Abatement by acetate of an ethanol withdrawal syndrome

Acetate, the main end product of ethanol metabolism in the liver and a substrate of the cerebral small-pool Krebs-cycle, was tested for its ability to abate an ethanol withdrawal syndrome. Male Sprague-Dawley derived rats were rendered physically dependent on ethanol by intragastric administration of ethanol at a dosage of 9 to 15 grams per kilogram per day over a 4-day period. Oral administration of acetate was effective in abating the tremulous component of the ethanol withdrawal syndrome.     

Suppression of an ethanol withdrawal syndrome in rats by 3-hydroxybutyrate

An ethanol withdrawal syndrome was elicited by withholding ethanol from physically dependent, male Sprague-Dawley rats. Ethanol dependence had been induced by intragastric administration of ethanol at a dosage of 9 to 15 grams per kilogram per day over a four-day period. Oral administration of 3-hydroxybutyrate, a compound which is elevated in blood of ethanol dependent rats and is a substrate of both the cerebral small-pool and large-pool Krebs-cycle, was effective in suppressing the tremulous component of the ethanol withdrawal syndrome. 3-Hydroxybutyrate did not function as a central nervous system depressant at the dose levels employed.     

Organic hydroperoxide-dependent oxidation of ethanol by microsomes: lack of a role for free hydroxyl radicals

Organic hydroperoxides can replace NADPH in supporting the oxidation of ethanol by liver microsomes. Experiments were carried out to evaluate the role of hydroxyl radicals in the organic hydroperoxide-catalyzed reaction. Maximum rates of ethanol oxidation occurred in the presence of either 0.5 mM cumene hydroperoxide or 2.5 mM t-butyl hydroperoxide and were linear for 2 to 4 min. The Km for ethanol was about 12 mM and Vmax was about 8 nmol ethanol oxidized/min/mg microsomal protein. Besides ethanol, the organic hydroperoxides supported the oxidation of longer-chain alcohols (1-butanol), and secondary alcohols (isopropanol). The organic hydroperoxide-supported oxidation of alcohols was not affected by several hydroxyl-radical scavengers such as dimethylsulfoxide, mannitol, or 2-keto-4-thiomethylbutyrate which blocked NADPH-dependent oxidation of alcohols by 50% or more. Iron-EDTA, which increases the production of hydroxyl radicals, increased the NADPH-dependent oxidation of ethanol, whereas desferrioxamine, which blocks the production of hydroxyl radicals, inhibited the NADPH-dependent oxidation of ethanol. Neither iron-EDTA nor desferrioxamine had any effect on the organic hydroperoxide-supported oxidation of ethanol. Cumene-and t-butyl hydroperoxide did not support microsomal oxidation of hydroxyl-radical scavengers. These results suggest that, in contrast to the NADPH-dependent oxidation of ethanol, free-hydroxyl radicals do not play a role in the organic hydroperoxide-dependent oxidation of ethanol by microsomes. Ethanol appears to be oxidized by two pathways in microsomes, one which is dependent on hydroxyl radicals, and the other which appears to be independent of these oxygen radicals.     

Genetic selection for ethanol withdrawal severity: Differences in replicate mouse lines

We report an ongoing within-family selective breeding project for the severity of handling-induced withdrawal seizures in mice made physically dependent on ethanol by inhalation. Two Withdrawal Seizure Prone (WSP) and two Withdrawal Seizure Resistant (WSR) lines have been subjected to five generations of selection, and two control (WSC) lines are maintained. Each WSP line had more severe and each WSR line had less severe withdrawal convulsions than its respective WSC line. Differences relative to control lines were more pronounced in the WSP lines and were not due to differences in effective dose of ethanol. Heritabilities were higher in the WSP lines than in the WSR lines. These lines will be useful for studying physiological determinants of ethanol dependence and withdrawal.     

