HAD Rats Research Articles

PRECLINICAL STUDY: Effects of concurrent access to multiple ethanol concentrations and repeated deprivations on alcohol intake of high‐alcohol‐drinking (HAD) rats

High-alcohol-drinking rats, given access to 10% ethanol, expressed an alcohol deprivation effect (ADE) only after multiple deprivations. In alcohol-preferring (P) rats, concurrent access to multiple ethanol concentrations combined with repeated cycles of EtOH access and deprivation produced excessive ethanol drinking. The current study was undertaken to examine the effects of repeated alcohol deprivations with concurrent access to multiple concentrations of ethanol on ethanol intake of HAD replicate lines of rats. HAD-1 and HAD-2 rats received access to 10, 20 and 30% (v/v) ethanol for 6 weeks. Rats from each replicate line were assigned to: (1) a non-deprived group; (2) a group initially deprived of ethanol for 2 weeks; or (3) a group initially deprived for 8 weeks. Following the restoration of the ethanol solutions, cycle of 2 weeks of ethanol exposure and 2 weeks of alcohol deprivation was repeated three times for a total of four deprivations. Following the initial ethanol deprivation period, deprived groups significantly increased ethanol intakes during the initial 24-hour re-exposure period. Multiple deprivations increased ethanol intakes, shifted preference to higher ethanol concentrations and prolonged the duration of the elevated ethanol intakes for up to 5 days. In addition, repeated deprivations increased ethanol intake in the first 2-hour re-exposure period as high as 5-7 g/kg (which are equivalent to amounts consumed in 24 hours by HAD rats), and produced blood ethanol levels in excess of 150 mg%. The results indicate that HAD rats exhibit 'loss-of-control' of alcohol drinking with repeated deprivations when multiple ethanol concentrations are available.

Disposition of Naltrexone after Intravenous Bolus Administration in Wistar Rats, Low-Alcohol-Drinking Rats and High-Alcohol-Drinking Rats

Reports have shown that interspecies differences in the metabolism and pharmacokinetics of naltrexone are a rule rather than exception. However, there is paucity of information on the disposition of naltrexone in selectively bred rat lines that reliably exhibit high and low voluntary alcohol consumption, and are often used to study alcohol-drinking behavior. We have characterized the pharmacokinetic profiles of naltrexone in selectively bred rat lines: high-alcohol-drinking (HAD-1) and low-alcohol-drinking (LAD-1) rats as well as the native Wistar strain. This study was carried out to establish a baseline pharmacokinetic profile of naltrexone in these rats prior to evaluating its pharmacokinetic profile in polymeric controlled-release formulations in our laboratory. The hypothesis is that alcohol-preferring and non-alcohol-preferring lines of rats should differ in the disposition of intravenously administered naltrexone. Naltrexone administration and blood collection were via the jugular vein. In a parallel experiment, naltrexone was administered via the jugular vein, but urine was collected using the Nalgene metabolic cage system. Data were analyzed by a noncompartmental approach. Results show a high clearance that is close to or higher than hepatic blood flow in all groups (Wistar > LAD-1 > HAD-1, but with a statistically significant difference only between Wistar and HAD-1). Volume of distribution (∼2.5–3 l/kg) and the half-life (∼1 h) were similar. Urinary elimination of naltrexone was small, but also showed differences between the rats: HAD-1 > LAD-1 > Wistar, but with a statistically significant difference only between HAD-1 and Wistar rats. This study has therefore established the baseline disposition characteristics of naltrexone in these strains of rats.

Schedule-induced Polydipsia in Lines of Rats Selectively Bred for High and Low Ethanol Preference

Ethanol drinking was assessed in the P/NP, HAD1/LAD1, and HAD2/LAD2 lines of rats under environmental conditions that produce schedule-induced polydipsia. Female rats (n = 8/line), maintained at 85% of free-feeding body weights, underwent daily 1-h sessions during which 45-mg food pellets were delivered every 60 s. Water, 2, 4, 8, 16, or 32% w/v ethanol solution was available from a single bottle for 8 consecutive sessions at each concentration, with blood-ethanol levels (BELs) determined after selected sessions. P and HAD2 rats drank more water and ethanol than their non-preferring counterparts, while HAD1 and LAD1 rats did not differ. Ethanol intake and BELs were positively correlated (r = 0.75) across lines. Finally, rats were allowed 14 daily choice sessions with 8% ethanol and water concurrently available. Water intake generally exceeded ethanol intake in all lines, while P rats drank similar amounts of both fluids. These line differences indicate pleiotropic effects of genes that mediate ethanol intake and schedule-induced behaviors.

