A major QTL for acute ethanol sensitivity in the alcohol tolerant and non-tolerant selected rat lines.

The Alcohol Tolerant and Alcohol Non-Tolerant rats (AT, ANT) were selectively bred for ethanol-induced ataxia as measured on the inclined plane. Here we report on a quantitative trait locus (QTL) study in an F(2) intercross population derived from inbred AT and ANT (IAT, IANT) and a follow-up study of congenics that were bred to examine one of the mapped QTLs. Over 1200 F(2) offspring were tested for inclined plane sensitivity, acute tolerance on the inclined plane, duration of the loss of righting reflex (LORR) and blood ethanol at regain of the righting reflex (BECRR). F(2) rats that were in the upper and lower 20% for inclined plane sensitivity were genotyped with 78 SSLP markers. Significant QTLs for inclined plane sensitivity were mapped on chromosomes 8 and 20; suggestive QTLs were mapped on chromosomes 1, 2 and 3. Highly significant QTLs for LORR duration (LOD = 12.4) and BECRR (LOD = 5.7) were mapped to the same locus on chromosome 1. Breeding and testing of reciprocal congenic lines confirmed the chromosome 1 LORR/BECRR QTL. A series of recombinant congenic sub-lines were bred to fine-map this QTL. Current results have narrowed the QTL to an interval of between 5 and 20 Mb. We expect to be able to narrow the interval to less than 5 Mb with additional genotyping and continued breeding of recombinant sub-congenic lines.

Developmental alterations of ethanol sensitivity in selectively bred high and low alcohol sensitive rats.

Initial sensitivity and acute tolerance to ethanol have been implicated as risk factors in the development of alcoholism in humans. These behaviors were investigated in rats selectively bred for differences in hypnotic sensitivity following their first dose of ethanol in two different experiments. In Experiment 1, developmental profiles of the association between initial sensitivity and acute tolerance induced by a single exposure to ethanol were examined using male and female high, low, and control alcohol sensitive (HAS, LAS, and CAS) rats. Dose-response curves were constructed for duration of the loss of the righting reflex and for blood ethanol concentration (BEC) at the regain of the righting reflex. Animals were tested with a single ethanol dose ranging from 1.5 to 5.0 g/kg at either 15, 25, 40, 70, 120, or 180 days of age (DOA). For each group, acute tolerance to ethanol was estimated by the slope of the regression line using dose of ethanol and mean BEC at regain. In general, all rat lines showed an increase in hypnotic sensitivity to ethanol with age. To a large degree, the lower sensitivity observed in 15 and 25 DOA HAS and LAS rats was associated with an increase in the development of acute ethanol tolerance relative to older rats. Divergence of the LAS and CAS lines was evident by 25 DOA and remained stable with advancing age. However, HAS rats did not differ significantly from CAS rats until 40 DOA, after which the magnitude of the difference continued to increase with age. In Experiment 2, rats were treated with alcohol at 25, 70, or 180 DOA. Rats at 70 or 180 DOA required less ethanol to disrupt their motor coordination on a rotating dowel (rotarod). Blood ethanol levels were determined at the loss and subsequent regain of the ability to negotiate the rotarod. Total duration of inability to negotiate the rotarod also was recorded. HAS rats were less able to remain on a rotarod while under the influence of alcohol relative to LAS and CAS rats regardless of age. However, no evidence of acute tolerance was observed in this experiment and, in fact, there was evidence of reverse tolerance in that all animals had lower BEC values at regain of ability than they did at loss.

Neurotensin levels in specific brain regions and hypnotic sensitivity to ethanol and pentobarbital as a function of time after haloperidol administration in selectively bred rat lines.

