Effects Of Chronic Ethanol Administration Research Articles

Ethanol and intestinal carcinogenesis in the rat

The effect of chronic ethanol administration on 1,2-dimethylhydrazine induced rectal carcinogenesis was investigated in 32 paired male Sprague-Dawley rats fed a nutritionally adequate liquid diet containing 36% of total calories either as ethanol or isocaloric carbohydrates. Chronic ethanol ingestion increased the total number of rectal tumors significantly (17 vs. 6, p<0.02), whereas no cocarcinogenic effect of ethanol was observed in other parts of the intestine. Alcohol did not influence tumor size or histopathology. A 47% increase in the activity of mucosal alcohol dehydrogenase in the distal colorectum was found between chronically ethanol fed and pair fed controls (0.241±0.019 vs. 0.164±0.020 μmol mg protein −1 hr −1, p<0.01). This could in part explain the cocarcinogenic effect of alcohol in this tissue. The data give experimental support to the epidemiologic findings of an increased incidence of rectal cancer in the alcoholic.

Effects of chronic ethanol intake on mobilization and excretion of cholesterol in baboons.

To investigate the effects of chronic ethanol administration on the mobilization and excretion of cholesterol, turnover and balance studies were carried out in baboons pair-fed cholesterol-free diets containing 50% of energy either as ethanol or as additional carbohydrate for several years. Ethanol feeding increased free cholesterol in all plasma lipoprotein fractions, and esterified cholesterol in very low density lipoprotein, intermediate density lipoprotein, and high density lipoprotein (HDL). The major increase occurred in HDL, mainly as esterified cholesterol. The latter was associated with decreased transfer of esterified cholesterol from HDL to low density lipoprotein. By contrast, the smaller increase in HDL-free cholesterol was associated with increased turnover in the plasma, increased splanchnic uptake, and increased fecal excretion of plasma cholesterol, mainly as neutral steroids. Cholesterol extraction predominated over release in the splanchnic vascular bed, suggesting that the excess of cholesterol excreted in the feces originated in extrasplanchnic tissues. Thus, these findings indicate that alcohol consumption favors mobilization of tissue free cholesterol for hepatic removal and excretion. By contrast the increase in HDL-cholesterol (mainly esterified) appears to be a poor indicator of cholesterol mobilization.

Effects of chronic ethanol administration and withdrawal on incorporation of arachidonate into membrane phospholipids

A plasma membrane fraction isolated from cerebral cortex of control and ethanol-treated rats was used to study the effects of chronic ethanol administration on uptake of arachidonate by membrane phospholipids. Upon incubation of the membranes with [(14)C] arachidonic acid in the presence of ATP, Mg(2+), and CoA, radioactivity was incorporated into all of the phospholipids, although a large proportion of the label was found in phosphatidylinositols (PI, 60%) and phosphatidylcholines (PC, 20%). Rats given ethanol (8-10 g/kg body wt) via intubation in the form of a liquid diet for 4 weeks showed an increase (17-20%) in arachidonate incorporation into PI and PC as compared to phosphatidylethanolamines (PE) and phosphatidylserines (PS). A similar increase in uptake activity was observed at 2 or 24 h upon withdrawal of ethanol, but uptake activity returned readily to that of control level by 72 h. The method described in this study is a sensitive and reliable procedure for monitoring the arachidonoyl turnover activity in neural membranes with respect to chronic ethanol induction and withdrawal.

Effect of chronic ethanol administration on production of and response to erythropoietin in the mouse.

Male adult mice were allowed to drink only a solution of 32% ethyl alcohol for 3 months. Hematocrit and hemoglobin concentration were lower in ethanol-treated than in control mice at the end of the experimental period. Red cell volume was not significantly different between both groups. Plasma volume was higher in experimental than in control mice. Therefore, the anemia found in ethanol-treated mice can be regarded as a dilution anemia. When ethanol-treated and control mice, both made polycythemic by hypertransfusion to suppress their endogenous erythropoietin formation, were injected with doses of erythropoietin in the range of 0.2 to 3.2 IRP units, the derived dose-response curves were markedly different because of a reduced response to the hormone by the treated mice. This finding suggests that the number of "erythropoietin-responsive cells" may be reduced as the result of ethanol, or that their response to the hormone may be delayed or inhibited. Plasma erythropoietin concentration in alcohol- treated mice, as determined in the posthypoxic polycythemic mouse bioassay, was higher than normal in both normoxic and hypoxic conditions, probably as the result of the impaired responsiveness to the hormone mentioned above.

