Effects of castration and testosterone administration on rat liver alcohol dehydrogenase activity

The effects of castration and testosterone administration on the activity of liver alcohol dehydrogenase and on the rate of ethanol elimination were determined in male Sprague-Dawley rats. Castration increased liver alcohol dehydrogenase activity. The total liver activity in castrated animals was 2.37 ± 0.229 (S.E.) $\text{mmoles}\text{hr}$ as compared with a value of 1.39 ± 0.125 $\text{mmoles}\text{hr}$ in sham-operated controls (P < 0.01). Testosterone administration partially suppressed the enhanced activity of liver alcohol dehydrogenase produced by castration. By contrast, in control animals testosterone administration resulted in a small paradoxical increase in liver alcohol dehydrogenase. The increase in the enzyme activity in castrated animals was associated with a parallel increase in the rate of ethanol elimination. Castrated and control animals showed decreases in free cytosolic and mitochondrial NAD⁺/NADH ratios after ethanol administration. These observations suggest that testosterone (and probably other as yet unknown factors modified by castration) affects liver alcohol dehydrogenase activity, and that the total enzyme activity can be a principal limiting factor in ethanol elimination.     

Role of the pituitary and neonatal androgenic imprinting in the hormonal regulation of liver alcohol dehydrogenase activity

Liver alcohol dehydrogenase activity is increased by thyroidectomy, orchidectomy, or hypophysectomy. We investigated the mechanisms of these hormonal effects by examining the effects of testosterone, dexamethasone and thyroid hormone on liver alcohol dehydrogenase activity in hypophysectomized rats and in cultured hepatocytes, and the effect of administration of androgens to neonatal female rats. Testosterone did not lower alcohol dehydrogenase activity in hypophysectomized rats, whereas dexamethasone and thyroxine produced moderate decreases in activity. Triiodothyronine reduced alcohol dehydrogenase activity of cultured hepatocytes from male and hypothyroid female rats in a dose-dependent fashion, confirming that thyroid hormone had pituitary-independent effects on the enzyme activity. Dexamethasone was required for the expression of alcohol dehydrogenase activity in cultured cells, and it increased the enzyme activity when present at supraphysiologic concentrations. Treatment of neonatal female rats with testosterone reduced the activity of the enzyme in adulthood. The difference in alcohol dehydrogenase activity in adult male and female rats appears to be determined in part by neonatal imprinting by androgens and in part by an effect of testosterone that is either mediated by or dependent upon the pituitary. Thyroid hormone reduces alcohol dehydrogenase activity by a direct effect on the liver.     

Action of metadoxine on isolated human and rat alcohol and aldehyde dehydrogenases. Effect on enzymes in chronic ethanol-fed rats

Metadoxine (pyridoxine-pyrrolidone carboxylate) has been reported to accelerate ethanol metabolism. In the present work we have investigated the effect of metadoxine on the activities of isolated alcohol and aldehyde dehydrogenases from rat and man, and on the activity of these enzymes in chronic ethanol-fed rats. Our results indicate that in vitro metadoxine does not activate any of the enzymatic forms of alcohol dehydrogenase (classes I and II) or aldehyde dehydrogenase (low-Km and high-Km, cytosolic and mitochondrial). At concentrations higher than 0.1 mM, metadoxine inhibits rat class II alcohol dehydrogenase, although this would probably not affect the physiological ethanol metabolism. Chronic ethanol intake for 5 weeks results in a 25% decrease of rat hepatic alcohol dehydrogenase (class I) activity as compared with the pair-fed controls. The simultaneous treatment with metadoxine prevents activity loss, suggesting that the positive effect of metadoxine on ethanol metabolism can be explained by the maintenance of normal levels of alcohol dehydrogenase during chronic ethanol intake. No specific effect of chronic exposure to ethanol or to metadoxine was detected on rat aldehyde dehydrogenase activity.     