Evidence indicates that sensitivity to ethanol is a good predictor of the development of alcoholism. Thus, identification of neuronal processes that regulate ethanol sensitivity has been the subject of much recent research. The present studies were designed to further test the hypothesis that neurotensinergic processes mediate, in part, hypnotic sensitivity to ethanol. Single doses of haloperidol were administered to lines of rats [selectively bred for high and low sensitivity (HAS and LAS, respectively) to hypnotic effects of ethanol] to produce increases in neurotensin (NT) levels in brain regions. At 20 h after administration, haloperidol produced dose-dependent increases in NT immunoreactivity levels in nucleus accumbens (NA) and caudate putamen (CP) in both HAS and LAS lines. Levels of NT in NA and CP returned to control values at 48 h after 4 mg/kg haloperidol. These studies used two measures of hypnotic sensitivity to ethanol: duration of loss of righting reflex (sleep time) and blood ethanol concentration at regain of righting reflex (BECRR). At 20 h, but not 48 h, after haloperidol treatment, both HAS and LAS rats displayed increases in ethanol-induced sleep time with concomitant decreases in BECRR. Pentobarbital-induced sleep time was not increased 20 h after administration of 4 mg/kg haloperidol; however, hypnotic sensitivity to both pentobarbital and ethanol was increased by acute (30-min) pretreatment with 1 mg/kg. These results suggest that NT levels in NA, acting via NT receptors, enhance hypnotic sensitivity to ethanol, but not pentobarbital.

Relationship of brain ethanol metabolism to the hypnotic effect of ethanol. I: Studies in outbred animals.

BACKGROUND: This study was designed to investigate the relationship between the ethanol-oxidizing capacity of the brain, accumulation of acetaldehyde, and ethanol-induced hypnosis in animals in vivo. METHODS: Randomly outbred albino rats were treated with ethanol, and the duration of ethanol-induced loss of the righting response (sleep time) was measured. They were killed 2 weeks later (without further in vivo administration of ethanol), and brain homogenates were prepared to measure the accumulation of acetaldehyde from ethanol added in vitro. In a similar way, we determined the sleep time and, 5 days later, the rates of acetaldehyde accumulation in brains of heterogeneous mice. RESULTS: Significant correlations between the duration of ethanol-induced sleep and acetaldehyde accumulation in vitro were found. The Km value of the process of acetaldehyde accumulation was lower in long-sleeping, as compared with short-sleeping, rats. A similar result was also obtained in genetically heterogeneous mice. Animals with a longer duration of ethanol-induced sleep had a higher level of the accumulation of ethanol-derived acetaldehyde in brain homogenates, as compared with the short-sleeping mice. Rats and mice with the intermediate duration of ethanol-induced sleep had an intermediate value of acetaldehyde accumulation in brain homogenates. There was no correlation between brain catalase activity and ethanol-induced loss of the righting response in either the rats or the mice. CONCLUSIONS: This study is a direct demonstration of the positive correlation between ethanol-derived acetaldehyde accumulation in vitro in the brain and a central (behavioral) effect of alcohol in outbred rats and mice in vivo.

Relationship of brain ethanol metabolism to the hypnotic effect of ethanol. II: Studies in selectively bred rats and mice.

BACKGROUND: To clarify the role of brain acetaldehyde in the hypnotic effect of ethanol, we compared the ethanol-oxidizing capacity (rate of acetaldehyde accumulation) and catalase and aldehyde dehydrogenase activity in the brains of animals genetically selected for different sensitivities to the hypnotic effect of ethanol. METHODS: We used high, low, or control alcohol-sensitive rats (HAS, LAS, and CAS) and short- and long-sleep mice (SS and LS), as well as SS x LS recombinant inbred mice with known strain differences in mean duration of ethanol-induced sleep. We studied the rate of accumulation of acetaldehyde from ethanol in brain homogenates of these animals and correlated those values with their hypnotic sensitivity to ethanol. RESULTS: Acetaldehyde accumulation from ethanol was significantly higher in the brain homogenates from HAS rats and LS mice with high sensitivity to the hypnotic effect of ethanol in vivo, compared with LAS rats and SS mice with low sensitivity to ethanol. A correlation was found between the duration of ethanol-induced sleep and the in vitro rate of accumulation of ethanol-derived acetaldehyde in the brains of recombinant SS x LS mice strains. There was no correlation of sleep time with brain catalase levels. There were no line differences in brain catalase or aldehyde dehydrogenase or in alcohol or aldehyde dehydrogenase activity in livers of LAS, CAS, and HAS rats or in SS and LS mice. CONCLUSIONS: A correlation between the brain acetaldehyde accumulation, but not catalase levels, and the central effect of ethanol was demonstrated in animals genetically differing in initial sensitivity to the hypnotic effect of ethanol.

Thiamine status in liver and brain of rats genetically selected for different sensitivity to hypnotic effect of alcohol.

