Effects of prolonged ethanol consumption on hepatic lipids and hepatotoxicity in C57BL/6 female mice.

Mice consume high levels of alcohol for a short period of time resulting in increased toxicity and lethality. The effects of lower doses that could be consumed without death for prolonged periods were studied. The effects of moderate doses of ethanol on indices of lipid peroxidation (LP), liver lipid accumulation and hepatotoxicity were studied in C57BL/6 mice. Three groups of mice were fed diets in which ethanol provided 0, 25 or 30% of the total calories for 3, 7, 10 and 13 weeks. Increased hepatic cholesterol, phospholipid and triglycerides, indicative of changes in liver lipid metabolism; and increased levels of hepatic malondialdehyde, conjugated dienes, lipid fluorescence, serum alanine aminotransferase and minimal changes in liver architecture indicative of LP and liver damages, were observed in mice fed the alcoholic diets. Such increases were time and dose dependent. These results suggest that continuous ingestion of lower levels of dietary ethanol in mice produces biochemical and hepatotoxic responses which are indicative of the health risk often associated with high alcohol intake.     

Metabolic effects of ethanol on primary cell cultures of rat skeletal muscle.

Individuals who have consumed alcohol chronically accumulate glycogen in their skeletal muscles. Changes in the energy balance caused by alcohol consumption might lead to alcoholic myopathy. Experimental models used in the past, such as with skeletal muscle biopsy samples of alcohol-dependent individuals or in animal models, do not distinguish between direct effects and indirect effects (i.e., alterations to the nervous or endocrine system) of alcohol. In the current study, we evaluated the direct effect of ethanol on skeletal muscle glycogen concentrations and related glycolytic pathways. We measured the changes in metabolite concentrations and enzyme activities of carbohydrate metabolism in primary cell cultures of rat skeletal muscle exposed to ethanol for two periods. The concentrations of glycolytic metabolites and the activities of several enzymes that regulate glucose and glycogen metabolism were measured. After a short exposure to ethanol (6 h), glucose metabolism slowed. After 48 h of exposure, glycogen accumulation was observed.     

Liver fat and plasma ethanol are sharply lower in rats fed ethanol in conjunction with high carbohydrate compared with high fat diets

The effects of high fat and high carbohydrate diets on alcohol metabolism were studied on blood alcohol and liver fat concentration. In Experiment 1, rats consumed an alcohol-containing liquid diet. Blood was collected for ethanol, glucose and lactate analyses and livers were excised for lipid determination. Blood ethanol and liver fat were lower when rats consumed the high carbohydrate diet. Glucose concentrations were lower in rats fed the high fat diet compared with those fed the high carbohydrate diet when ethanol was consumed. In Experiment 2, rats consumed a high fat, ethanol-containing diet for 13 d. Half of the rats were switched to a high carbohydrate, ethanol-containing diet for an additional 11 d. The same analyses were carried out as for Experiment 1. Switching the high fat-fed rats to the high carbohydrate diet reversed the high blood ethanol and high liver fat values, even though the rats consumed significantly more alcohol with the high carbohydrate diet. In Experiment 3 the same high fat and high carbohydrate diets without ethanol were consumed for 2 wk, at which time ethanol was administered acutely, intraperitoneally, at 2 g/kg. Blood was analyzed for ethanol, glucose and lactate 30, 60 and 120 min after injection. Rats fed the high carbohydrate diet had lower blood ethanol but higher lactate at 120 min compared with those fed the high fat diet. The results suggest that the rate of ethanol elimination is slower in rats fed high fat than in those fed high carbohydrate diets, resulting in elevated blood ethanol and liver fat levels for the former.     

Ethanol decreases progesterone synthesis in human placental cells: mechanism of ethanol effect.