Effects of short deprivation and re-exposure intervals on the ethanol drinking behavior of selectively bred high alcohol-consuming rats

Alcoholics generally display cycles of excessive ethanol intake, abstinence and relapse behavior. Using an animal model of relapse-like drinking, the alcohol deprivation effect (ADE), our laboratory has shown that repeated 2-week cycles of ethanol deprivation and re-exposure, following an initial 6-week access period, result in a robust ADE by alcohol-preferring (P) and high alcohol-drinking (HAD-1 and HAD-2) rats. These rat lines have been selectively bred to prefer a 10% ethanol solution over water. The present study examined whether P and HAD rats would display an ADE using much shorter ethanol deprivation and re-exposure intervals. Rats were given either continuous or periodic concurrent access to multiple concentrations (10%, 20%, and 30% [vol/vol]) of ethanol. The periodic protocol involved access to ethanol for 12 days followed by four cycles of 4 days of deprivation and 4 days of re-exposure to ethanol access. High–alcohol-drinking rats displayed a robust 24-h ADE upon first re-exposure (HAD-1: ∼5 vs. 8 g/kg/day; HAD-2: ∼6 vs. 9 g/kg/day, baseline vs. re-exposure), whereas P rats (∼7 vs. 8 g/kg/day) displayed a modest, nonsignificant, increase in 24-h intake. In a separate group of rats, ethanol intake and blood alcohol concentrations after the first hour of the fourth re-exposure cycle were HAD-1: 2.0 g/kg and 97 mg%, HAD-2: 2.3 g/kg and 73 mg%, and P: 1.2 g/kg and 71 mg%; with all three lines displaying a robust first hour ADE. These findings suggest that (a) an ADE may be observed with short ethanol deprivation and re-exposure intervals in HAD rats, and (b) the genetic make-up of the P and HAD rats influences the expression of this ADE.

Chronic ethanol consumption increases dopamine uptake in the nucleus accumbens of high alcohol drinking rats

Past research has indicated that chronic ethanol exposure enhances dopamine (DA) neurotransmission in several brain regions. The present study examined the effects of chronic ethanol drinking on dopamine transporter (DAT) function in the nucleus accumbens (Acb) of High-Alcohol-Drinking replicate line 1 (HAD-1) rats. HAD rats were given concurrent 24-h access to 15% ethanol and water or water alone for 8 weeks. Subsequently, DA uptake and the V max of the DAT were compared between the two groups using homogenates of the nucleus accumbens. DA uptake was measured following a 2 min incubation at 37°C in the presence of 8 nM [ 3H]DA. For kinetic analyses, DA uptake was assessed in the presence of 5 concentrations of [ 3H]DA ranging from 8 nM to 500 nM. Analyses of the data revealed a significant increase in DA uptake in the ethanol group compared to water controls. Kinetic analyses revealed the change in DA uptake to be a consequence of an increase in the V max of transport. These findings demonstrate that chronic free-choice oral ethanol consumption in HAD-1 female rats increases DA uptake in the Acb by increasing the V max of the transporter. However, it is not known whether the ethanol-induced change in V max is caused by differences in the actual number of available transporter sites or from a difference in the velocity of operation of a similar number of transporters. Overall, the data indicate that chronic ethanol consumption by HAD-1 rats produces prolonged neuroadaptations within the mesolimbic DA system, which may be important for the understanding of the neurobiological basis of alcoholism.

Effects of multiple alcohol deprivations on operant ethanol self-administration by high-alcohol-drinking replicate rat lines