BACKGROUND: The mechanisms of the different sensitivity or resistance of animals and humans to alcohol are still not completely understood. For further biochemical characterization of animals genetically selected for high-alcohol sensitivity (HAS) and low-alcohol sensitivity (LAS) with the hypnotic effect of alcohol, the thiamine status and thiamine metabolizing enzymes in these animals have been studied. METHODS: We investigated thiamine diphosphate and thiamine triphosphate levels as well as the activity of thiamine-dependent enzyme, transketolase, and thiamine-metabolizing enzymes, thiamine kinase, and thiamine triphosphatase in the liver and brain of HAS, LAS, and CAS (control) rats by standard biochemical techniques. RESULTS: It was found that the activity of transketolase, and the level of the coenzyme form of thiamine, thiamine diphosphate (TDP), were significantly lower in HAS versus LAS rats. The activation of transketolase by the exogenous TDP (TDP-effect) was significantly higher in the liver and brain regions of HAS rats compared with LAS rats. The level of TDP in the liver and cerebellum of HAS rats was significantly lower compared with LAS rats. These results indicate a severe deficiency of TDP in HAS rats. HAS rats have a significantly lower activity of thiamine triphosphatase, the additional source of TDP. Accordingly, HAS rats have much higher thiamine triphosphate levels in the liver and brain, compared with LAS rats. There were no significant differences between groups with respect to the thiamine diphosphatase and thiamine kinase activity. Most of the above parameters had the intermediate values in CAS rats, compared with LAS and HAS rats. These data indicate the possible role of the thiamine phosphate esters and related enzymes in the mechanisms that bring about the differential sensitivity to the hypnotic effect of alcohol. CONCLUSIONS: HAS rats have the genetically mediated thiamine diphosphate deficiency and increased thiamine triphosphate levels, probably due to reduced activity of thiamine triphosphatase in the liver and brain, compared with LAS rats. It can be related with the higher initial sensitivity of HAS rats to hypnotic effect of ethanol.

Phenotypic and genotypic relationships between ethanol tolerance and sensitivity in mice selectively bred for initial sensitivity to ethanol (SS and LS) or development of acute tolerance (HAFT and LAFT).

BACKGROUND: Genetically based risk for development of alcoholism in humans seems to be related to initial sensitivity and/or acute tolerance to ethanol. The genetic basis for the development of tolerance has received less attention than other ethanol-related behaviors. We have selected lines of mice, according to genetics, which are differentially sensitive to the initial hypnotic effect of ethanol (Short Sleep and Long Sleep, SS and LS) and other lines that differentially develop acute functional tolerance to ethanol (High and Low Acute Functional Tolerance, HAFT and LAFT). We review reports of the relationship between initial sensitivity and two forms of tolerance as measured using different behavioral measures and different time scales. The goal of the study was to investigate alcohol tolerance as measured by different behavioral tests conducted over different time periods and relate these variables to hypnotic sensitivity. METHODS: We investigated the phenotypic and genotypic relationships between different measures of tolerance to ethanol in the SS and LS mice. We used two measures of tolerance: (a) The time an animal can remain on a stationary dowel or roto-rod at 5-min intervals up to 30 minutes after a single low dose of ethanol (Acute Single Dose Tolerance, ASDT-dowel or ASDT-roto-rod); and (b) The difference in blood ethanol levels taken when a mouse could repeatedly regain balance on a stationary dowel or roto-rod after successive doses of ethanol (Acute Functional tolerance, AFT-dowel or AFT-roto-rod). The time course in AFT was much longer, up to 2 hours. We carried out the same studies on the High and Low Acute Functional Tolerance (HAFT and LAFT) mice. RESULTS: SS and LS mice differ in hypnotic sensitivity as measured by sleep time, and they differ in all forms of acute tolerance that were measured except in AFT-dowel. Although there were phenotypic correlations between AFT-dowel and ASDT-roto-rod in the Heterogeneous Stock (HS) of mice, provisional Quantitative Trait Loci (determined with Recombinant Inbred mice from a SS X LS cross) for the two phenotypes did not overlap, which indicated that there was little or no genetic correlation between the measures. HAFT and LAFT mice do not differ in hypnotic sensitivity as measured by sleep time measurements nor in ataxic sensitivity as measured on the dowel. The HAFT and LAFT mice both developed tolerance when tested in the 30-minute time frame, but the differences between the lines was largely in the rate of development of tolerance and not the amount developed. On the other hand, when tolerance was measured over 2 hr on the dowel or roto-rod, the HAFT and LAFT animals developed different levels of tolerance. CONCLUSIONS: We concluded that measures of tolerance depended on both the time of ethanol's action and the behavioral task used. It seemed that the measures of tolerance used in this study had different genetic bases in mice. Presumably, tolerance will also vary in humans depending on the behavioral measure, and tolerance will also have different genetic bases for the different behavioral measures in humans.