Fetal alcohol syndrome (FAS) is a set of signs and symptoms in offsprings born to mothers who abuse alcohol during pregnancy. We postulated that impairment in the placental endocrine function contribute to FAS. In this study, we examined in vitro effects of ethanol on the placental cells' (cytotrophoblast cells) capacity to synthesize progesterone. Cytotrophoblast cells were isolated from normal term placenta and were incubated (2 x 10(6)) with 20-, 30-, and 40-mM doses of ethanol for 6 h. Progesterone was measured in the incubate by RIA. The results showed that, at the 20-mM dose of ethanol, progesterone synthesis was significantly decreased (p less than 0.01), at the 30-mM dose level there was a further decrease of 20%. The differences between 30- and 40-mM ethanol dose levels were not significant. To determine the mechanism of ethanol effects on progesterone synthesis, cytotrophoblast cells were preincubated with 30 mM ethanol followed by 10 microliters of LDL (10 microliters LDL = 80 micrograms cholesterol) and vice versa. The results showed that ethanol effects on progesterone synthesis was dependent on whether ethanol was added prior to or following the addition of LDL in the medium. If ethanol was added in the medium prior to LDL, progesterone synthesis was decreased significantly (p greater than 0.01); however, when ethanol was added after the LDL, ethanol had no effect on progesterone synthesis. In the experiment where ethanol and LDL were added simultaneously in the medium, ethanol blunted the stimulatory effect of LDL on progesterone synthesis.(ABSTRACT TRUNCATED AT 250 WORDS)     

Opposite effects of acute and chronic administration of alcohol on gastric emptying and small bowel transit in rat

The effects of acute and chronic administration of a large dose of alcohol on gastric emptying and small bowel transit were studied in rats. The development of tolerance to the acute effect of alcohol on gastrointestinal motility during chronic alcohol administration was also investigated. Gastric emptying and small intestinal transit were assessed by the Phenol Red recovery method. Acutely, ethanol was given in a dose of 2.5 g/kg body wt by gavage 30 min before the test meal. Chronically, ethanol was administered by two different methods: (1) a dose of 2.5 g/kg body wt was administered by gavage daily for 10 days; (2) animals received 15% ethanol in their drinking water for 30 days. A single large dose of alcohol inhibited gastric emptying and small bowel transit. Treatment with a large dose of alcohol for 10 days did not change the gastric emptying significantly, but inhibited the small intestinal transit. Alcohol consumption in drinking water for 30 days accelerated gastric emptying and small bowel transit. Tolerance to the acute inhibitory effect of a single large dose of alcohol on gastrointestinal motility did not develop during chronic alcohol treatment.     

The effects of ethanol on blood toluene concentrations

The metabolism of toluene was found to be inhibited by a large dose of alcohol in seven volunteers exposed to 80 ppm of toluene under experimental conditions. When alcohol was taken during exposure, blood toluene concentrations were 42.5% greater on average than during exposure with no alcohol. This is probably due to competition for alcohol dehydrogenase which is required for the breakdown of both ethanol and toluene. In men exposed to toluene at work, it was found that blood toluene concentrations were lowest in those who drank regularly. These results suggest that continued alcohol intake increases the metabolism of toluene, presumably by inducing the microsomal oxidising enzyme system in the liver.     

Effect of chronic ethanol administration on disposition of ethanol and its metabolites in rat.

