that had experienced the ethanol odor paired with external distress drank significantly more of the ethanol solution than their controls. These results provide further evidence that olfactory experiences with ethanol can lead to changes in ethanol ingestion, and indicate that the nature of the unconditioned stimulus is critical in establishing the ingestive effect.     

Acute alcohol intoxication paired with aversive reinforcement: ethanol odor as a conditioned reinforcer in rat pups

Previous research has suggested that infant rats process ethanol sensory properties during acute alcohol intoxication. The present study was designed in order to examine if alcohol odor could act as an aversive conditioned stimulus after the organism experiences the state of intoxication paired with nociceptive stimulation (footshock). In a first experiment 11-day-old pups received intragastric alcohol administration (1.5 g/kg). At different postabsorptive intervals footshock was presented (0-30, 30-60, 60-90, or 90-120 min). An explicitly unpaired control group which experienced footshock prior to the state of intoxication was also employed. All animals were subsequently tested in terms of alcohol intake and ethanol locational odor preferences. Both assessments indicated that pups which were exposed to the unconditioned peripheral stimulus 30-60 min after receiving ethanol expressed strong alcohol aversions. In a second experiment pups were exposed to footshock during this postabsorptive interval. Twenty four hours later, pups experienced ambient ethanol odor paired with soft or rough texture surfaces. Differential texture aversions were registered in experimental animals when compared with controls which suffered the state of intoxication explicitly unpaired with footshock, or unpaired presentations of ethanol odor and the tactile stimuli under consideration. These results appear to support the hypothesis concerning orosensory processing during an acute state of intoxication. Additionally it seems that the hedonic value of sensory attributes of this drug varies as a function of associative processes occurring during such a state.     

Importance of tactile and olfactory cues to the inhibition of the grasshopper mouse's attack through toxicosis

This study examined the effect that toxicosis paired with the presence of a distinctive texture had on the inhibition of the grasshopper mouse's predatory attack. The first experiment measured the context in which tactile cues would be most effective by presenting prey with various combinations of added stimuli. The combination of distinctive tactile, olfactory, and gustatory cues produced the longest lasting inhibition. Because certain manipulations also had an unintended weak odor associated with them, the second experiment measured the importance of a weak or strong odor to inhibition of a mouse's attack. A combination of strong odor and distinctive texture paired with toxicosis inhibited an attack more effectively than a weak odor and a similar texture. In a third experiment, toxicosis was paired with an acraea moth caterpillar which has a highly distinctive texture. This produced the longest lasting inhibition of mouse's attack observed thus far. The ecological significance of a combination of a distinctive texture and odor to the inhibition of the grasshopper mouse's attack is discussed.     

Influence of rearing conditions on voluntary ethanol intake and response to stress in rats

The effects of exposure to four environmental rearing conditions on subsequent voluntary ethanol intake and response to immobilization stress were examined. Male weanling rats were reared in an enriched environment, with a female partner, with a male partner, or individually, for 90 days. At 111 days of age, voluntary consumption of ethanol in increasing concentrations (3 to 9%, v/v) was assessed. Following the ethanol-exposure period, rats were randomly divided into stressed and nonstressed groups and exposed to 3 h of immobilization. Results indicated that the enriched animals consumed greater amounts of ethanol as compared to all other groups, suggesting that the enriched environment and not handling, housing conditions, or the presence of another male or female is responsible for the observed increase in ethanol drinking behavior. Ulcer data indicated that among environmentally enriched rats, ethanol attenuated stress ulcer development relative to their non-ethanol-exposed but stressed controls. In nonstressed enriched rats, ethanol alone exacerbated stomach damage. We suggest that environmental rearing conditions markedly influence the complex interaction between ethanol intake and the response to stress.     

Effect of chronic ethanol administration on intermale aggression in rats

An experiment was performed to study the effect of chronic ethanol administration on intermale aggression in rats using a 24-hour resident-intruder test. During the resident-intruder test residents displayed virtually all of the agonistic behaviors, and intruders displayed virtually all of the defensive behaviors. Intruders treated with ethanol displayed more defensive behavior and elicited more agonistic behavior than control intruders. Twenty minutes into the resident-intruder test intruders showed the greatest increase in corticosterone (338% vs. 129%), while residents showed the greatest increase in testosterone (103% vs. 18%). On the 2nd day of the resident-intruder test intruders lost more weight than residents (21.5 g vs. 10.2 g). Plasma corticosterone levels remained elevated for the intruders, and in particular for those intruders displaying defensive behaviors regardless of the resident's behavior. Plasma testosterone levels remained elevated for those residents that were paired with intruders that displayed defensive behaviors regardless of the resident's behavior. The frequency and severity of biting attacks by ethanol residents was significantly greater than that of control residents. In addition, the locus of biting attack shifted from the upper back of intruders paired with control residents to the flanks, tail, lower feet, and ventral surface on intruders paired with ethanol residents.     