Previously, we reported that the expression of an alcohol deprivation effect (ADE) under 24-h free-choice alcohol-drinking access in high-alcohol-drinking (HAD) replicate lines of rats is dependent upon repeated cycles of alcohol access and forced abstinence. In the present study, operant techniques (including progressive ratio measures) were used to examine the effects of initial deprivation length and number of deprivation cycles on the magnitude and duration of the ADE in HAD rats to test the hypothesis that repeated deprivations increase the reinforcing effects of ethanol. Adult male HAD-1 and HAD-2 rats were trained in two-lever operant chambers to concurrently self-administer 15% ethanol (v/v) on a fixed-ratio (FR)-5 schedule and water on an FR-1 schedule of reinforcement in daily 1-h sessions. Following 10 weeks of daily 1-h sessions, the HAD-1 and HAD-2 rats were randomly assigned to one of four groups ( n = 6–8/group/line): nondeprived, or deprived of alcohol for 2, 5, or 8 weeks. Following this initial period, the deprived groups were given 15% ethanol again in the operant chambers for a 2-week period, following which they were deprived again for 2 weeks (all three deprived groups). Following the fifth deprivation, the rats underwent a progressive ratio test to determine the breakpoints for the nondeprived and deprived groups. The expression of an ADE under operant conditions in HAD rats was dependent upon exposure to repeated cycles of ethanol access and abstinence. Additionally, repeated deprivations increased both the magnitude and the duration of the ADE as indicated by increased responding on the ethanol lever for more sessions. Breakpoint values for the deprived groups were 1.5-fold and twofold higher than the value for the nondeprived group for the HAD-1 and HAD-2 rats, respectively. The results suggest that repeated alcohol deprivations increased the expression of an ADE and the reinforcing effects of ethanol in both HAD replicate lines of rats, and these effects were more pronounced in the HAD-2 line than the HAD-1 line.

Decreased reward during acute alcohol withdrawal in rats selectively bred for low alcohol drinking

We have previously hypothesized that increased sensitivity to the dysphoric-like or aversive effects of alcohol withdrawal following an initial exposure to alcohol might be associated with a genetic propensity to avoid alcohol. A decrease in brain reward function, as measured by an elevation in intracranial self-stimulation (ICSS) reward threshold, is one of the few methods available to model dysphoric-like or aversive effects of drug withdrawal in rats. We compared brain reward function during withdrawal following an initial exposure to alcohol in alcohol-naïve rats selectively bred for high (HAD1 line) versus low (LAD1 line) voluntary alcohol consumption. Male HAD1 ( n = 5) and LAD1 ( n = 6) rats were implanted with unilateral electrodes in the medial forebrain bundle and trained to bar press for delivery of a 100 μA current that varied in frequency from 45 to 200 Hz. Responding for ICSS was generally stable within subjects across multiple experimental sessions on a given day and across several consecutive days prior to alcohol or water administration. ICSS responding was assessed in both rat lines prior to and at 12, 14, 16, 18, 20, and 24 h following a single intragastric infusion of alcohol (4.0 g/kg body weight) or water. Rats of the LAD1 line, but not those of the HAD1 line, exhibited a decrease in brain reward function as evidenced by a decrease in bar-press responding for ICSS and an increase in ICSS stimulation threshold during alcohol withdrawal. The results suggest that rats selectively bred for low alcohol drinking may experience dysphoric-like effects during withdrawal from an initial exposure to alcohol, while rats selectively bred for high alcohol drinking may not.

EFFECTS OF CHRONIC ALCOHOL TREATMENT ON ACOUSTIC STARTLE REACTIVITY DURING WITHDRAWAL AND SUBSEQUENT ALCOHOL INTAKE IN HIGH AND LOW ALCOHOL DRINKING RATS

The purpose of the present study is to determine whether the inverse genetic association between alcohol withdrawal magnitude and genetic propensity for alcohol drinking that we have previously identified in alcohol-naive rats given alcohol acutely, would also be seen following chronic alcohol exposure. The effect of forced, chronic alcohol treatment on subsequent voluntary alcohol drinking was also examined. Male rats from the high alcohol drinking (HAD2) and low alcohol drinking (LAD2) lines received two intragastric (IG) infusions of alcohol (3.0 g/kg BW; 25% v/v) or an equal volume of water, separated by 5 h, every day for 20 consecutive days (chronic alcohol treatment). Acoustic startle reactivity was assessed at 10, 14, and 18 h after the second infusion on days 1, 5, 10, 15, and 20. After acoustic startle testing was completed, all rats received two IG infusions of 3.0 g alcohol/kg BW, separated by 5 h, and blood alcohol content was assessed at 10, 14, and 18 h after the second alcohol infusion. All rats were then given a 24 h free-choice between alcohol and water for 8 weeks. Startle magnitude to a 120 dB tone was suppressed during alcohol withdrawal in both alcohol-treated HAD2 and LAD2 rats after 5, 10, and 15 days of alcohol treatment. Forced, chronic alcohol treatment produced metabolic tolerance in both the HAD2 and LAD2 lines and significantly suppressed subsequent voluntary alcohol intake in rats of the HAD2 line. Reduced acoustic startle reactivity during alcohol withdrawal in both HAD2 and LAD2 rats is consistent with our previous findings in the HAD2 but not the LAD2 line and may reflect reduced CNS excitability during withdrawal from forced alcohol exposure. Forced alcohol exposure robustly retarded the expression of a genetic predisposition toward alcohol drinking in rats selectively bred for high alcohol intake.