We studied the effects of chronic alcohol intake on the disposition of alcohol and its metabolites in the rat. We used male Wistar rats for all of the experiments in this study. Using a pair-feeding process, rats were fed a liquid diet containing alcohol or without alcohol for 6 weeks. Ethanol solutions (0.5, 1.0, 1.5, and 2.0 g/kg body weight [BW]) were administered as a bolus, intravenously. We then measured blood ethanol and acetate concentrations. Simultaneous multiline fitting was performed using mean blood alcohol concentration (BAC)-time curves fitted to the one-compartment open model with parallel first-order and Michaelis-Menten elimination kinetics. At low doses (0.5, 1.0, and 1.5 g/kgBW), no differences were observed between the alcohol group and the control group with respect to ethanol elimination rate, area under the curve of ethanol (AUC(EtOH)), and mean residence time of ethanol (MRT(EtOH)). At higher doses (2.0 g/kgBW), ethanol elimination rate in the alcohol group was significantly higher than in the control group (P<.5%). These findings were also substantiated by corresponding changes in AUC(EtOH) and MRT(EtOH). At low doses, no differences were observed between the alcohol group and the control group with respect to plateau concentration of acetate (AcT) (concentration of steady state=C(ss)AcT), area under the curve of AcT (AUC(AcT)), and mean residence time of AcT (MRT(AcT)). However, at higher doses, although there were no differences in C(ss)AcT, both AUC(AcT) and MRT(AcT) were significantly lower in the alcohol group when compared to the control group (P<.5%). Chronic alcohol consumption increases ethanol oxidation and AcT metabolism in rats, as observed at high blood alcohol concentrations (BACs). These effects were observed at BACs of 3.5-4.5 mg/ml, and were not observed at lower doses. Thus, with general alcohol consumption, interindividual differences and intra-individual changes in alcohol metabolism may not take into account increased or accelerated metabolism due to alcohol tolerance.     

Opposite effects of dietary saturated and unsaturated fatty acids on ethanol-pharmacokinetics,triglycerides and carnitines.

The objective of this study was to determine the effects of saturated fatty acid (SFA) and unsaturated fatty acid (UFA) diets on ethanol pharmacokinetics. Hepatic ADH and plasma carnitines were also evaluated as possible indicators of the mechanism involved.

Sprague-Dawley male rats were fed modified AIN76 diets containing 10% coconut oil (SFA) or corn oil (UFA) for 120 days. A single dose (3 g/kg bw) of ethanol (13% solution) was orally administered using a gastric canula on day 30, 90, 105 and 120. Tail vein blood samples were collected at various intervals following ethanol dose and were analyzed for blood-ethanol concentration (BEC). In an analogous trial rats were given these diets for 70 days and blood samples were collected on day 35 and 63 for triglycerides, cholesterol and carnitine determination. The animals were killed on day 70 to collect liver for ADH determination.

Compared to the UFA group, the SFA group exhibited significantly higher BEC, larger area under the curve, longer half-life of ethanol, and lower rates of ethanol elimination. Plasma carnitines were also higher in the SFA vs UFA group. However, hepatic ADH activity was not different between the groups.

Dietary SFA protects liver from alcohol injury by retarding ethanol metabolism, and carnitine may be involved.     

Effects of D1 and D2 dopamine receptor agents on ethanol consumption in the high-alcohol-drinking (HAD) line of rats

Dopamine receptor agonists and antagonists were tested for effects on alcohol drinking in female HAD rats (n = 10) given limited access (4 h/day) to a 10% (v/v) ethanol solution. Food and water were available ad libitum. Subcutaneous drug injections were given 30–60 min before the ethanol access periods. The D₂ agonist quinpirole (0.04–2.0 mg/kg) caused a dose-dependent decrease in alcohol drinking throughout the 4-h period. Spiperone, a D₂ antagonist, had no effect during the initial part of the session, but by the fourth hour, the 10 μg/kg dose tended to increase alcohol intake and the 30 μg/kg dose reduced intake. The D₁ antagonist SCH-23390 (3–30 μg/kg) dose-dependently decreased ethanol drinking during the first hour of access. The D₁ agonist SKF-38393 (2–6 mg/kg) also decreased alcohol intake, but it was less effective than SCH-23390. The findings implicate both D₁ and D₂ receptors in the reinforcing effects of alcohol drinking by the HAD line of rats.     