Selectively bred lines of mice show response and drug specificity for genetic regulation of acute functional tolerance to ethanol and pentobarbital.

Genetic regulation of acute tolerance to ethanol may be associated with ethanol consumption and other ethanol-related behaviors in rodents. We have used lines of mice, selectively bred for high and low acute functional tolerance (HAFT and LAFT, respectively) to ethanol-induced loss of balance to test this hypothesis. Replicate HAFT and LAFT lines differ in AFT to ethanol-induced loss of balance by 4.4- and 5-fold, respectively. Frequency distributions and mean AFT scores for those lines, F(1), and backcrosses show a dominance for the HAFT phenotype. Time courses for acquisition and decay showed that AFT to ethanol-induced loss of balance developed rapidly, could be maintained up to 6 h with repeated doses, and decayed 6 h after peak tolerance and discontinuance of ethanol administration. The lines did not differ in initial sensitivity as measured by brain ethanol concentration at loss of balance, indicating that initial sensitivity and AFT to loss of balance were not coselected traits. Surprisingly, HAFT versus LAFT lines did not differ in development of AFT to loss of righting response, or hypothermia, indicating different mechanisms or neuronal systems mediate genetic influences on these measures. Voluntary ethanol consumption was low in both of the replicate lines, but HAFT lines consumed greater amounts of ethanol than LAFT lines. The HAFT and LAFT lines developed AFT to pentobarbital-induced loss of balance, however, there were no line differences in rates or extent of the AFT development. These results show that genetic regulation of AFT development is drug- as well as response-specific.

An examination of ALDH2 genotypes, alcohol metabolism and the flushing response in Native Americans.

OBJECTIVE: The study was designed to examine the relationship between aldehyde dehydrogenase (ALDH2) genotype and the flushing response in a population of Native Americans. METHOD: Objective measures of the flushing response were obtained by monitoring skin temperature, heart rate, blood pressure, as well as blood alcohol concentrations, in flushing and nonflushing Native Americans (n = 105) as well as in Oriental (n = 15) and white (n = 15) control subjects following a dose of alcohol (0.2 or 0.4 gm/kg). ALDH genotypes were determined via polymerase chain reaction followed by hybridization to 32P or biotin-labeled allele-specific oligonucleotide probes. RESULTS: There were no ALDH2 mutations detectable in Native Americans reporting the flushing response, nor any objective evidence of an Oriental-like response to alcohol. The rate of alcohol metabolism was shown to be the same among whites, Native flushers and Native nonflushers. CONCLUSIONS: The results demonstrate that the flushing reaction experienced by Native Americans appears to be milder and less unpleasant than the "Oriental" flushing reaction, with little effect on drinking frequency and amount. In addition, the flushing is not mediated by the ALDH2 mutation or elevated blood acetaldehyde. A critical analysis of the discrepancies in the literature regarding alcohol metabolism in Native Americans is provided.

Distribution and kinetics of ethanol metabolism in rat brain.

It was found that the accumulation of acetaldehyde produced from 50 mM ethanol in rat brain homogenates takes place in all major brain regions. The velocity varied between 3.5 to 7.1 nmol/mg of protein/hr. The rate increased in the following order: brain hemispheres, striatum, brainstem, hypothalamus, and cerebellum. Significant regional differences in this process were found: in the initial period of incubation (5 min), acetaldehyde accumulation was maximal in the brain hemispheres; but, in the 30- to 60-min period, it became significantly higher in the cerebellum. Inhibition of this process by the catalase inhibitor, 3-amino-1,2,4-triazole (8 mM), was minimal in the brainstem (27%) and maximal (57%) in the cerebellum, despite nearly complete inhibition of catalase. This would indicate that processes other than catalase activity must contribute to acetaldehyde accumulation.