Effects of Chronic Ethanol Feeding on Sympathetic Innervated Organs: Temporal Sequence of Biochemical, Functional, and Trophic Changes

In noradrenergic nerves as well as in effector organs of the sympathetic nervous system, chronic ethanol feeding induced biochemical, functional, and trophic changes. The time sequence of modifications in the activities of tyrosine hydroxylase, phenylethanolamine-N-methyl transferase, monoamine oxidase, endogenous norepinephrine levels, as well as the neuronal uptake mechanism and secretory responses to norepinephrine in the submaxillary gland of the rat, confirms that the effects of chronic ethanol administration are related to an increase in sympathetic tone.

Alterations in brain monoamine oxidase kinetics after chronic ethanol administration and withdrawal

Chronic ethanol administration to rats can produce a number of biochemical changes associated with brain neurotransmitter function (Ellingboe, 1978; Holman, 1983). For example, 5-HT* turnover is increased after 3 weeks of ethanol treatment, but 1 week after withdrawal this is reversed (Badawy et al., 1980). These findings are of interest as 5-HT has been implicated in ethanol self-administration (Rockman et al., 1982) and in depression, which may be similar to the mood disturbances seen in alcoholics (Coppen & Wood, 1982). In the brain, the major catabolic route for 5-HT is oxidative deamination by M A 0 [monoamine: 0, oxidoreductase (deaminating), EC 1.4.3.41, and in particular the A form of this enzyme (Youdim & Ashkenazi, 1982). Thus it is conceivable that changes in 5-HT metabolism are the result of alterations in the activity of MAO. In the present study, the effects of chronic ethanol administration and withdrawal on M A 0 kinetics in the brain stem (pons plus medulla oblongata), corpus striatum and hippocampus were examined. Male Sprague-Dawley rats (15@-180g) bred at NESCOT were housed three to a cage with free access to Labsure CRM diet [RHM (South) Ltd., Poole, Dorset, U.K.]. Ethanol was diluted in the drinking water to give a daily intake of about lOg/kg body wt. for 28 days (Badawy et al., 1980). At I days before the end of the experiment, half the ethanol group were withdrawn by replacing their drug solution with water. M A 0 was assayed at 37OC with air as the gas phase, by a modification of the meihod of Fowler & Tipton (1982). Each incubation contained supernatant (400-500 pg of protein), 13H]-5-HT (100-1OOop~) and 10mM-potassium phosphate buffer, pH7.2, in a final volume of lml. The reaction was stopped after 20min by the addition of 500 pl of 2 M-HCI, and the acid metabolite extracted into 4ml of ethyl acetate by vigorous shaking for 5 min. The radioactivity was counted at an efficiency of 48% in a Searle-Nuclear Chicago Mk I1 liquid-scintillation spectrometer. Under the assay conditions described, M A 0 activity was equivalent to the initial velocity, and most of the 3H-labelled product extracted with a single portion of ethyl acetate (i.e. 8 1 k 2%; n = 18). Vma. was unaltered after chronic ethanol treatment or withdrawal in any of the brain regions studied (Table 1). However, on withdrawal, K , was increased 2.5 times in the brain stem and corpus striatum, suggesting competitive inhibition (Table 1). Ethanol (100mM) in uitro had no effect on the enzyme, whereas the IC,, values for the MAO-A inhibitor

The effect of chronic ethanol administration on lipogenesis in liver and adipose tissue in the rat.

Rates of lipogenesis de novo have been studied in liver and epididymal fat pads of male rats chronically treated with ethanol. A solution of ethanol (150 ml/l) was administered as the only drinking fluid for 3 months with a standard solid diet; both food and drink were available ad lib. Lipogenesis in vivo was measured by the incorporation of tritiated water into lipid fractions: non-saponifiable lipid and fatty acids. Non-saponifiable lipid, both in liver and in adipose tissue, was unaffected by ethanol treatment. However, fatty acid synthesis de novo was significantly enhanced in both liver and adipose tissue, by 150 and 300% respectively. Plasma triacylglycerol and non-esterified fatty acid levels were unchanged and plasma glucose concentration slightly increased by ethanol administration. The rate of lipogenesis increased when insulin: glucagon increased twofold due to the effect of ethanol.