First pass metabolism of ethanol: an important determinant of blood levels after alcohol consumption

Alcohol consumption by rats fed ethanol-containing liquid diets is considerably greater than the measured rate of ethanol elimination from the blood, suggesting that a significant fraction of the alcohol ingested does not enter the systemic circulation. To assess the possibility of a first pass metabolism of ethanol, we compared the areas under the blood ethanol concentration curves after administration of various doses through various routes in alcohol-fed and control rats. In both groups, blood ethanol concentrations were significantly lower after intragastric than after intraportal or intravenous (femoral) administrations and, to a lesser extent, than after an intraduodenal dose. By contrast, the rise in blood acetate, a product of ethanol oxidation, was faster after intragastric administration. Moreover, absorption of the ethanol dose was virtually complete at the time of ethanol disappearance from the blood. The fraction of the dose that did not enter the systemic circulation was proportionally greater with the smaller doses. These results indicate that there is a significant first pass metabolism of ethanol which takes place in the gastrointestinal tract and particularly in the stomach, where alcohol dehydrogenase (ADH) activity is the highest. Chronic alcohol administration decreased ADH activity (probably secondary to gastric mucosal injury) and also decreased the magnitude of the first pass metabolism. The amount of ethanol ingested which does not enter the systemic circulation accounts for the apparent dissociation between alcohol consumption, blood ethanol levels and rate of blood ethanol elimination in alcohol-fed animals.     

Assessment of GABA-B, metabotropic glutamate, and opioid receptor involvement in an animal model of binge drinking

Drinking to intoxication or binge drinking is a hallmark characteristic of alcohol abuse. Although hard to model in rodents, the scheduled high alcohol consumption (SHAC) procedure generates high, stable ethanol intake and blood ethanol concentrations in mice to levels consistent with definitions of binge drinking. The purpose of the present studies was to determine the effects of pharmacological manipulation of the opioidergic, glutamatergic, and γ-aminobutyric acid (GABA)ergic systems on binge drinking with the SHAC procedure. Parallel manipulations were conducted in mice trained in operant self-administration of either sucrose or ethanol. For the SHAC procedure, genetically heterogeneous Withdrawal Seizure Control mice were given varying periods of fluid access, with a 30-min ethanol session every third day (total of seven). Mice were pretreated intraperitoneally with naltrexone (0, 0.6, or 1.25 mg/kg), baclofen (0, 2.5, or 5.0 mg/kg), or 2-methyl-6-(phenylethynyl)-pyridine (MPEP; 0, 3.0, or 10.0 mg/kg) before each ethanol session. For the operant self-administration procedure, separate groups of C57BL/6 mice were trained to complete a single response requirement (16 presses on the active lever) to gain 30 min of access to an ethanol or a sucrose solution. Mice received pretreatments of the same doses of naltrexone, MPEP, or baclofen before the self-administration sessions, with saline injections on intervening days. Naltrexone produced a dose-dependent decrease in binge drinking, and the highest dose also significantly decreased operant self-administration of ethanol and sucrose. Both doses of baclofen significantly decreased binge alcohol consumption, but the higher dose also tended to decrease water intake. The highest dose of baclofen also significantly decreased operant self-administration of sucrose. MPEP (10 mg/kg) significantly decreased binge alcohol consumption and sucrose self-administration. These results indicate that manipulation of the opioidergic, glutamatergic, and GABAergic systems significantly decreased binge drinking.     

Alcohol, the cerebral circulation and strokes

Inasmuch as ethanol is thought to exert its major effects on the CNS, it is important to determine whether this abused substance can exert any direct action on cerebral blood vessels. Since chronic ingestion of alcohol: (1) can produce a loss (and degeneration) of neurons and glial cells in the brain, and (2) is associated, often, with hallucinations in human subjects particularly those undergoing withdrawal, it is possible that ethanol could produce hypoxia in select regions of the brain. The available indirect evidence in man and animals, albeit equivocal, does indicate that ethanol in certain concentrations might produce deficits in cerebral blood flow in select regions of the brain. Direct in-situ observations on the rat brain, using high-resolution, quantitative TV image-intensification microscopy, indicates that administration of ethanol, irrespective of the route of administration (e.g., perivascularly, intraarterially or systemically), produces graded concentration-dependent spasms of arterioles and venules. Concentrations of ethanol approximately greater than 250 mg/dl produce intense spasms resulting in rupture of these vessels. Recent in-situ studies in conscious dogs, using radiolabelled microspheres, also indicate that ethanol can produce deficits in regional brain blood flow. Studies with isolated canine middle cerebral and basilar arteries clearly demonstrate that low concentrations of ethanol (e.g., (less than 10 mM) can produce concentration-dependent spasms by a direct vascular action. Collectively, these new findings could be used to support the concept that heavy use of alcohol or binge-drinking can produce stroke-like effects. Specific calcium antagonists prevented or reversed the alcohol-induced cerebrovasospasms in rats and may prove valuable in treating the hypertension and strokes observed in heavy users of alcohol.     