Acute alcohol intoxication paired with appetitive reinforcement: effects upon ethanol intake in infant rats

A recent study suggested that infant rats process alcohol odor and/or taste during acute ethanol intoxication probably due to ethanol elimination via respiration and salivation. The present set of experiments was meant to analyze the possibility that this orosensory processing may act as a conditioned stimulus when an appetitive reinforcer is paired with the state of intoxication. In the first experiment it was observed that intragastric administration of a mildly intoxicating ethanol dose (1.5 g/kg), paired during postabsorptive time intervals with oral infusion of sucrose, was sufficient to promote a significant preference to ethanol. In Experiment 2 different doses of ethanol were either paired or explicitly unpaired with sucrose administration. The result reported in Experiment 1 was replicated and it was observed that a higher dose (3.0 g/kg) unpaired with the reinforcer resulted in alcohol aversions in terms of alcohol consumption patterns. However, when the reinforcer was paired with this dose, the aversion was inhibited. Finally, in the third experiment results indicated that preexposure to alcohol odor eliminates sucrose-conditioned alcohol preferences. These results indicate that, in physiologically immature rats, alcohol preference can be regulated by prior associative experiences involving the state of intoxication and consequences internal and/or inherent to this state.     

Sex differences in the alcohol deprivation effect in rats

Few studies have investigated the relationship between early onset of alcohol drinking and relapse in adulthood, which may be larger in females, as revealed by studies using rats as subjects. The present study assesses the alcohol deprivation effect (ADE) in adult rats of both sexes that began alcohol consumption during preadolescence or adolescence. Female and male Wistar rats received free-choice access to water, 5, 10 and 20% (v/v) ethanol solutions during the ethanol exposures. The effects of age and sex on ADE in adult rats were assessed after repeated periods of abstinence. The results showed that female but not male rats increased ethanol intake after the second and third deprivation (showing ADE), irrespective of the onset-age of alcohol drinking.     

Ethanol impairs memory of a simple discrimination in adolescent rats at doses that leave adult memory unaffected

Adolescent rats are less sensitive than adults to the hypothermic, anxiolytic, motor impairing, hypnotic, and lethal effects of ethanol. In vitro experiments nevertheless suggest that hippocampal neural activity is more affected by ethanol in preweanling or adolescent rats than in adults. These data are complemented by in vivo results showing that pretraining ethanol impairs learning in adolescent rats at doses that do not affect adult learning. In order to determine if posttraining ethanol affects memory differently in adolescents than in adults, Sprague-Dawley albino rats of both ages were trained in an appetitively motivated odor discrimination in which they were required to dig in scented sand for sweetened cereal reward. Immediately after training subjects received intraperitoneal injections of 0, 0.5, or 1g/kg ethanol (12.6%). At test, 48h later, subjects were presented with unbaited discriminanda and the time (s) spent digging in the S+ and S- was measured. Adolescents, but not adults, showed impaired discrimination performance if training was followed by ethanol. A subsequent experiment discounted the possibility that impaired adolescent performance was due to ethanol-induced conditioned taste or odor aversions. It thus appears that relative to adults, memory in adolescent rats is more strongly affected by ethanol in a test of appetitive conditioning that excludes ethanol's effects on sensory and motivational influences during the learning experience.     

Olfactory aversive conditioning in the newborn (3-hr-old) rat impairs later suckling for water and milk

An olfactory conditioning paradigm tested whether newborn rats can acquire a conditioned aversion to olfactory events associated with their first postnatal meal 3-5 hr after birth. Exposure to lemon odor (conditioned stimulus [CS]) paired with intraoral infusions of 0.1% quinine (unconditioned stimulus) resulted in explicit conditioning. Responsiveness to a surrogate nipple providing water in the presence of the CS was significantly lower than the 3 control conditions. The conditioning dramatically suppressed responsiveness to a surrogate nipple providing milk, which normally is expressed voraciously in terms of sustained nipple attachment and milk intake. These findings suggest that as early as 3-5 hr after birth newborn rats are capable of aversive conditioning to odors in the context of suckling behavior.     