Tyrosine hydroxylase activity in the brain and adrenal gland of rats following chronic administration of ethanol.

The effects of chronic administration and ethanol withdrawal on the activity of tyrosine hydroxylase were examined in the adrenal gland and six brain regions, including the frontal cortex, hippocampus, locus coeruleus, striatum, substantia nigra, and hypothalamus. Ethanol was administered to rats by oral intubation every 8 hr for 48 hr in amounts based upon their state of intoxication. One hr after the last intubation, tyrosine hydroxylase activity was not significantly different from control values in either the adrenal gland or brain. Forty hr following the last ethanol intubation, tyrosine hydroxylase activity was significantly increased above control values in both the adrenal gland and locus coeruleus. Tyrosine hydroxylase activity in the remaining five brain areas was unaffected by ethanol treatment. Immunotitration studies indicate that the increases in tyrosine hydroxylase activity found in the adrenal gland and locus coeruleus 40 hr after ethanol administration were due to an increase in enzyme protein. These data indicate that high blood ethanol concentrations maintained over a period of time (48 hr) do not affect adrenal gland or brain tyrosine hydroxylase activity. However, withdrawal from ethanol following 48 hr of treatment does produce an increase in tyrosine hydroxylase activity in the adrenal gland and locus coeruleus, similar to that seen following other stresses.

Development and use of a rat albumin cDNA clone to evaluate the effect of chronic ethanol administration on hepatic protein synthesis.

A rat albumin cDNA probe (pBR alb 149) was developed in order to investigate the molecular mechanisms responsible for changes in hepatic protein synthesis after chronic administration of ethanol to rats. Rats fed a diet for up to 1 year in which 36% of calories were from ethanol, developed fatty livers but not cirrhosis. Cell-free protein synthesis with liver membrane-bound polysomes of ethanol-fed rats was increased as compared to control membrane-bound polysomes, whereas protein synthesis with free polysomes was unchanged. Total RNA extracted from liver membrane-bound polysomes and translated in a rabbit reticulocyte mRNA-dependent system showed a marked increase in albumin synthesis in the ethanol-fed group. Analysis of RNA molecules separated according to molecular weight by gel electrophoresis and hybridized with recombinant-cloned albumin cDNA demonstrated an increase in full-sized albumin mRNA species in ethanol-fed animals. Therefore, chronic ethanol administration appears to increase albumin synthesis by increasing the steady-state level of biologically active albumin mRNA in liver membrane-bound polysomes. Despite development of fatty liver, the protein synthesis machinery functions normally.

Effects of chronic ethanol administration on hepatic glycoprotein secretion in the rat

The effects of chronic ethanol feeding on protein and glycoprotein synthesis and secretion were studied in rat liver slices. Liver slices from rats fed ethanol for 4–5 wk showed a decreased ability to incorporate [14C]glucosamine into medium trichloracetic acidprecipitable proteins when compared to the pair-fed controls; however, the labeling of hepatocellular glycoproteins was unaffected by chronic ethanol treatment. Immunoprecipitation of radiolabeled secretory (serum) glycoproteins with antiserum against rat serum proteins showed a similar marked inhibition in the appearance of glucosamine-labeled proteins in the medium of slices from ethanol-fed rats. Minimal effects, however, were noted in the labeling of intracellular secretory glycoproteins. Protein synthesis, as determined by measuring [14C]leucine incorporation into medium and liver proteins, was decreased in liver slices from ethanol-fed rats as compared to the pair-fed controls. This was the case for both total proteins as well as immunoprecipitable secretory proteins, although the labeling of secretory proteins retained in the liver slices was reduced to a lesser extent than total radiolabeled hepatic proteins. When the terminal sugar, [14C]fucose, was employed as a precursor in order to more closely focus on the final steps of hepatic glycoprotein secretion, liver slices obtained from chronic ethanol-fed rats exhibited impaired secretion of fucose-labeled proteins into the medium. When ethanol (5 or 10 mM) was added to the incubation medium containing liver slices from the ethanol-fed rats, the alterations in protein and glycoprotein synthesis and secretion caused by the chronic ethanol treatment were further potentiated. The results of this study indicate that liver slices prepared from chronic ethanol-fed rats exhibit both impaired synthesis and secretion of proteins and glycoproteins, and these defects are further potentiated by acute ethanol administration.