Responses to ethanol challenge in long- and short-sleep mice prenatally exposed to alcohol

Individual sensitivity to alcohol may influence the severity of functional deficits due to prenatal alcohol exposure. To examine this hypothesis, Long-Sleep (LS) and Short-Sleep (SS) mice, selectively bred for differences in ethanol-induced narcosis, were intubated with either 2.9 g/kg ethanol (E) or an isocaloric amount of sucrose (S) twice per day on days 7 through 15 of pregnancy. An untreated control group (C) was maintained for each line. Offspring were fostered to lactating Rockland-Swiss mice at birth. Males and females from each litter were challenged with an acute dose of ethanol (3.8 g/kg) at 30 days of age. Measures of sleep time duration, waking blood ethanol concentrations (BEC), rectal temperatures, heart rate, and ethanol clearance were obtained to examine whether the acute effects of ethanol are altered by prenatal alcohol exposure. Prenatal alcohol exposure did not differentially affect responses to ethanol challenge within either genotype. Ethanol-induced hypothermia, heart-rate depression, and sleep time did differ between genotypes, with LS more affected than SS mice. Ethanol clearance rates were faster for SS than LS mice. These results suggest postnatal pharmacological responses to acute ethanol challenge are not altered by prenatal alcohol exposure in LS and SS mice. Prenatal alcohol-exposed offspring of both mouse genotypes showed lower average heart rate responses than controls, suggesting this measure may be a sensitive indicator of prenatal alcohol effects in mice.     

Response to an ethanol challenge dose on sleep time and blood alcohol level in Wistar rats prenatally exposed to ethanol during gestational day 8

Pregnant albino rats were treated during the eighth day of gestation (GD 8), with two IP injections, spaced by 4 hours, of either ethanol (2.9 g/kg in 24% v/v saline solution) or saline. Maternal blood alcohol levels reached a peak of 457 mg/dl 60 min after the second dose. At the age of 45 days, an equal number of male and female offspring were injected with 3.5 g/kg ethanol and sleep time and blood ethanol levels were determined upon awakening. Ethanol metabolic rate was studied in other individuals injected with the same dose of ethanol and the slope of the linear descending portion of the curves was calculated. Animals that received ethanol in utero exhibited shorter sleep time and higher blood ethanol levels at the moment of awakening than controls. The rate of ethanol metabolism was similar in both groups. These results show that an acute intoxication with ethanol during GD 8 induced long-term changes in the CNS of offspring which caused reduced sensitivity to ethanol hypnotic effects.     

Ontogeny of ethanol elimination and ethanol-induced hypothermia.