Circadian activity rhythms in selectively bred ethanol-preferring and nonpreferring rats

Chronic alcohol intake is associated with dramatic disruptions in sleep and other circadian biological rhythms in both humans and experimental animals. In human alcoholics, these disruptions persist during extended abstinence and appear to promote relapse to drinking. Whereas chronic ethanol intake alters fundamental properties of the circadian pacemaker in unselected rats, nothing is known concerning circadian pacemaker function in selectively bred ethanol-preferring and nonpreferring rats, which are the most widely accepted animal models of genetic predisposition to alcoholism. The present experiments were designed to characterize free-running circadian activity (wheel-running) rhythms under both constant darkness and constant light in selectively bred ethanol-preferring (P, HAD2) and nonpreferring (NP, LAD2) rats. Differences in circadian organization between ethanol-preferring and nonpreferring animals were seen for both pairs of selected lines (P vs. NP; HAD2 vs. LAD2), but these differences were not identical in the two line pairs. For example, although P rats showed shorter free-running periods than NP rats only in constant light, HAD2 rats showed shorter free-running periods than LAD2 rats only in constant darkness. In addition, ethanol-preferring HAD2 rats showed a high rate of rhythm “splitting” that was not seen in any of the other three lines. Taken together, these results suggest that the circadian pacemakers of P and NP rats differ mainly in light sensitivity, whereas those of HAD2 and LAD2 rats differ in their intrinsic period.

Neuropeptide Y in the Paraventricular Nucleus of the Hypothalamus Increases Ethanol Intake in High‐ and Low‐Alcohol‐Drinking Rats

The behavioral effects of neuropeptide Y (NPY) attributed to its actions in the hypothalamus are complex and include effects on feeding, sedation, and the hypothalamic-pituitary-adrenal axis. NPY infused into the paraventricular nucleus (PVN) increases ethanol intake in unselected rats. High-alcohol-drinking (HAD1) and low-alcohol-drinking (LAD1) rats differ in basal NPY levels in the PVN, and HAD1, but not LAD1, rats exhibit decreases in ethanol intake after infusion of NPY into the ventricles. This study examined whether NPY infused into the PVN alters ethanol intake in HAD1 and LAD1 rats. Female HAD1 (n = 14) and LAD1 (n = 18) rats were given 24-hr free-choice continuous access to 15% (v/v) ethanol and water for 6 weeks and then implanted bilaterally with cannulas aimed at the PVN. Two weeks later, rats received a series of microinfusions, each separated by 1 week, that included four doses of NPY (0.0, 0.25, 0.5, and 1.0 microg). Ethanol, water, and food were available ad libitum after infusions. All rats received a final microinfusion of 1.0 microg of NPY, after which ethanol and water, but no food, were made available for 2 hr. During the 2 hr after infusion, NPY yielded dose-dependent increases in both water and food consumption. With food concurrently available, the 0.25- and 1.0-microg doses of NPY did not alter baseline ethanol intake, whereas the 0.5-microg dose increased ethanol intake. Infusion of 1.0 microg of NPY in the absence of food yielded a decrease in water intake and an increase in ethanol intake relative to the same dose in the presence of food. Twenty-four hours after infusion, there were no effects of NPY on water and food intake, and increases in ethanol intake were no longer apparent. Increases in ethanol intake after infusion of NPY into the PVN may depend on NPY dose and whether food is concurrently available.