Early (preweanling) recognition of alcohol's orosensory cues resulting from acute ethanol intoxication

Three experiments were conducted in order to analyze the possibility that during acute alcohol intoxication, preweanling rats process ethanol's orosensory cues. Intragastric administration of an ethanol dose equivalent to 3.0 g/kg resulted in peak blood alcohol levels greater than 250 mg% (Experiment 1). Twenty-four hours after receiving this dose, 11-day-old pups expressed an ethanol odor aversion in a locational test. This aversion response was not observed when pups were previously administered an ethanol dose resulting in blood alcohol levels lower than 150 mg% (Experiment 2). Nevertheless when subjects received such a lower dose and 30 to 60 min after administration suffered nociceptive stimulation, alcohol aversions were also detected in terms of alcohol ingestion patterns and ethanol odor aversions (Experiment 3). These results appear to indicate that orosensory ethanol processing takes place during acute ethanol intoxication probably due to ethanol elimination via respiration, salivation, and/or hematogenic olfaction processes.     

Immunotoxic cholinergic lesions in the basal forebrain reverse the effects of entorhinal cortex lesions on conditioned odor aversion in the rat

Conditioned odor aversion (COA) corresponds to the avoidance of an odorized-tasteless solution (conditioned stimulus, CS) previously paired with toxicosis. COA occurs only when the interstimulus interval (ISI) is kept short, suggesting that the memory trace of the odor is subject to rapid decay. Previous experiments have shown that the entorhinal cortex (EC) is involved in the acquisition of COA, since lesion of the EC rendered COA tolerant to long ISI. Because EC lesions induce a septo-hippocampal cholinergic sprouting, the present experiment investigated whether COA tolerance to long ISI may be linked to this sprouting reaction. In a first experiment, male Long-Evans rats subjected to bilateral excitotoxic EC lesions combined to intracerebroventricular infusions of the selective cholinergic immunotoxin 192 IgG-saporin were exposed to odor-toxicosis pairing using a long ISI (120 min). Results showed that EC-lesioned rats displayed COA with the long ISI but not the control groups. In rats with EC combined to 192 IgG-saporin lesions, histological analysis demonstrated no evidence for cholinergic septo-hippocampal sprouting. In a second experiment, animals with 192-IgG saporin lesion showed a marked COA with a short ISI (5 min). These results suggest that the COA with the long ISI found in rats with EC lesions might involve a functional activity related to the EC lesion-induced hippocampal cholinergic sprouting. As the injection of 192 IgG-saporin alone did not affect COA with a short ISI, our data also point to a possible role of hippocampal cholinergic neurons in the modulation of memory processes underlying COA.     

The effects of prenatal stress on motivation in the rat pup

Exposure to prenatal stress (PNS) has been shown to induce a set of psychological and behavioral changes in developing offspring. We used the rodent model to investigate whether PNS produces changes in the ability of the pup to express social motivation. We used a set of behavioral tasks including monitoring ultrasonic vocalizations after isolation, a conditioned place preference, and a novel and familiar odor approach test. Pregnant Long-Evans rats were exposed to an unpredictable, variable stressor twice daily during the third week of gestation. Isolation vocalizations were assessed on postnatal day (PND) 10. Pup affinity for the dam was evaluated on PND 15. Typically, pups display a selective preference for an odor-paired environment only after the odor has been associated with the dam. This previous association produces a positive conditioned stimulus (CS). Normally, pups exposed to a neutral CS (odor paired with cotton balls) do not form this place preference. Results indicate that PNS exposed pups had significantly increased distress vocalizations and an equal preference for the positive and neutral conditioned stimuli. This type of alteration in forming early preferences could be detrimental because of decreases in the specificity of social learning and an impaired responsiveness in social relationships.     

The impact of emotional stress early in life on adult voluntary ethanol intake-results of maternal separation in rats

The combination of genetic and environmental factors determines the individual vulnerability for excessive ethanol intake, possibly leading to dependence. The environmental influences early in life represent examples of determinant factors for adult behaviour and can be protective as well as risk factors. Maternal separation is one model to examine the long-term consequences of early environmental experiences on neurochemistry and behaviour, including drug-taking behaviour in experimental animals. In the present review, findings from studies using repeated short and prolonged periods of maternal separation, with emphasis on effects on voluntary ethanol intake in rats with or without a genetic predisposition for high voluntary ethanol intake, are summarized. Despite some contradictory results, the general picture emerging shows that short periods of maternal separation during the postnatal period result in a lower adult voluntary ethanol intake in male rats. Prolonged periods of maternal separation were found to induce a high voluntary ethanol intake in male rats, including rats with a genetic predisposition for high ethanol intake. Results from the literature also show that changes were not just related to time of separation but were also related to the degree of handling. Interestingly, in terms of voluntary ethanol intake, female rats were generally not affected by postnatal maternal separation. The reasons for these sex differences need further investigation. In terms of neurobiological consequences of maternal separation, conclusive data are sparse and one of the future challenges will, therefore, be to identify and characterize underlying neurobiological mechanisms, especially in the individual animal.     