Effect of chronic ethanol administration on bromobenzene liver toxicity in the rat

Female Sprague-Dawley rats were pair-fed a nutritionally adequate liquid diet containing either ethanol or isocaloric carbohydrate for 3 weeks. In vitro studies showed that chronic ethanol pretreatment preferentially increased the liver microsomal biotransformation of bromobenzene to p-bromophenol (via the toxic 3,4-epoxide) rather than to o-bromophenol (via the nontoxic 2,3-epoxide) and could thus potentiate bromobenzene hepatotoxicity. Bromobenzene administration (500 mg/kg body weight, ip), after an overnight fast, was associated in ethanol-pretreated rats with greater and accelerated liver glutathione depletion and subsequent decrease in liver cytochrome P-450 content than in controls. As assessed histologically and by determination of the rise in activities of serum enzyme markers of liver necrosis, chronic ethanol pretreatment, however, mainly resulted in earlier onset and resolution of bromobenzene-induced liver necrosis, with only a mild increase in the maximal severity of liver lesions. These results suggest that the twofold increase in liver microsomal bromobenzene 3,4-epoxidation by ethanol, being much less than that seen after phenobarbital pretreatment in our animal model and in that of others, is apparently not sufficient to markedly affect the severity of bromobenzene-induced liver toxicity.

Effects of Chronic Ethanol Administration and Cold Exposure on Rabbit Adrenocorticoid 11β-Hydroxylase and 18-Hydroxylase Activities

Effects of Chronic Ethanol Administration and Cold Exposure on Rabbit Adrenocorticoid 11β-Hydroxylase and 18-Hydroxylase Activities

Effects of chronic ethanol administration on the composition of membrane lipids in the mouse

The relative proportions of the phospholipid fatty acids of erythrocyte membranes in mice were changed by chronic ethanol treatment and were not related to effects of the drug on nutrition, body temperature or experimental stress. Similar changes were observed using two different routes of ethanol administration and they did not reflect the metabolic effects of ethanol seen in the phospholipid fatty acids of whole liver. The observed increased content of saturated fatty acids and decreased content of polyunsaturated acids support the concept of adaptive changes taking place in the membrane during tolerance development to compensate for an increased membrane fluidity caused by ethanol. However, an increased content of the mono-unsaturated acid, octadecenoic (oleic), was found and there was no change in the cholesterol/phospholipid ratio. Other contrasting types of plasma membrane in mice showed different patterns of change in their phospholipid fatty acids during chronic ethanol administration. It is suggested that changes in membrane lipid composition could only partly account for an adaptation to ethanol-induced membrane disordering.

Ethanol intoxication and withdrawal among three age groups of C57BL/6NNIA mice

Data have not been forthcoming on the effects of chronic ethanol administration on intoxication and severity of withdrawal using animals representative of the life-span of a particular species. The purpose of this study was to examine ethanol intoxication and withdrawal among three age groups (3, 14, 25 months) of C57BL/6NNIA male mice. Ethanol was administered in a liquid diet for 14 days. Pair-fed control groups and laboratory chow groups were also employed. Blood ethanol levels, signs of intoxication and withdrawal, liquid diet consumption, and body weight were measured. Old mice were significantly more intoxicated than younger mice. However, young mice consumed more ethanol as compared to the older mice. Blood ethanol levels did not differ among the three age groups, although variability was high within each age group. The ethanol liquid diet groups did not show a decrease in body weight. Withdrawal was more severe for old animals than younger animals. The greater effects of ethanol observed in the old animals do not appear to be attributable to age differences in blood ethanol levels, amount of ethanol consumed, or body weight loss.