Ontogeny of ethanol elimination rates and ethanol-induced hypothermia were examined as possible mechanisms contributing to the marked reduction in ethanol sensitivity early in life (Little et al., 1996; Silveri & Spear, 1998) and the notable gender difference in ethanol sleep-time seen in adult animals (Silveri & Spear, 1998). Elimination rates and brain/blood ethanol levels were determined following doses of 1.5 or 4.5 g/kg ethanol in male and female Sprague-Dawley rats at postnatal days (P)16, 26, 36, or 56. Animals were sacrificed at 40, 80, or 160 min post-injection, with ethanol elimination rates estimated from the slope of the regression of blood and brain alcohol levels across the three sampling periods. P16 animals exhibited the slowest rate of ethanol metabolism, while no gender effects were evident at any age. Observed ontogenetic increases in ethanol hypothermia were not systematically related to the ontogeny of ethanol metabolism. Factors other than ontogenetic changes in ethanol metabolism, hypothermia, or the distribution of ethanol between brain and blood must underlie the relative insensitivity to ethanol often reported in young and adolescent organisms, a fruitful area for future studies given the frequent use and misuse of alcohol by human adolescents.     

Wheat gluten-based diet retarded ethanol metabolism by altering alcohol dehydrogenase and not carnitine status in adult rats.

The objective of this study was to determine the effect of a lysine-deficient diet on carnitine status in adult rats and subsequently on ethanol metabolism. Adult male rats were fed either the AIN-76 diet (NS), the AIN-76 diet with wheat gluten (WG) replacing casein, the WG diet plus 0.8% L-lysine (LS), or the LS diet plus 0.5% L-carnitine (CS) for 30 days. On the 31st day the rats were given an oral dose of ethanol and blood-ethanol concentrations (BEC) were monitored for the next 8 hours. One week later the rats were given a second dose of ethanol and urine was collected until killed, 3 hours post-ethanol administration (PEA). Besides growth retardation and hypoproteinemia, BEC were significantly elevated in the WG group compared to the other group at hours 3-8 PEA. There were no significant differences in BEC between the LS and CS groups; however, their BEC were significantly higher than that of the NS group. The BEC were inversely related to liver alcohol dehydrogenase (ADH) activities which were significantly lower in WG, LS and CS groups than in the NS group. Plasma, liver and urine carnitine values were significantly higher in the CS group than in the NS, WG and LS groups, wherein the values were similar. It is concluded that the WG diet reduced ADH activity and attenuated ethanol metabolism without significantly altering blood, liver and urinary carnitines in the adult rat.     

Neonatal Alcohol Exposure Produces Hyperactivity in High-Alcohol-Sensitive But Not in Low-Alcohol-Sensitive Rats

Children of women who consume high amounts of alcohol during their pregnancies vary greatly in physical and behavioral outcomes. Although many factors, such as dose and timing of exposure, undoubtedly contribute to this variation, one important determinant may be genetic differences in the response to alcohol. The present study examined activity levels in high alcohol sensitivity (HAS) and low alcohol sensitivity (LAS) rats following neonatal alcohol exposure. These lines were selectively bred for extremes in ethanol-induced “sleep times.” The HAS and LAS offspring were exposed to alcohol via an artificial rearing procedure using the “pup-in-the-cup” technique. Rat pups were exposed to ethanol (6 g/kg/day) from postnatal day (PD) 4 through 7 and faded to a dose of 3 g/kg/day on PD 8 and 9. An artificially reared gastrostomy control group (GC) and a normally reared suckle control group (SC) were also included. Activity level was measured on PD 18 through PD 21 for 30 min daily in automated activity monitors. Neonatal ethanol exposure produced overactivity in HAS rats, relative to their controls, but the same ethanol treatment had no effect on the LAS rats. Importantly, there were no differences in blood alcohol concentrations (around 420 mg/dl) between the two lines during the treatment period. These data suggest that genetic differences in response to alcohol may be a predictor for some of the behavioral teratogenic effects of alcohol.     