The relation of abstinence analgesia to the development of cardiac disorders in the ethanol withdrawal syndrome in rats

Ethanol was administrated intragastrically (25%, w/v) to Wistar male rats. They received 7-10 g ethanol/kg b.wt. daily in 2 fractional doses for 6 days. In 20-24 hours after the last ethanol administration behavioral and neurological signs of withdrawal syndrome and pain latent period were measured. Analgesia was determined using the tail flick and hot plate tests. Two days later systolic function of the isolated perfusing heart and creatine phosphokinase outflow were examined. Rats had longer latent pain period than in control. Heart perfusion revealed a decrease of systolic pressure, dp/dt of systolic and diastolic pressure, increase of enzyme outflow. Kendall's correlation analysis revealed a positive relationship between intensity of withdrawal symptoms and analgesia index in the hot plate test (tau = +0.343, p. 0.01) and a lack of relationship in the tail flick test. There was negative relationship between the analgesia index and the indices of heart disorders. It is proposed that analgesia index can be used as a predictor of the cardiac muscle injury caused by the alcoholic abstinent syndrome.     

Profiling impaired hepatic endoplasmic reticulum glycosylation as a consequence of ethanol ingestion

Alcoholic liver disease (ALD) is a prominent cause of morbidity and mortality in the United States. Alterations in protein folding occur in numerous disease states, including ALD. The endoplasmic reticulum (ER) is the primary site of post-translational modifications (PTM) within the cell. Glycosylation, the most abundant PTM, affects protein stability, structure, localization, and activity. Decreases in hepatic glycosylation machinery have been observed in rodent models of ALD, but specific protein targets have not been identified. Utilizing two-dimensional gel electrophoresis and liquid chromatography-tandem mass spectrometry, glycoproteins were identified in hepatic microsomal fractions from control and ethanol-fed mice. This study reports for the first time a global decrease in ER glycosylation. Additionally, the identification of 30 glycoproteins within this fraction elucidates pathway-specific alterations in ALD impaired glycosylation. Among the identified proteins, triacylglycerol hydrolase (TGH) is positively affected by glycosylation, showing increased activity following the addition of sugar moieties. Impaired TGH activity is associated with increased cellular storage of lipids and provides a potential mechanism for the observed pathologies associated with ALD.     

Estrogen neuroprotection against the neurotoxic effects of ethanol withdrawal: potential mechanisms

Ethanol withdrawal (EW) produces substantial neurotoxic effects, whereas estrogen is neuroprotective. Given observations that both human and nonhuman female subjects often show less impairment following EW, it is reasonable to hypothesize that estrogens may protect females from the neurotoxic effects of ethanol. This article is based on the assumption that the behavioral deficits seen following EW are produced in part by neuronal death triggered by oxidative insults produced by EW. The EW leads to activation of protein kinase C, especially PKCepsilon, which subsequently triggers apoptotic downstream events such as phosphorylation of nuclear factor-kappaB (NFkappaB) complex. On phosphorylation, active NFkappaB translocates to the nucleus, binds to DNA, and activates caspases, which trigger DNA fragmentation and apoptosis. In contrast, estrogens are antioxidant, inhibit overexpression of PKCepsilon, and suppress expression of NFkappaB and caspases. Estrogen treatment reduces the behavioral deficits seen during EW and attenuates molecular signals of apoptosis. The effects of ethanol and estrogen on each step in the signaling cascade from ethanol exposure to apoptosis are reviewed, and potential mechanisms by which estrogen could produce neuronal protection against the neurotoxicity produced by EW are identified. These studies serve as a guide for continuing research into the mechanisms of the neuroprotective effects of estrogen during EW and for the development of potential estrogen-based treatments for male and female alcoholics.     

A rapid increase in lipoprotein (a) levels after ethanol withdrawal in alcoholic men.