Old-age deficits in the sense of smell as gauged by thresholds, magnitude matching, and odor identification

Twenty elderly subjects (70-90 years old) and 20 young control subjects (18-24 years old) underwent three kinds of olfactory testing: absolute thresholds to three odorants (d-limonene, iso-amyl butyrate, benzaldehyde), magnitude matching of these odorants to salt tastes, and odor identification of 30 common substances. For all three odorants elderly subjects' mean threshold significantly exceeded that of the young by about ninefold for d-limonene, about threefold for benzaldehyde, and about twofold for iso-amyl butyrate. These threshold differences predict approximate concentration differences necessary to arouse the same estimated odor strength above the threshold for the elderly and the young. Young subjects also scored better than the elderly in odor identification, even when subjects were given four alternatives from which to select the correct label. Unimpaired olfactory functioning is uncommon in the elderly; correlational tests show that as a group the young have better olfactory ability and show more interindividual uniformity.     

Acute ethanol ingestion impairs appetitive olfactory learning and odor discrimination in the honey bee

Invertebrates are valuable models for increasing our understanding of the effects of ethanol on the nervous system, but most studies on invertebrates and ethanol have focused on the effects of ethanol on locomotor behavior. In this work we investigate the influence of an acute dose of ethanol on appetitive olfactory learning in the honey bee (Apis mellifera), a model system for learning and memory. Adult worker honey bees were fed a range of doses (2.5%, 5%, 10%, or 25%) of ethanol and then conditioned to associate an odor with a sucrose reward using either a simple or differential conditioning paradigm. Consumption of ethanol before conditioning significantly reduced both the rate of acquisition and the asymptotic strength of the association. Honey bees also exhibited a dose dependent reduction in arousal/attention during conditioning. Consumption of ethanol after conditioning did not affect recall 24h later. The observed deficits in acquisition were not due to the affect of ethanol on gustatory sensitivity or motor function. However, honey bees given higher doses of ethanol had difficulty discriminating amongst different odors suggesting that ethanol consumption influences olfactory processing. Taken together, these results demonstrate that an acute dose of ethanol affects appetitive learning and olfactory perception in the honey bee.     

Eyeblink classical conditioning and interpositus nucleus activity are disrupted in adult rats exposed to ethanol as neonates

Neonatal exposure to ethanol in rats, during the period of brain development comparable to that of the human third trimester, produces significant, dose-dependent cell loss in the cerebellum and deficits in coordinated motor performance. These rats are also impaired in eyeblink conditioning as weanlings and as adults. The current study examined single-unit neural activity in the interpositus nucleus of the cerebellum in adults following neonatal binge ethanol exposure. Group Ethanol received alcohol doses of 5.25 g/kg/day on postnatal days 4–9. Group Sham Intubated underwent acute intragastric intubation on postnatal days 4–9 but did not receive any infusions. Group Unintubated Control (from separate litters) did not receive any intubations. When rats were 3–7 mo old, pairs of extracellular microelectrodes were implanted in the region of the interpositus nucleus. Beginning 1 wk later, the rats were given either 100 paired or 190 unpaired trials per day for 10 d followed by 4 d of 100 conditioned stimulus (CS)-alone trials per day. As in our previous study, conditioned response acquisition in Group Ethanol rats was impaired. In addition, by session 5 of paired acquisition, Group Sham Intubated and Group Unintubated Control showed significant increases in interpositus nucleus activity, relative to baseline, in the CS–unconditioned stimulus interval. In contrast, Group Ethanol failed to show significant changes in interpositus nucleus activity until later in training. These results indicate that the disruption in eyeblink conditioning after early exposure to ethanol is reflected in alterations in interpositus nucleus activity.     