Contribution of frontal cerebral blood flow measured by (99m)Tc-Bicisate spect and executive function deficits to predicting treatment outcome in alcohol-dependent patients

Aim: To determine whether inhibition and working memory deficits, and reduced regional cerebral blood flow (rCBF) (previously shown to be related), measured at the end of a detoxification programme, predict alcoholic relapse 2 months later. METHODS: Twenty uncomplicated alcoholic inpatients were investigated at the end of detoxification, at least 7 days since the last dose of diazepam, and a mean of 18.8 days since the last drink. Their performance was assessed on the inhibition (Hayling) test, working memory (Alpha-span task), episodic memory (California Verbal Learning Test) and abstract reasoning (Progressive Matrices). Frontal CBF was assessed at the same time with a semiquantitative (99m)Tc-Bicisate SPECT procedure. Patients were contacted 2 months later. Patients who abstained (n = 9) did not differ from those who relapsed (n = 11) on age, gender, smoking, duration of alcohol misuse, number of previous detoxifications, amount of ethanol consumed the month prior to admission to the detoxification programme, state anxiety, trait anxiety, or depression. RESULTS: Relapsed subjects had shown a lower uptake of (99m)Tc-Bicisate in the bilateral medial frontal gyrus (n = 9; mean ratio +/- SD = 0.69 +/- 0.006) than abstainers (n = 11; 0.85 +/- 0.19), and poorer performance on the Alpha-span task and the Hayling test. The other tests were not different. CONCLUSIONS: Inhibition and working memory deficits, associated with low levels of CBF in the medial frontal gyrus, are related to the difficulty of maintaining short-term abstinence from alcohol.     

Alcohol consumption and cerebral blood flow among older adults

A substantial epidemiological literature now supports the existence of a J or U-shaped association between alcohol consumption and a broad range cardiovascular health outcomes including stroke. Although it is well documented that alcoholics exhibit both global and regional cerebral hypoperfusion in the sober state, little is known regarding the effects of a broader range of alcohol consumption on cerebral blood flow (CBF). The present study employed positron emission tomography with H(2)(15)O to assess quantitative global and regional CBF in 86 participants (51 men and 35 women; mean age 60.1) as a function of self-reported weekly alcohol consumption (none, <1, 1 to <7, 7 to <15, and >15 drinks per week). Analyses controlling for age, gender, and vascular health (carotid intima-media thickness) revealed that, relative to the weighted population mean, global CBF was greater in the lightest alcohol consumption group (<1 per week) and lower in the heaviest (>15 per week). Effects did not vary across regions of interest. This report is the first to describe an inverted J-shaped relationship between alcohol consumption and CBF in the absence of stroke.     

Ethanol effects on GABA-gated current in a model of increased alpha4betadelta GABAA receptor expression depend on time course and preexposure to low concentrations of the drug.

Several recent studies have suggested that alphabetadelta subtypes of gamma-aminobutyric acid type A (GABAA) receptors (delta-GABAR) are a target for low dose ethanol (<30 mM). However, there are also conflicting reports suggesting that only high doses of the drug (100 mM) modulate these receptors. In addition, the studies which have demonstrated a clear effect of low dose ethanol on delta-GABAR find different effective concentrations for this effect. Here, we test the hypothesis that the apparent disparity in effective concentration is due to time-course effects when low (1-3 mM) dose ethanol is preapplied. To this end, we tested ethanol effects on native GABAR in CA1 hippocampus in a model of increased alpha4betadelta GABAR expression following 48h administration of the GABA-modulatory steroid THP (3alpha-OH-5beta-pregnan-20-one) to adult, female rats. GABA(EC20)-gated current was recorded with whole-cell patch clamp procedures from acutely isolated pyramidal cells. We assessed ethanol's effect on GABA-gated current using either (1) 2-5 min application of ethanol in increasing concentrations (0.1-30 mM) or (2) coadministration of ethanol with GABA. Two minute application of 1-3 mM ethanol produced optimal potentiation of GABA-gated current following steroid treatment, with higher concentrations less effective. In contrast, 30 mM ethanol produced optimal effects when ethanol was not preapplied. However, following preapplication of 1mM ethanol, 30 mM ethanol decreased the peak GABA-gated current. These findings suggest that ethanol may act at multiple interacting sites to affect GABAR efficacy and desensitization. These data also suggest that ethanol effects on GABA-gated current are affected by the time course of exposure and previous exposure to low concentrations of the drug.