EFFECTS OF DISULFIRAM AND THREE RELATED COMPOUNDS ON THE ACTIVITY OF ALCOHOL DEHYDROGENASE AND ALDEHYDE DEHYDROGENASE IN RAT LIVER AND GASTRIC MUCOSA

SUMMARY 1. Disulfiram or related compounds were administered to rats by stomach tube and the activities of alcohol dehydrogenase and aldehyde dehydrogenase in the liver and the gastric mucosa were determined 43 h later.
2. In liver, disulfiram did not affect alcohol dehydrogenase activity but reduced aldehyde dehydrogenase activity. The disulfiram metabolite diethyldithiocarbamate was without action.
3. In gastric mucosa, disulfiram had no effect on aldehyde dehydrogenase activity but enhanced alcohol dehydrogenase activity. Diethyldithiocarbamate had a similar effect.
4. The increase in alcohol dehydrogenase activity in the gastric mucosa might explain the hyperacetaldehydaemia produced by ethanol in disulfiram-treated patients.     

Depression of alcohol dehydrogenase activity in rat hepatocyte culture by dihydrotestosterone

Hepatocytes harvested from castrated rats retained a higher alcohol dehydrogenase (EC 1.1.1.1) activity than hepatocytes harvested from normal rats during 7 days of culture. Dihydro-testosterone (1 μM) decreased the enzyme activity, after 2 and 5 days of culture, in hepatocytes from castrated and control animals respectively. Dihydrotestosterone decreased the enzyme activity to similar values in both groups of hepatocytes by the end of 7 days of culture. Testosterone (1 μM) had no effect on the enzyme activity in normal hepatocytes and only a transitory effect in decreasing the enzyme activity in hepatocytes from castrated animals. The increases in alcohol dehydrogenase activity after castration and their suppression by dihydrotestosterone were associated with parallel changes in the rate of ethanol elimination. Additions of substrates of the malate-aspartate shuttle or dinitrophenol did not modify ethanol elimination. These observations indicate that dihydrotestosterone has a direct suppressant effect on hepatocyte alcohol dehydrogenase and that the enzyme activity is a major determinant of the rate of ethanol elimination.     

Effects of long-term ethanol treatment on aldehyde and alcohol dehydrogenase activities in rat liver.

Administration of intoxicating doses of ethanol by gavage for 3 weeks caused weight loss and reduced hepatic aldehyde dehydrogenase activity in the soluble, mitochondrial and microsomal fractions. Rats receiving equivalent amounts of ethanol as a constituent of a liquid diet for 5 weeks gained weight and showed no changes in aldehyde dehydrogenase activity. Alcohol dehydrogenase activity was decreased in the rats treated by gavage and unchanged in those given ethanol in the diet, but in spite of this the rate of ethanol elimination was accelerated in both groups. In the livers of two strains of rats genetically selected for their difference in voluntary alcohol consumption, the mitochondrial and microsomal aldehyde dehydrogenase activities had previously been shown to be significantly higher in the alcohol-consuming (AA) than in the alcohol-avoiding (ANA) rats. Similar differences were now found after long-term intragastric ethanol administration, although in both strains the absolute levels of aldehyde dehydrogenase were reduced. Profound reduction of mitochondrial low-K_m aldehyde dehydrogenase activity and high blood acetaldehyde were observed, especially in the ANA rats. This suggests a possible connection between the low activity of this enzyme and the increased acetaldehyde level.     

The effect of testosterone propionate on energy metabolism indices and the catecholamine content of the vascular wall

The effect of testosterone propionate (1 mg/kg) on the specific activity of lactate dehydrogenase, malate dehydrogenase, contents of pyruvic and lactic acids, levels of catecholamines in target tissues and non-target tissues of rats was studied. Deficit of androgens and their single compensation by administering testosterone propionate lead to an inhibition of energy formation, a decrease of enzyme activity and contents of metabolites in tissues of the seminal vesicles. The course administration of testosterone propionate increases the activity of malate dehydrogenase in the aorta and myocardium. Acetylsalicylic acid (40 mg/kg) induces an increase of noradrenaline levels in the heart and aorta along with an increase of androgen level in the organism.     

Voluntary ethanol drinking by the RAT: effects of 2-aminoethylisothiouronium Salt, a modifier of NAD:NADH and norelegnine, a beta-carboline derivative.