Effects of concurrent access to a single concentration or multiple concentrations of ethanol on ethanol intake by periadolescent high-alcohol-drinking rats

The objectives of the current study were to assess the effects of access to different concentrations of ethanol and sex of the animal on ethanol consumption of high-alcohol-drinking (HAD-1 and HAD-2) rats during adolescence [postnatal days (PNDs) 30 through 60]. At the beginning of adolescence (PND 30), the rats were given concurrent access to either a single concentration [15% volume/volume (vol./vol.)] or multiple concentrations [10%, 20%, and 30% (vol./vol.)] of ethanol and water. Analyses of ethanol consumption data revealed significant ( P < .025) main effects of line, ethanol condition, and week, and a significant line by sex by ethanol condition by week interaction. For the first week, both male and female HAD-1 and HAD-2 rats consumed more ethanol under the multiple ethanol concentration condition than under the single ethanol concentration condition. However, across the second through fourth weeks, this pattern was seen primarily in male and female HAD-1 rats and to a lesser degree in female HAD-2 rats. In general, female rats consumed more fluids than consumed by male rats, and male rats displayed a higher preference for ethanol over water ratio than observed for their female counterparts. In addition, in comparison with HAD-2 rats, HAD-1 rats drank more ethanol and displayed a higher preference for ethanol ratio. Overall, the current study results indicate that, compared with access to a single concentration (which is used in most studies), concurrent access to multiple concentrations of ethanol produced significantly higher ethanol intakes in periadolescent HAD rats, supporting the suggestion that this ethanol drinking condition would have a greater impact on neuronal development. In addition, although the replicate lines were selectively bred by using the same criteria and foundation stock, the higher ethanol intakes of the HAD-1 line, compared with intakes for the HAD-2 line, seen in the current study support the suggestion that there are some differences in their genetic make-up, affecting ethanol intake, which are expressed during periadolescence.

Confirmation of alcohol preference quantitative trait loci in the replicate high alcohol drinking and low alcohol drinking rat lines.

Selective breeding has been employed to develop replicate high-alcohol-drinking (HAD1 and HAD2) and low-alcohol-drinking (LAD1 and LAD2) rat lines from the heterogeneous N/Nih rat. Within-family selection and a rotational breeding design were used to discourage inbreeding (Li et al., 1993). A genome screen was previously performed using 459 HAD1xLAD1 F2 progeny to identify quantitative trait loci (QTLs) on rat chromosomes 5, 10, 12 and 16 that contribute to alcohol preference and consumption in these non-inbred rat models of alcoholism. To confirm these QTLs in the replicate lines, 16 HAD2 and 16 LAD2 rats were genotyped for microsatellite markers within each of these QTL intervals. Review of the genotypic data support confirmation of the QTLs on chromosomes 5 and 10; several markers in the QTL region display different alleles in the HAD2 and LAD2 rats, suggesting linkage disequilibrium between the microsatellite markers and the QTL. Although the QTL on chromosome 12 had the highest LOD score in the HAD1 and LAD1 studies, little evidence supported confirmation of this QTL based on the genotyped markers. Further evaluation of each of these QTL regions is ongoing in a sample of HAD2xLAD2 F2 progeny currently being generated that will be used to assess the evidence of linkage in each of these QTL regions.

Effects of Neuropeptide Y on Sucrose and Ethanol Intake and on Anxiety‐Like Behavior in High Alcohol Drinking (HAD) and Low Alcohol Drinking (LAD) Rats

Background: In a previous study, neuropeptide Y (NPY) administered into the lateral ventricles decreased ethanol intake in alcohol‐preferring (P) rats but not in alcohol‐nonpreferring (NP) or unselected Wistar rats. The purpose of the present investigation is to extend these findings in selectively‐bred high‐alcohol‐drinking (HAD)1 and low‐alcohol‐drinking (LAD)1 rats by examining the effects of intracerebroventricularly administered NPY on the elevated plus maze test of anxiety and on ethanol and sucrose intake.Methods: Female HAD1 and LAD1 rats were surgically implanted with cannula into the lateral ventricle. Following recovery, a test of anxiety was conducted in which the rats (n= 12–13/group) received either artificial cerebrospinal fluid (aCSF) or NPY (10 μg) 10 min prior to a 5‐min test on an elevated plus maze. Following anxiety testing, 11 HAD and 11 LAD rats were trained to self‐administer ethanol (8% w/v), and 5 HAD and 8 LAD rats were trained to self‐administer sucrose (2.5%) during daily 2‐hr sessions. A within‐subject design was used in which the rats were pretreated once a week with aCSF, 5 μg NPY, or 10 μg NPY prior to the drinking sessions.Results: HAD and LAD rats treated with aCSF did not differ in time spent in open arms of the plus maze. NPY increased time spent on the open arms to similar degrees in both rat lines. HAD rats consumed more ethanol and sucrose than LAD rats. NPY increased sucrose intake in both rat lines. However, the same doses of NPY reduced ethanol intake in HAD but not in LAD rats.Conclusion: The plus maze results indicated that selective breeding for high and low alcohol preference in the HAD1 and LAD1 rats, respectively, did not yield differences in anxiety‐like behavior and in response to the anxiolytic effects of NPY. The increases in sucrose intake were consistent with the known orexigenic effects of NPY. The decreased ethanol intake following NPY administration in HAD rats was similar to previous observations with P rats and is consistent with the hypothesis that ethanol intake and NPY activity may be inversely related.