Plasma concentrations of lipoprotein (a) (Lp(a)) were studied in 11 male alcoholics at the end of a drinking period and monitored during subsequent abstinence. Lp(a) levels showed a daily increase for four consecutive days after the beginning of abstinence, the values for the third and the fourth day being significantly higher than those of the first day (p less than 0.05 and p less than 0.01, respectively). The changes in Lp(a) showed no association with the changes in low density lipoprotein (LDL) and high density lipoprotein (HDL) cholesterol levels. In one alcoholic subject with a heterozygous form of familial hypercholesterolemia who was monitored for 11 days, the Lp(a) levels rose up to the fourth day and remained at a high level thereafter. These results suggest that ethanol ingestion may be associated with a lowering of Lp(a) levels, which may contribute to the delayed progression of atherosclerosis observed in alcohol drinkers. Ethanol intake may be added to the short list of factors that affect the quite stable, genetically determined Lp(a) concentrations in the plasma.     

Yohimbine-precipitated clonidine withdrawal: an experimental model of the antihypertensive drug withdrawal syndrome.

In this study, a model of the clonidine withdrawal syndrome in normotensive rats was used to investigate the mechanisms and sites of the cardiovascular responses associated with this withdrawal. Clonidine (400 micrograms.kg-1.day-1), an alpha 2-adrenergic receptor agonist, was administered to rats via indwelling osmotic minipumps for 7 days. Withdrawal was precipitated by an intravenous injection of the alpha 2-adrenergic receptor antagonist yohimbine under alpha-chloralose anaesthesia, and the blood pressure and heart rate responses were recorded. Yohimbine (0.25, 0.50, and 1.0 mg/kg i.v.) in clonidine-treated rats provoked an immediate rise in blood pressure and heart rate. Similar injections in saline-treated rats produced slight hypotension and modestly increased the heart rate. Intracerebroventricular (i.c.v.) yohimbine injection (30 or 120 micrograms/kg in 10 microL volume) failed to elicit signs of withdrawal in clonidine-treated animals, but a subsequent intravenous injection of yohimbine (0.5 mg/kg) provoked brisk signs of withdrawal. hexamethonium (2 mg/kg) pretreatment did not abolish the increase in the heart rate, but it delayed the blood pressure increase. Pretreatment with atropine sulfate (1 mg/kg) did not block the yohimbine-induced increase in heart rate or blood pressure. This study demonstrates that yohimbine can effectively produce cardiovascular signs of withdrawal in rats chronically exposed to clonidine. The lack of i.c.v. yohimbine suggests that the antagonist-precipitated withdrawal may not have a central origin.(ABSTRACT TRUNCATED AT 250 WORDS)     

Utilization of by-products from ethanol production as substrate for biogas production

The aims of this work were to determine the specific biogas yields of steam-exploded sugarcane straw and bagasse as well as to estimate their energy potential under Brazilian conditions. Steam-explosion was carried out under different time and temperature conditions. The specific biogas yields were analyzed in batch-tests according to VDI 4630.Results have shown that steam-explosion pre-treatment increased the specific biogas yields of straw and bagasse significantly compared to the untreated material. The utilization of these by-products can contribute to 5% of the total energy consumption and thereby higher energy independence in Brazil. Further efforts in defining the optimum pretreatment conditions with steam-explosion as well as implementing this technology in large scale plants should be made.     

The ultraviolet sensitivity of Cockayne syndrome cells is not a consequence of reduced cellular NAD content

Cells from individuals with Cockayne syndrome (CS) are hypersensitive to the lethal effects of ultraviolet light (uv) and show a number of abnormal biochemical responses following uv-irradiation. Fujiwara et al. recently reported that the NAD contents of CS fibroblasts were lower than those of normal fibroblasts, and that addition of NAD to the cellular growth medium rectified most of the abnormal responses of CS cells to uv-irradiation. In our experiments, however, the cellular NAD contents of normal and CS fibroblasts were similar, and addition of NAD to the growth medium had no effect on the hypersensitivity of CS cells to uv-irradiation, nor did it restore the inability of CS cells to recover normal rates of DNA or RNA synthesis following uv-irradiation.     