Voluntary intake of ethanol solution (ETOH) was decreased in rats administered 2-aminoethylisothiouronium bromide hydrobromide (AET), an agent reported to alter NAD:NADH ratios in rat liver. Repeated administration of same dose of AET to ETOH-naive rats produced a significant inhibition of liver aldehyde dehydrogenase. Ethanol intake was decreased in rats given noreleagnine (NLG), a beta-carbone derivative reported to inhibit monoamine oxidase. Repeated administration of NLG exerted a significant inhibitory effect on liver alcohol dehydrogenase activity. It is concluded that the observed reduction of ethanol under AET which inhibits liver aldehyde dehydrogenase may reflect an antabuse-like reaction and the reduction of ethanol intake under NLG may be due, in part, to a build-up of alcohol in the blood and brain through inhibition of ethanol metabolism. The results are discussed in reference to the possible mechanism of action underlying voluntary intake of ethanol in rats, implicating alteration of NAD:NADH ratios in the biochemical processes underlying alcohol intake of rats.     

Demonstration of a functional blood-testis barrier to acetaldehyde. Evidence for lack of acetaldehyde effect on ethanol-induced depression of testosterone in vivo

In vitro studies have shown that acetaldehyde is a more potent inhibitor of testicular steroidogenesis than ethanol. The present study examined the in vivo role of acetaldehyde in ethanol-induced reduction of testosterone by (1) determining the levels of acetaldehyde to which the testes were exposed subsequent to acute ethanol administration to mice; and (2) examining the effect of ethanol on testosterone in animals subsequent to drug pretreatment which decreased or increased ethanol-derived acetaldehyde. Ethanol-induced (3 g/kg) depression of testosterone was dependent upon gonadotropin stimulation. The increase in hCG-induced testosterone was suppressed (P less than 0.01) in ethanol- as compared to saline-treated animals [39.8 +/- 2.6 (S.E.M.) vs 28.1 +/- 2.3 ng/ml]. Pargyline (100 mg/kg) or cyanamide (8.4 mg/kg) increased (P less than 0.05) plasma and testicular acetaldehyde, while having no effect on the testosterone response to ethanol. Similarly, 4-methylpyrazole (25 mg/kg) reduced blood and testicular acetaldehyde to nondetectable levels, while having no effect on testosterone. Testicular acetaldehyde was lower (P less than 0.001) than plasma levels (14 +/- 2 vs 2.0 +/- 0.2 microM). This functional blood-testis barrier to acetaldehyde could be explained by testicular aldehyde dehydrogenases in the mitochondria (Km for acetaldehyde = 1.5 microM) and in the cytosol (Km = 123 microM) whose maximal activities totaled to more than 25-fold greater than that of testicular alcohol dehydrogenase (ADH). ADH was concentrated in the Leydig cells, while aldehyde dehydrogenase was evenly distributed in the testis. Ethanol prevented further hCG-induced rises in testosterone rather than inhibiting testosterone production to below pre-ethanol values. The above data argue against a significant role of acetaldehyde in the in vivo response of testosterone to ethanol. Ethanol appears to impair gonadotropin-testicular receptor interaction in vivo.     

Effects of testosterone on neuromuscular transmission in rat isolated urinary bladder

Testosterone was examined for its effects on neuromuscular transmission in rat and shrew urinary bladder. In isolated preparations of detrusor muscle from sexually immature male rats (8-10 weeks old) at concentrations of 100-300 microM, it inhibited neuromuscular transmission in a concentration-dependent manner and it also inhibited responses to applied carbachol and diadenosine pentaphosphate (Ap(5)A, a P2X receptor agonist). Ethanol (at or above 38 mM), the solvent for testosterone, also caused significant inhibition of neurogenic contractions as well as carbachol- and Ap(5)A-induced contractions. In older, sexually mature male rats (over 16 weeks old), testosterone and ethanol had similar effects to those observed in the young male rat, although both were slightly less potent. In young virgin female rats (8-12 weeks old), testosterone and ethanol inhibited neuromuscular transmission; testosterone was approximately 1000 times more potent than in male rats, with a threshold concentration of 30 nM. In the insectivore, Suncus murinus, testosterone (0.1 microM-1 x mM) caused inhibition of neurogenic and chemogenic responses, but ethanol had no significant effect. Flutamide (50 microM), a genomic testosterone-receptor antagonist, did not inhibit any of the responses to testosterone. It is concluded that testosterone acts predominantly on a postjunctional nongenomic receptor to inhibit urinary bladder detrusor muscle contraction.     