Moderate doses of ethanol partially reverse avoidance learning deficits in high-alcohol-drinking rats

We previously reported that ethanol-naive high-alcohol-drinking (HAD1 and HAD2) rats exhibited selective deficits in active avoidance learning, as compared to low-alcohol-drinking (LAD1 and LAD2) rats, in a signaled bar-pressing task [Alcohol. Clin. Exp. Res. 24 (2000) 1778]. In the current study, we used appetitive and aversive learning tasks to assess whether administration of ethanol influences approach and avoidance learning in HAD and LAD rats. Rats were administered 0.0, 0.5, 1.0, or 1.5 g ethanol/kg body weight during appetitive and aversive conditioning sessions. We found that ethanol impaired acquisition of the appetitive conditioned response in a dose-dependent manner in both HAD and LAD rats, with 1.5 g/kg ethanol producing the greatest deficits. Notably, moderate doses of ethanol (0.5 and 1.0 g/kg) partially reversed avoidance learning deficits in HAD rats, but only when appetitive conditioning preceded aversive conditioning. The highest dose (1.5 g/kg EtOH) abolished avoidance responding altogether in HAD rats. Avoidance responding in LAD rats was not affected by any dose of ethanol. These results are consistent with previous studies suggesting that alcohol preference may be associated with increased fear or anxiety, but the conditions under which ethanol produces a reduction of fear and anxiety in HAD rats appear to be relatively complex.

Differential effects of saccharin supplementation on alcohol and water consumption by Wistar rats from different sources

The study was aimed to elucidate whether the dissimilar effects of concurrent presentation of sweet water on alcohol consumption previously reported for Wistar rats from different sources remain unchanged when alcohol is also flavored with 0.1% saccharin. Male Wistar rats from Laboratory of Experimental Biological Models (LEBM) and Krukovo animal farm (K) stocks having had 2 months free access to food, tap water, and 10% alcohol were given four consecutive two-bottle drinking tests: alcohol versus water, alcohol versus sweet water, sweet alcohol versus water, or sweet alcohol versus sweet water. The test order was quasi-random and each test lasted 4 days. In Wistar (K) rats, flavoring of either water or alcohol solution increased consumption of each of the fluids and decreased intake of concurrently available fluids. The elevation in water intake induced by its sweetening was antagonized by flavoring of alcohol solution. In Wistar (LEBM) rats, flavoring of either water or alcohol increased consumption of each of the fluid, but did not change the intake of alternative fluids. The stable alcohol consumption by Wistar (LEBM) rats and its suppression seen in Wistar (K) rats induced by concurrent presentation of flavored water parallel the patterns previously observed among P, sP, and HAD rats suggesting the existence among alcohol-consuming animals of typological diversity of alcohol motivation.

Ethanol effects on local cerebral glucose utilization in high-alcohol-drinking and low-alcohol-drinking rats