Role of GABA in the actions of ethanol in rats selectively bred for ethanol sensitivity.

Rats from the N/Nih heterogeneous stock have been selectively bred for high (HAS) or low (LAS) initial sensitivity to injected ethanol as measured by duration of the loss of the righting reflex. The selection for ethanol sensitivity in these lines apparently has reached a maximum. These lines are useful to elucidate the central nervous system mechanisms of the genetic differences between the lines and also provide clues to the mechanisms of ethanol's action. We have found that: 1) ethanol, etomidate, and ketamine but not propofol produce different sleep times and brain levels of the drug on awakening between these two lines; 2) only ethanol, etomidate, and ketamine produced significant differences between the HAS and LAS rats in GABA-mediated stimulation of chloride uptake into brain microsacs; 3) GABA, propofol, and etomidate decreased the Kd for flunitrazepam binding to whole-brain membranes but equally in both lines. Neither ethanol nor ketamine had an effect; 4) only GABA, ethanol, and etomidate increased the Kd for TBPS binding and only GABA decreased Bmax of TBPS binding. As with the previous selection for ethanol sensitivity in mice (short and long sleep) these lines of rats have very marked line differences in GABA-mediated events, and these are correlated with the sedative effects of ethanol. From these and previous studies we know that the major differences between selected lines of mice and rats are that the mouse lines are not differentially sensitive to halothane or pentobarbital while the rat lines are. However, the mouse lines are differentially sensitive to propofol and the rat lines are not. These data should be useful in dissecting the actions of ethanol at the GABA(A) receptor.

Acute tolerance to the ataxic effects of ethanol in short-sleep (SS) and long-sleep (LS) mice.

The objective of this series of studies was to examine the relationship between alcohol sensitivity and the development of very rapid acute tolerance to alcohol in mice. In order to measure acute tolerance to alcohol, a behavioral test was developed using a rotorod. In the first study, mice selectively bred for resistance (short sleep, SS) or sensitivity (long sleep, LS) to the acute hypnotic effects of ethanol were used, as well as mice from the base population (heterogeneous stock, HS). Mice were trained to run on the rotorod at a speed of 14 rpm to a criterion of 200 s, in four daily training sessions. On the test day, baseline measurements of rotorod performance were taken and mice were injected i.p. with alcohol in doses from 0 to 2.5 g/kg. Animals were tested at 1-min intervals for the first 5 min following injection, then at 5-min intervals for a total of 30 min. The results demonstrated that SS and HS mice developed tolerance within 10 min following the alcohol injections. LS mice did develop some acute tolerance, but at a much slower rate than the SS or HS mice. In the second study, the effects of intoxicated practice on the rates of acute tolerance development were examined in the SS, HS and LS mice at a dose of 2.0 g/kg alcohol. A total of ten groups of each strain were given a different number of practice trials (ranging from one to ten) on the rotorod prior to a final test session at 30 min post-injection. The results provide evidence that SS and HS mice are capable of developing acute tolerance independent of practice. That is, the group of animals injected at 0 time and tested ten times up to 30 min were no better at the 30-min time point than the group injected at 0 time and tested only once at 30 min. On the other hand, the LS mice showed a modest practice effect, developing additional tolerance to the ataxic effects of alcohol with increasing intoxicated practice. Overall, these studies demonstrated that mice can develop acute tolerance within minutes following alcohol exposure, and that this ability is correlated with the initial sensitivity to alcohol.

Phenobarbital sensitivity in HAS and LAS rats before and after chronic administration of ethanol.

Rats selectively bred for high alcohol sensitivity (HAS) or low alcohol sensitivity (LAS) were tested for initial sensitivity to hypnotic doses of ethanol and a locomotor-altering dose of phenobarbital. Following 6 weeks of either a pair-fed control or 33% ethanol-derived calorie diet, animals were tested again for tolerance to ethanol and cross-tolerance to phenobarbital. HAS and LAS rats did not differ in baseline open field or Rotarod activity before chronic ethanol treatment. However, HAS rats were more sensitive to 50 mg/ kg phenobarbital relative to LAS rats. Both control- and ethanol-diet rats appeared to be less sensitive to phenobarbital after the 6-week treatment period. Chronic ethanol-exposed HAS and LAS rats demonstrated tolerance to ethanol and cross-tolerance to phenobarbital, and in particular LAS rats were even more active in the open field following phenobarbital relative to controls. In summary, significant differences in response to phenobarbital were observed between HAS and LAS rats. These observations suggest that initial sensitivity and tolerance to ethanol are associated with differences in phenobarbital sensitivity and are influenced by similar genes.