Increased hepatic retinal dehydrogenase activity after phenobarbital and ethanol administration

Cytosolic retinal dehydrogenase (EC 1.2.1.36, retinal: NAD+ oxidoreductase) activity was measured by assessing the conversion of retinal to retinoic acid by HPLC. In vitro, acetaldehyde, chloral hydrate and disulfiram were found to be inhibitors, whereas 95% of the activity remained in the presence of cyanide or in the absence of oxygen. In rats, retinal dehydrogenase activity prevailed over that of retinal oxidase. By contrast, in deermice, 80% of retinal oxidation was due to the oxidase rather than the retinal dehydrogenase activity in a normal strain (ADH+) as well as in one lacking alcohol dehydrogenase (ADH-). In ADH- deermice, retinal oxidase activity was greater than in ADH+ animals. In vivo, in the rat, chronic ethanol administration resulted in a significant increase of the dehydrogenase activity in the liver, but not in other tissues. After phenobarbital administration, hepatic retinal dehydrogenase activity was increased 8-fold, but no extrahepatic induction was observed. Conversely, feeding rats with a diet devoid of the precursor for the substrate (retinal) by replacing retinyl acetate with an equivalent amount of retinoic acid resulted in decreased retinal dehydrogenase activity. Under conditions in which retinal dehydrogenase activity is rate-limiting for the metabolism of retinal to retinoic acid, its induction after phenobarbital or ethanol administration may contribute to hepatic vitamin A depletion.     

Effects of Phenobarbital and Ethanol on Rat Liver Aldehyde Dehydrogenases

The aldehyde dehydrogenases and the drug-metabolizing system (the mixed function oxidase) were studied in Wistar rats treated with ethanol and phenobarbital. Phenobarbital treatment had no effect on the microsomal aldehyde dehydrogenase activity, whereas the mitochondrial aldehyde dehydro-genase activity was slightly decreased and the activities of ethylmorphine demethylase, NADPH-cytochrome c reductase and the concentration of cytochrome P-450 were increased two to threefold. Ethanol treatment caused no changes in the activities of the aldehyde dehydrogenases, and the drug metabolizing system was only slightly affected. Very little aldehyde dehydrogenase activity was found in the cytosol from livers of either control, ethanol-treated or phenobarbital-treated rats. The results are discussed with particular reference to the oxidation of acetaldehyde and other aldehydes during ethanol metabolism.     

Effect of medroxyprogesterone acetate on steroid reductases in rat liver

Male and female rats were treated with medroxyprogesterone acetate (17 alpha-acetoxy-6 alpha-methyl-pregn-4-ene-3,20-dione, MPA) 600 mg/kg body weight i.p. daily for seven days. The steroid metabolizing enzymes in liver microsomes and liver homogenate were measured following MPA treatment and in control rats. The specific activity of NADPH- and NADH-5 alpha-reductase in female rats and of NADPH-5 alpha-reductase in male rats decreased by the treatment. NADP+- and NAD+-3-hydroxysteroid dehydrogenase activities were lower in female MPA rats if compared to untreated animals. In male rats only NADP+- and NAD+-3 beta-hydroxysteroid dehydrogenase activities (substrate 5 beta-dihydrotestosterone) were diminished by MPA administration. There was no effect on the cytoplasmatic 5 beta-reductases. Plasma concentrations of luteinising hormone (LH), testosterone and androstenedione were lowered by MPA treatment.     