Divergent ethanol-drinking behavior in rats selectively bred for high- or low-ethanol-drinking behavior could be related to differences in the sensitivity of the CNS to ethanol. In the current study, we examined the effects of acute (i.e., single injection) ethanol administration on local cerebral glucose utilization (LCGU) within selected brain regions of high-alcohol-drinking (HAD) and low-alcohol-drinking (LAD) rats. Adult, male, HAD and LAD rats from replicate line 2 were injected intraperitoneally with saline, or ethanol, at doses of 0.25 g/kg or 1.0 g/kg, during their dark cycle; 10 min later, [ 14C]-2-deoxyglucose ([ 14C]-2-DG; 125 μCi/kg) was injected into the femoral vein. Timed arterial blood samples were collected over 45 min and assayed for plasma glucose, ethanol, and [ 14C]-2-DG levels. Rats were then decapitated, and their brains were quickly extracted and frozen in isopentane at –50°C. Coronal brain sections were prepared and apposed to x-ray film for 2 days, and image densities were determined by using quantitative autoradiography. Data were collected from several key limbic (nucleus accumbens, ventral tegmental area, olfactory tubercle, amygdala, hippocampus, ventral pallidum, and septum), basal ganglia, cortical (medial prefrontal, frontal, parietal, temporal, occipital, entorhinal, piriform, and cingulate), and subcortical (thalamus, habenula, and superior colliculus) structures. After administration of both low (0.25 g/kg) and moderate (1.0 g/kg) doses of ethanol, LCGU values were lower, relative to those for saline controls, in several CNS regions (lateral septum; posterior cingulate, frontal, parietal, and temporal cortices; dorsomedial striatum; and dorsomedial thalamus) of LAD but not HAD rats. These findings may indicate that certain CNS regions of LAD-2 rats are more sensitive than regions of HAD-2 rats to the effects of low-to-intermediate doses of ethanol.

Neurophysiological Profiles of Replicate Line 2 High‐Alcohol‐Drinking (HAD‐2) and Low‐Alcohol‐Drinking (LAD‐2) Rats

Rationale A select number of electrophysiological findings have been demonstrated to differentiate rat lines selectively bred for high and low ethanol preference.Objective In the present study, EEGs and event‐related potentials (ERPs) of high‐alcohol‐drinking (HAD) and low‐alcohol‐drinking (LAD) rats from replicate line 2 (HAD‐2 and LAD‐2) were assessed to determine if their neurophysiological profiles are similar to selected lines previously evaluated.Methods Rats obtained from Indiana University were implanted with cortical and amygdalar recording electrodes. Baseline EEG and ERPs were assessed in ethanol‐naïve HAD‐2 and LAD‐2 rats. Animals subsequently were trained to self‐administer ethanol by using a sucrose‐substitution procedure.Results Compared with LAD‐2 rats, HAD‐2 rats displayed greater parietal cortical power in the 6 to 32 Hz frequency range of the EEG. Greater parietal cortical peak frequency in the 2 to 4 Hz range and decreased frontal, parietal, and amygdalar peak frequencies in the 16 to 32 Hz frequency range were also seen. Compared with LAD‐2 rats, HAD‐2 rats had decreased P2 latency of ERPs recorded in the parietal cortex. HAD‐2 rats also had greater frontal, parietal, and amygdalar P2 amplitudes, greater frontal and parietal cortical P1 amplitudes, and greater parietal cortical P3 amplitudes compared with LAD‐2 rats. As anticipated, HAD‐2 rats consumed significantly greater levels of sucrose, sucrose‐ethanol, and ethanol over the course of the sucrose‐substitution procedure compared with LAD‐2 rats.Conclusions These data suggest that increased cortical power is associated with high ethanol preference in a number of selectively bred rat lines. However, unique electrophysiological characteristics may index alcohol preference in each line.

Neurophysiological Profiles of Replicate Line 2 High-Alcohol-Drinking (HAD-2) and Low-Alcohol-Drinking (LAD-2) Rats

Rationale A select number of electrophysiological findings have been demonstrated to differentiate rat lines selectively bred for high and low ethanol preference. Objective In the present study, EEGs and event-related potentials (ERPs) of high-alcohol-drinking (HAD) and low-alcohol-drinking (LAD) rats from replicate line 2 (HAD-2 and LAD-2) were assessed to determine if their neurophysiological profiles are similar to selected lines previously evaluated. Methods Rats obtained from Indiana University were implanted with cortical and amygdalar recording electrodes. Baseline EEG and ERPs were assessed in ethanol-naïve HAD-2 and LAD-2 rats. Animals subsequently were trained to self-administer ethanol by using a sucrose-substitution procedure. Results Compared with LAD-2 rats, HAD-2 rats displayed greater parietal cortical power in the 6 to 32 Hz frequency range of the EEG. Greater parietal cortical peak frequency in the 2 to 4 Hz range and decreased frontal, parietal, and amygdalar peak frequencies in the 16 to 32 Hz frequency range were also seen. Compared with LAD-2 rats, HAD-2 rats had decreased P2 latency of ERPs recorded in the parietal cortex. HAD-2 rats also had greater frontal, parietal, and amygdalar P2 amplitudes, greater frontal and parietal cortical P1 amplitudes, and greater parietal cortical P3 amplitudes compared with LAD-2 rats. As anticipated, HAD-2 rats consumed significantly greater levels of sucrose, sucrose-ethanol, and ethanol over the course of the sucrose-substitution procedure compared with LAD-2 rats. Conclusions These data suggest that increased cortical power is associated with high ethanol preference in a number of selectively bred rat lines. However, unique electrophysiological characteristics may index alcohol preference in each line.