Effect of ethanol on the neurotransmitters glutamate and GABA

The effect of chronic ethanol administration, 5 g/kg per day for 2 or 4 weeks, on the neurotransmitters glutamate and GABA was investigated in rats. An increase in GABA was found in the striatum, hippocampus, and substantia nigra, 8 or 12 h after the last ingestion of ethanol, this being masked by injection of barbiturate. In addition an increase of glutamate has been found in the examined brain areas. This is consistent with the known effect of ethanol on brain Dopa metabolism and release. Finally, the relevance of the results for the pathogenesis of withdrawal seizures and delirious states is discussed.

The effect of chronic ethanol administration of high affinity 3H-nicotinic binding in rat brain.

The effect of chronic ethanol administration on high affinity 3H-nicotinic binding rat brain was investigated. Chronic ethanol administration for 5 months included a significant decrease in the number of specific nicotinic binding sites in the hippocampus, whereas a significantly increase in the number of nicotinic binding sites both in the hypothalamus and in the thalamus was found. Similar effects were observed both after administration of a lower concentration (8% W/V) ethanol and a higher concentration (16% W/V) indicating that the duration of the administration seems to be of importance for the biochemical changes observed.

Differential effect of chronic ethanol administration on rates of protein synthesis on free and membrane-bound polysomes in vivo in rat liver during dependence development

Administration of ethanol thrice daily to rats in amounts sufficient to induce a high degree of physical dependence resulted in a 20% decrease in the rate of protein synthesis on liver membrane-bound polysomes in vivo after 3 days of treatment without affecting the rate on free polysomes. The inhibition was attributable to a decrease in the rate of polypeptide elongation as evidenced by comparable decreases in nascent chain synthesis and completed protein release without any change in leucine uptake by liver. Chronic ethanol treatment did not affect the quantity or distribution of free and membrane-bound polysomes, the DNA concentration, or the weight of liver. The inhibition of protein synthesis on membrane-bound polysomes cannot, therefore, be readily ascribed to ethanol-induced nutritional deficiencies or to some nonspecific toxic effect of ethanol.

Effect of chronic ethanol administration on gamma-glutamyltransferase activities in plasma and in hepatic plasma membranes of male and female rats.

The effects of chronic ethanol administration on the activities of gamma-glutamyltransferase (GGT) in plasma and in hepatic plasma membranes of male and female rats are studied. The effects of alcohol on the lipid level in plasma are also investigated. After 4 weeks of treatment, GGT activity significantly increases in plasma either in male rats (131%, p less than 0.02) or in female ones (64%, p less than 0.05). In addition, chronic alcohol consumption simultaneously increases beta-lipoprotein and triglyceride levels in plasma only in male rats (181%, p less than 0.05 and 171%, p less than 0.01, respectively). In the liver, a significant elevation of GGT activity is observed in plasma membranes (146% in male rats, p less than 0.02, and 84% in female rats, p less than 0.02) but neither in homogenates nor in microsomal fractions. So, the variation of enzymatic activity in plasma as well as in hepatic plasma membranes is higher in male than in female rats. These results demonstrate, as for phenobarbital, that alcohol provokes an induction of GGT in rat liver only in the plasma membrane fraction.

Stimulation of thymidine incorporation in isolated rat intestinal mucosal cells by feeding an ethanol-containing liquid diet.

The effect of chronic ethanol administration on the incorporation of 3H-thymidine into DNA of isolated intestinal cells in the rat was investigated. Chronic ethanol consumption increased significantly DNA synthesis in intestinal mucosal cells. Whether this enhancement of DNA synthesis as a measure for cellular regeneration is due to the local effect of ethanol via tissue injury, to a direct stimulatory effect of ethanol on enzyme activities involved in DNA metabolism or to a change in the intestinal cell population with a shift to immature crypt cells by ethanol is not known.

Effect of chronic ethanol administration on plasma levels of LH and the estrous cycle in the female rat.

Intragastric intubation of 4 or 8 g/kg of body weight of ethanol per day for 12 consecutive days disrupted the normal estrous cycle in female rats as monitored by vaginal epithelial cell changes. Plasma luteinizing hormone levels were unchanged suggesting that blood ethanol in the 50--100 mg/100 ml range acted as a direct gonadal toxin and not at the hypothalamic-pituitary gland level.