The functional consequence of RhoA knockdown by RNA interference in rat cerebral arteries

Uridine triphosphate (UTP) constricts cerebral arteries by activating transduction pathways that increase cytosolic [Ca(2+)] and myofilament Ca(2+) sensitivity. The signaling proteins that comprise these pathways remain uncertain with recent studies implicating a role for several G proteins. To start clarifying which G proteins enable UTP-induced vasoconstriction, a small interfering RNA (siRNA) approach was developed to knock down specified targets in rat cerebral arteries. siRNA directed against G(q) and RhoA was introduced into isolated cerebral arteries using reverse permeabilization. Following a defined period of organ culture, arteries were assayed for contractile function, mRNA levels, and protein expression. Targeted siRNA reduced RhoA or G(q) mRNA expression by 60-70%, which correlated with a reduction in RhoA but not G(q) protein expression. UTP-induced constriction was abolished in RhoA-depleted arteries, but this was not due to a reduction in myosin light chain phosphorylation. UTP-induced actin polymerization was attenuated in RhoA-depleted arteries, which would explain the loss of agonist-induced constriction. In summary, this study illustrates that siRNA approaches can be effectively used on intact arteries to induce targeted knockdown given that the protein turnover rate is sufficiently high. It also demonstrates that the principal role of RhoA in agonist-induced constriction is to facilitate the formation of F-actin, the physical structure to which phosphorylated myosin binds to elicit arterial constriction.     

Down's syndrome: a pathology involving the lack of balance of reactive oxygen species

Mongolism, or Down's syndrome (DS) is an inborn pathology due to trisomy 21. Apart from mental deficiency and anatomical abnormalities, premature aging is characteristic for this disease. This paper mainly reviews data on the generation of reactive oxygen species (ROS) and antioxidative defense mechanisms in DS, and on the sensitivity of DS cells to model sources of ROS. Changes in the antioxidative defense of DS cells are due mainly to the location of the gene for (Cu,Zn)-superoxide dismutase on chromosome 21. It is suggested that a disturbance of balance of ROS may be a key point in the pathogenesis of DS.     

The superior colliculus, a neuronal network resistant to the Gaba withdrawal syndrome

Chronic intracortical perfusion of GABA (Gamma Amino Butyric Acid) and its subsequent withdrawal generates the GABA withdrawal syndrome (GWS). This particular epileptic model has been observed in the motor cortex of monkeys and rats. Our purpose was to study the GWS in the motor cortex (MC), dorsal hippocampus (DH), and superior colliculus (SC). Thirty chronically-implanted adult Wistar rats were separated into 3 groups of 10 (8 experimental and 2 controls). The first group received GABA in MC, the second in the DH and the third in the SC. GABA was released in doses of 10 to 60 micrograms/microliter/h for 6 days employing osmotic mini-pumps. Two control rats per group received saline solution in the above-mentioned structures. Rats perfused in the MC showed GWS after interruption of the GABA flow. The group perfused in the DH showed paroxysmal discharges and epileptic seizures during perfusion. They also later showed GWS. No epileptic effects were observed in the SC-perfused group during either the GABA perfusion or during withdrawal. None of the six control animals showed epileptic effects. Our results show that the SC offers a strong resistance to GWS. This could be explained by the particular neuronal network structure of rat SC.     

Mobile D-loops are a preferred substrate for the Bloom's syndrome helicase

The Bloom's syndrome helicase, BLM, is a member of the highly conserved RecQ family, and possesses both DNA unwinding and DNA strand annealing activities. BLM also promotes branch migration of Holliday junctions. One role for BLM is to act in conjunction with topoisomerase IIIα to process homologous recombination (HR) intermediates containing a double Holliday junction by a process termed dissolution. However, several lines of evidence suggest that BLM may also act early in one or more of the recombination pathways to eliminate illegitimate or aberrantly paired DNA joint molecules. We have investigated whether BLM can disrupt DNA displacement loops (D-loops), which represent the initial strand invasion step of HR. We show that mobile D-loops created by the RecA recombinase are a highly preferred substrate for BLM with the invading strand being displaced from the duplex. We have identified structural features of the D-loop that determine the efficiency with which BLM promotes D-loop dissociation. We discuss these results in the context of models for the role of BLM as an ‘anti-recombinase’.