Ethanol- and Sucrose-Reinforced Appetitive and Consummatory Responding in HAD1, HAD2, and P Rats

Introduction Ethanol-preferring (P) rats and high-alcohol-drinking (HAD1 and HAD2) rats have been selectively bred to consume greater amounts of ethanol than nonselected rat strains. These three rat lines also show increased levels of responding for ethanol in operant paradigms that assess a combined appetitive/consummatory response. Methods The present experiment used a model of reinforced responding that procedurally separates the appetitive, or seeking, response requirement from consummatory responding to compare seeking and intake responding in P and HAD rats. Subjects (n= 7 or 8 per group) were trained to make 25 lever-press responses, which were followed by 20 min of access to a sipper tube spout containing either 10% ethanol (10E) or (in separate groups of subjects) 3% sucrose (3S). After training, a single nonreinforced session was conducted to assess the limit to appetitive responding under extinction conditions. After this single nonreinforced session, three successive across-session breakpoint determinations were made for 10E and 3S by increasing the response requirement over days until subjects failed to complete the requirement. A final extinction session was then conducted. Results Appetitive responding during both the nonreinforced and breakpoint sessions indicated that P rats made significantly more responses overall than HAD rats in both the ethanol and sucrose groups. P rats also consumed more sucrose than HAD rats, with no differences in ethanol consumption between the lines (1.0–1.5 g/kg/20 min). Appetitive responding in the HAD rats in the ethanol groups was comparable to that reported previously for nonselected Long-Evans rats. Conclusions These findings indicate that appetitive and consummatory processes are distinct and that P rats have an increased tendency to both seek and drink ethanol and sucrose solutions, making this selected line useful when modeling both “craving” and “loss of control” related behaviors involved in excessive alcohol consumption.

Inverse Genetic Association Between Alcohol Preference and Severity of Alcohol Withdrawal in Two Sets of Rat Lines Selected for the Same Phenotype

The purpose of the present study was to determine whether alcohol-naïve rats selectively bred for alcohol preference or nonpreference differ in alcohol withdrawal severity using two sets of rat lines selectively bred for the same phenotype. Alcohol-naïve male rats from the high alcohol drinking (HAD1) and low alcohol drinking (LAD1) rat lines and from the alcohol preferring (P) and nonpreferring (NP) rat lines received an intragastric infusion of alcohol (4.0 g/20.3 ml/kg; 25% v/v) or an equal volume of water once a day for 10 consecutive days. Alcohol withdrawal severity was assessed at using a behavioral rating scale and a radiant heat assay measured analgesia at 10, 12, 14, 16, 18, and 24 hrs following infusion of alcohol or water on days 1, 5, and 10 of treatment. Data were analyzed using body weight as a co-factor to correct for differences in body weight between the HAD1/LAD1 and P/NP lines. Acute (1 day) but not repeated alcohol treatment (5 or 10 days) produced mild behavioral signs of withdrawal in LAD1 but not in HAD1 rats. HAD1 and LAD1 rats showed alcohol-induced analgesia after 1 and 5 days of alcohol treatment that disappeared by day 10 in both lines. Repeated alcohol treatment (5 days) produced mild behavioral signs of withdrawal in NP but not in P rats. Neither P nor NP rats showed alcohol-induced analgesia after 1, 5, or 10 days of alcohol treatment. An inverse genetic association was found between alcohol preference and severity of alcohol withdrawal in two sets of rat lines selected for the same phenotype. The pattern of alcohol withdrawal that emerged over the course of the 10 days of alcohol treatment differed between the two lines selected for low alcohol drinking (LAD1 and NP), suggesting that unique sets of genes may influence alcohol withdrawal severity in the two lines.