Effect of exogenous GM1 on ethanol sensitivity in selectively bred mouse lines.

Ethanol sensitive long-sleep (LS) and ethanol resistant short-sleep (SS) mice are lines that have been genetically selected for differential central nervous system sensitivities to the hypnotic effect of ethanol. Because they were genetically selected only for differences in sensitivity to ethanol hypnosis, biochemical and physiological differences between them are likely related to their differential ethanol sensitivity. The synaptosomal and whole brain concentration of GM1 ganglioside was previously shown to differ significantly between the lines. Further, GM1 alters membrane responses to ethanol, including a differential effect on LS and SS synaptosomal membrane disordering. Therefore, GM1 was administered intracerebroventricularly (i.c.v.) with micro-osmotic pumps, to partially bypass the blood-brain barrier and to test its effect on CNS sensitivity to ethanol hypnosis in LS and SS mice. In the first experiment, 3 days' infusion of GM1 (20 micrograms/microliters, 24 microliters/day), saline control and treated LS and SS mice were tested for both regaining of the righting reflex and waking brain ethanol concentration. Incorporation of 3H-GM1 into brain membranes was verified by scintillation spectroscopy. GM1 did not alter ethanol sensitivity or brain ethanol concentration at time of waking in LS mice. Conversely, SS mice treated with GM1 were significantly more sensitive to ethanol hypnosis than saline controls as measured by the time to regain the righting reflex ("sleep time") and waking brain ethanol concentrations. In the second experiment, GM1-treated SS mice were again significantly more sensitive to ethanol hypnosis than saline controls. GM1 incorporation into the contralateral and ipsilateral cerebral hemispheres was determined by high-performance liquid chromatography.

A description of alcohol/drug use and family history of alcoholism among urban American Indians.

The patterns of alcohol consumption, family history of alcoholism, and lifetime and current diagnoses of substance dependence were determined in a sample of American Indians (n = 105) living in Denver. Subjects were recruited through flyers, posters, and advertisements placed in local newspapers, the Denver Indian Center, and Denver Indian Health and Family Services. Subjects were interviewed regarding their education, employment, past and present drug and alcohol use (including frequency/quantity, beverage type, and pattern of intake) and family history of alcoholism. The drug and alcohol sections of the Diagnostic Interview Schedule were administered in order to determine lifetime and current prevalence of substance dependence. Although there are limits to the generalizability of these data due to the use of a non-random sampling method, the results indicate that approximately half of the sample (50.5%) were abstinent or irregular drinkers with moderate intake (3.3 drinks/occasion). Binge drinkers (3.8%) consumed large amounts of alcohol per occasion, with a mean of 21.6 drinks. Also, 45.5% of the sample were regular drinkers (at least once/wk) with a mean of 11 standard drinks/occasion. The rate of current alcohol dependence (33.3%) and other drug dependence (18.1%) was relatively high with cocaine and cannabis the primary drugs of abuse. The most striking aspect of the sample was the very high rate of family history of alcoholism (60.6% with at least one alcoholic parent) and only 11.1% with no primary or secondary alcoholic family members.

Ethanol metabolism in the brain.

Acetaldehyde is suspected of being involved in the central mechanism of central nervous system depression and addiction to ethanol, but in contrast to ethanol, it can not penetrate easily from blood into the brain because of metabolic barriers. Therefore, the possibility of ethanol metabolism and acetaldehyde formation inside the brain has been one of the crucial questions in biomedical research of alcoholism. This article reviews the recent progress in this area and summarizes the evidence on the first stage of ethanol oxidation in the brain and the specific enzyme systems involved. The brain alcohol dehydrogenase and microsomal ethanol oxidizing systems, including cytochrome P450 II E1 and catalase are considered. Their physicochemical properties, the isoform composition, substrate specificity, the regional and subcellular distribution in CNS structures, their contribution to brain ethanol metabolism, induction under ethanol administration and the role in the neurochemical mechanisms of psychopharmacological and neurotoxic effects of ethanol are discussed. In addition, the nonoxidative pathway of ethanol metabolism with the formation of fatty acid ethyl esters and phosphatidylethanol in the brain is described.