Extinction of conditioned odor potentiation of startle

Several recent studies with rats (Sprague-Dawley strain) have documented that an odor previously paired with shock potentiates the acoustic startle response, a phenomenon referred to as conditioned odor potentiation of startle (OPS). A surprising finding in these studies was that OPS did not extinguish even though the odor was present throughout the 25-min test session. Therefore, the present study more fully examined extinction of OPS. The results of Experiment 1, which employed both within-subject and between-group comparisons, showed that extinction of OPS occurred in adult rats only after several days of testing. Experiment 2 used the between-group procedure and found similar results with 23-day-old rats, the youngest age that exhibits the OPS effect. Experiment 2 also showed that giving rats 15 odor-shock pairings at 16 days of age, an age where they acquire the odor-shock association but cannot express it via OPS, does not increase subsequent resistance to extinction following odor-shock pairings at 23 days of age. Taken together, the results of this study show that (1) although OPS is somewhat resistant to extinction, it does extinguish with repeated tests and (2) suggests that there are no age differences in the rate of extinction of OPS.     

Bilateral lesions of the insular cortex or of the prefrontal cortex block the association between taste and odor in the rat

The neural basis for the association between taste and odor was investigated in the rat. First, behavioral procedures to study the mechanisms underlying the association between qualitative aspects of the odor and taste in rats were developed. Rats were presented with several pairings of the 0.3 mol/L NaCl solution and a flavor, and pairings of distilled water and another flavor. Then the rats received an IP injection of a furosemide to develop sodium deficiency. On the next day, the rats were presented with either of two types of odor-flavored water: water in which the flavor had been paired with NaCl, or water in which the flavor (grape or coffee) had been paired with distilled water. Normal rats avoided ingesting the water flavored with the odor previously paired with NaCl. Sodium-deprived rats, however, ingested the water flavored with that odor. Rats with lesions in either the insular cortex or in the prefrontal cortex neither preferred nor avoided the water flavored with the odor paired with NaCl. It was concluded that rats acquire association between taste and odor, and that the insular and the prefrontal cortices of the rats were involved in this association.     

Sex differences in adolescent ethanol drinking to behavioral intoxication

Rodent models have been especially useful for investigating adolescent ethanol exposure. However, there is a paucity of studies examining sex differences in behavioral intoxication from adolescent ethanol drinking. Here, we used an ethanol drinking model to investigate if adolescent rats of both sexes readily drink ethanol to measurable behavioral intoxication, indicated by increased impulsive action and motor incoordination. Beginning on postnatal day (P) 28, male and female Long‐Evans rats were given 30‐min access to a solution of sucrose (20%) or sweetened ethanol (20% sucrose +15% ethanol) every other day until P60 and once after 2 weeks of forced abstinence (on P75). On alternate (nondrinking) days, rats were reinforced with a food pellet for making a cued nosepoke response. Beginning on P56, rats were tested in this task after drinking sessions to assess ethanol‐induced changes in impulsive action, defined as premature responding prior to cue presentation. Motor coordination was assessed before and after drinking sessions using an incline plane test. Adolescent male and female rats readily consumed ethanol to behavioral intoxication, measured as reduced motor coordination. Following forced abstinence, females displayed greater ethanol‐induced impulsive action. These studies provide evidence for sex differences in behavioral intoxication following adolescent ethanol drinking.     

Age-related changes in contextual associative learning

The hippocampus plays a critical role in processing contextual information. Although age-related changes in the hippocampus are well documented in humans, nonhuman primates, and rodents, few studies have examined contextual learning deficits in old rats. The present study investigated age-related differences in contextual associative learning in young (6 mo) and old (24 mo) rats using olfactory stimuli. Stimuli consisted of common odors mixed in sand and placed in clear plastic cups. Testing was conducted in two boxes that represented two different contexts (Context 1 and Context 2). The contexts varied based on environmental features of the box such as color (black vs. white), visual cues on the walls of the box, and flooring texture. Each rat was simultaneously presented with two cups, one filled with Odor A and one filled with Odor B in each context. In Context 1, the rat received a food reward for digging in the cup containing Odor A, but did not receive a food reward for digging in the cup containing Odor B. In Context 2, the rat was rewarded for digging in the cup containing Odor B, but did receive a reward for digging in the cup containing Odor A. Therefore, the rat learned to associate Context 1 with Odor A and Context 2 with Odor B. The rat was tested for eight days using the same odor problem throughout all days of testing. The results showed no significant difference between young and old rats on the first two days of testing; however, young rats significantly outperformed old rats on Day 3. Young rats continued to maintain superior performance compared to old rats on Days 4-8. The results suggest that aging results in functional impairments in brain regions that support memory for associations between specific cues and their respective context.