Effect of a low-protein diet on acetaldehyde metabolism in rats

The effect of dietary changes on liver alcohol and aldehyde dehydrogenase activities as related to effects on ethanol and acetaldehyde metabolism was investigated. Feeding rats for 8 weeks on diets rich in carbohydrate or fat, but with normal protein content, induced minor changes relative to giving a balanced diet. A low-protein, high-carbohydrate diet (5 per cent and 80 per cent of calory content, respectively) caused a significant reduction of both alcohol and aldehyde dehydrogenase activities in the liver. The activity of the high-Km aldehyde dehydrogenase in the microsomal and soluble fractions appeared to be more reduced than that of the low-Km enzyme in the mitochondrial fraction. The tail blood acetaldehyde was significantly higher in rats on the protein deficient diet in spite of their reduced ethanol elimination rates. The results suggest that protein deficiency deranges acetaldehyde metabolism and may thus increase the possible contribution of acetaldehyde to the effects caused by ethanol metabolism.     

Testosterone reverses ethanol-induced deficit in spatial reference memory in castrated rats

The present study was designed to evaluate the effects of ethanol, testosterone and combination of ethanol and testosterone, on spatial reference memory and beta-endorphin (beta-EN) levels in castrated rats. Male Sprague-Dawley rats (120-150 g) were used in this study, Animals were castrated and ethanol, testosterone or combination of the drugs were administered to rats at 09:00 h. The drugs were administered after a training period of 5 days and spatial reference memory was evaluated for 7 days using the Morris water maze. One hour after the last injection, animals were sacrificed, their brains removed and dissected into cortex, hypothalamus, hippocampus and midbrain. The beta-EN levels in these brain regions were determined by radioimmunoassay. The time to find the platform (latency period) was significantly increased in ethanol-treated rats, indicating that ethanol induces deficit in spatial reference memory. On the other hand, testosterone administration improved spatial reference memory by significantly decreasing the latency period. In addition, there was a significant decrease in latency period in the animals treated with combination of ethanol and testosterone. Results also indicate that administration of ethanol resulted in a significant increase in beta-EN levels in the hippocampus and in the cortex while concurrent administration with testosterone abolished this increase. These findings clearly indicate that administration of testosterone did not only improve memory but also abolished the spatial memory deficit induced by ethanol in castrated rats.     

Castration and tolerance induces changes in the levels of the activity of acetylcholinesterase in the isolated vas deferens of the rat

Changes in the activity of acetylcholinesterase (AChE) of the isolated vas deferens from normal, castrated, morphine and ethanol-tolerant rats were studied. Three days after the termination of treatment with morphine and on the last day of treatment with ethanol, a significant inhibition of the activity of AChE was detected. This reduction in the enzymatic activity persisted in morphine-tolerant rats for 15 days, but not for 30 days, at which time the levels of AChE were determined to be normal. However, in ethanol-tolerant rats, there were no significant changes found at days 15 or 30. The activity of AChE was decreased significantly in castrated rats, but this effect was reversed by treatment with testosterone. During withdrawal from morphine or ethanol, the levels of AChE were significantly increased. The results indicate that morphine and ethanol may be inducing changes in the feedback mechanism which regulates the levels of AChE at post-synaptic sites, and these changes could play an important role in the development of tolerance to morphine and to ethanol.     

Inhibition of testosterone synthesis by ethanol and acetaldehyde

Ethanol administration decreases plasma testosterone levels acutely in man and animals. The specific mechanism responsible for this effect is, at present, unknown. We have examined the hypothesis that ethanol or acetaldehyde directly inhibits the enzymes required for testosterone synthesis from pregnenolone. Microsomes prepared from testes of normal adult rats were isolated and assayed, in the presence and absence of ethanol or acetaldehyde, for activity of 3β-hydroxy-Δ⁵-steroid dehy-drogenase (E.C.1.1.1.145)/3-oxosteroid Δ⁵–Δ⁴ isomerase (EC5.3.3.1) complex, testosterone 17β-dehydrogenase (ECl.1.1.64), and 17α-hydroxyprogesterone aldolase (EC4.1.2.30), three of the four enzymes required for the biosynthesis of testosterone from pregnenolone. Ethanol at levels commonly seen in the blood of chronic alcohol-ingesting men was shown to inhibit the activity of 17α-hydroxyprogesterone aldolase in a concentration-dependent manner. In addition, acetaldehyde was shown to act as an inhibitor of 17α-hydroxyprogesterone aldolase in the presence of androstenedione.     