Genetic selection and characterization of mouse lines for acute functional tolerance to ethanol.

Rapid adaptation to central nervous system inhibitory effects of ethanol is observed in animals and humans and this acute functional tolerance (AFT) is influenced by genotype in rodents. Studies have been conducted to identify neurochemical processes influencing AFT to ethanol, but little is known regarding genetic regulation of AFT or genetic influences on processes that mediate acquisition of AFT. Our study was designed to develop, by selective breeding, lines of mice that differ in acquisition of AFT to ethanol; such mouse lines will be valuable in identifying the neuroadaptive processes mediating AFT. AFT is defined as the difference in blood ethanol concentration (BEC) at regaining balance on a stationary dowel rod after two consecutive doses of ethanol, 1.75 followed by 2.0 g/kg. Starting with a genetically heterogeneous foundation stock (HS/lbg), seven generations of selective breeding has been completed for high (HAFT1), low (LAFT1), and control lines and four generations have been completed for the replicate HAFT2 and LAFT2 lines. The lines do not differ in initial sensitivity to ethanol; however, the means for AFT scores differ by 2.3- and 4.3-fold for females and males, respectively (106.5 vs. 46.5 mg ethanol/dl blood) for females and 106.2 vs. 24.8 mg/dl for males). Frequency distributions for HAFT1 and LAFT1 show only modest overlap in AFT scores. The lines differ in rates of acquisition of AFT, but not in rates of ethanol clearance. Heritabilities were 0.04 and 0.26 for HAFT1 and LAFT1 lines, respectively, indicating that the selection was asymmetrical. Evidence is provided indicating that practice during intoxication has little effect on acquisition of AFT in HAFT1 and LAFT1 lines.

Effect of administered ethanol on protein kinase C in human platelets.

There are numerous reports of the effect of ethanol on protein kinase C (PKC) in animals or with in vitro systems. However, the effect of ethanol on PKC in humans has not been extensively investigated despite the large number of studies involving PKC and human platelets. In this study, we administered ethanol to human volunteers and determined the level of PKC before and after a 0.4 g/kg dose of ethanol. We studied Native Americans and Caucasians of both sexes. There was an increases in PKC activity 60 min after ethanol administration. There were no ethnic, age, nor gender differences detected, nor was there any correlation between family history of alcoholism and the basal or stimulated platelet PKC levels. Neither was there any correlation of basal or stimulated PKC activity with the genotypes for ADH2, ADH3, ALDH2, CYP2E1, and CYP1A2.

Initiation of ethanol self-administration by the sucrose-substitution method with HAS and LAS rats.

This study was performed to examine ethanol self-administration in rats bred for different sensitivities to the sedative effects of alcohol [the Colorado High Alcohol Sensitive (HAS) and Low Alcohol Sensitive (LAS) rats]. Four rats from each replicate line of the HAS and LAS rats (n = 16) were obtained from the University of Colorado, and initiation to self-administer ethanol by the sucrose-substitution procedure was attempted. Before the initiation procedure was conducted, home-cage ethanol intake and preference ratio did not differ between LAS and HAS rats. During the initiation procedure, the LAS rats came to self-administer 10% ethanol (v/v) at similar levels as outbred Wistar rats initiated with the same procedure (approximately 0.4 g/kg/session). The HAS rats, however, failed to initiate (approximately 0.08 g/kg/ session after completing the sucrose-substitution procedure) and lever pressing was reduced even more in the HAS rats when the ethanol concentration presented was > 10% (v/v). Three of the eight HAS rats stopped lever pressing completely when the ethanol concentration was raised to 15%. After initiation, home-cage preference ratio differed significantly between the LAS and HAS rats (LAS > HAS, p < 0.03). That the LAS rats did not consume greater amounts of ethanol compared with outbred Long-Evans or Wistar rats is contrary to our hypothesis, based on recent human data suggesting that a lower sensitivity to ethanol could result in increased alcohol intake. The finding that the HAS rats could not be initiated, while selectively bred ethanol nonpreferring rats can, is also contrary to our hypothesis. Further studies related to ethanol self-administration with the HAS line could provide important information related to the genetics of alcohol nonacceptance.