Enzyme Activities of Myocardial Energy Metabolism during Prolonged Digoxin Treatment in Rats

Abstract The effect of prolonged digoxin treatment (1 mg/kg day for 8 days) on the activity levels of some enzymes of energy metabolism (phosphofructokinase, lactate dehydrogenase, citrate synthase, succinate dehydrogenase) in rat myocardium was studied. In the control animals receiving the solvent mixture (glycerol: ethanol:water in 1:1:1) a transient decrease in the lactate dehydrogenase and citrate synthase activity levels was observed. In the hearts of digoxin treated rats the level of activity of phosphofructokinase was permanently lowered by the fourth day and the level of activity of citrate synthase permanently increased after the first day of treatment. A transient increase in the activity level of succinate dehydrogenase in the myocardium of digoxin treated animals was seen between days 1 and 6. In this study a permanent decrease in phosphofructokinase and an increase in citrate synthase activity levels in rat heart muscle was noted during prolonged digoxin treatment.     

Enzyme activities of myocardial energy metabolism during prolonged digoxin treatment in rats.

The effect of prolonged digoxin treatment (1 mg/kg day for 8 days) on the activity levels of some enzymes of energy metabolism (phosphofructokinase, lactate dehydrogenase, citrate synthase, succinate dehydrogenase) in rat myocardium was studied. In the control animals receiving the solvent mixture (glycerol:ethanol:water in 1:1:1) a transient decrease in the lactate dehydrogenase and citrate synthase activity levels was observed. In the hearts of digoxin treated rats the level of activity of phosphofructokinase was permanently lowered by the fourth day and the level of activity of citrate synthase permanently increased after the first day of treatment. A transient increase in the activity level of succinate dehydrogenase in the myocardium of digoxin treated animals was seen between days 1 and 6. In this study a permanent decrease in phosphofructokinase and an increase in citrate synthase activity levels in rat heart muscle was noted during prolonged digoxin treatment.     

Pharmaceutical agents known to produce disulfiram-like reaction: effects on hepatic ethanol metabolism and brain monoamines

Several pharmaceutical agents produce ethanol intolerance, which is often depicted as disulfiram-like reaction. As in the case with disulfiram, the underlying mechanism is believed to be the accumulation of acetaldehyde in the blood, due to inhibition of the hepatic aldehyde dehydrogenases. In the present study, chloramphenicol, furazolidone, metronidazole, and quinacrine, which are reported to produce a disulfiram-like reaction, as well as disulfiram, were administered to Wistar rats and the hepatic activities of alcohol and aldehyde dehydrogenases (1A1 and 2) were determined. The expression of aldehyde dehydrogenase 2 was further assessed by Western blot analysis, while the levels of brain monoamines were also analyzed. Finally, blood acetaldehyde was evaluated after ethanol administration in rats pretreated with disulfiram, chloramphenicol, or quinacrine. The activity of aldehyde dehydrogenase 2 was inhibited by disulfiram, chloramphenicol, and furazolidone, but not by metronidazole or quinacrine. In addition, although well known for metronidazole, quinacrine also did not increase blood acetaldehyde after ethanol administration. The protein expression of aldehyde dehydrogenase 2 was not affected at all. Interestingly, all substances used, except disulfiram, increased the levels of brain serotonin. According to our findings, metronidazole and quinacrine do not produce a typical disulfiram-like reaction, because they do not inhibit hepatic aldehyde dehydrogenase nor increase blood acetaldehyde. Moreover, all tested agents share the common property to enhance brain serotonin, whereas a respective effect of ethanol is well established. Therefore, the ethanol intolerance produced by these agents, either aldehyde dehydrogenase is inhibited or not, could be the result of a "toxic serotonin syndrome," as in the case of the concomitant use of serotonin-active medications.