Intragastric self-infusion of ethanol by ethanol-preferring and -nonpreferring lines of rats.

An ethanol-preferring line of rats, developed by selective breeding, consumed as much as 9.4 +/- 1.7 grams of ethanol per kilogram of body weight per day through intragastric self-infusions, yielding blood ethanol concentrations of 92 to 415 milligrams per 100 milliliters. By contrast, the ethanol- nonpreferring line self-administered only 0.7 +/- 0.2 gram per kilogram per day. These findings indicate that the reinforcing effect of ethanol is postabsorptive and is not mediated by the drug's smell or taste. Hence the ethanol-preferring line of rats may be suitable animal model of alcoholism.

Dependence-induced alcohol drinking by alcohol-preferring (P) rats and outbred Wistar rats.

BACKGROUND: Chronic intermittent alcohol vapor exposure and selective breeding procedures have been used separately for many years to model specific aspects of alcohol dependence. The purpose of the present investigation was to combine these 2 approaches by exposing alcohol-preferring (P) rats to chronic intermittent alcohol vapor for extended periods of time and then testing them for operant alcohol responding in parallel with a group of outbred Wistar rats at multiple time points following the termination of vapor exposure. METHODS: P rats (n = 20) and Wistar rats (n = 18) were trained to respond for 10% (w/v) ethanol in an operant situation, then divided into groups matched for intake levels. Animals were then exposed to chronic intermittent alcohol vapor (14 hours ON/10 hours OFF) or air for 8 weeks. Rats were then tested for operant alcohol responding under various conditions and at multiple time points during alcohol withdrawal (6 hours) and protracted abstinence (1 to 15 days). RESULTS: Chronic alcohol vapor exposure produced similar increases in operant alcohol responding in P rats and Wistar rats during acute withdrawal and protracted abstinence. CONCLUSIONS: These results illustrate the separate and combined effects of genetic selection for high alcohol preference and dependence on alcohol drinking behavior. Furthermore, these results confirm past findings that dependent rats consume more alcohol than nondependent controls well into abstinence following extended periods of alcohol vapor exposure.

The role of acetaldehyde in mediating the deleterious effect of ethanol on pyridoxal 5'-phosphate metabolism.

Previous studies in vivo and with isolated perfused rat livers have suggested that the deleterious effect of ethanol on hepatic pyridoxal 5'-phosphate metabolism is mediated by acetaldehyde. Inasmuch as acetaldehyde has no effect on the synthesis of pyridoxal phosphate, it has also been postulated that acetaldehyde accelerates pyridoxal phosphate degradation by displacing this coenzyme from binding proteins, which protect it against hydrolysis. To test these hypotheses, studies have been performed with isolated rat hepatocytes, subcellular fractions of rat liver, and human erythrocytes. Ethanol oxidation lowered the pyridoxal phosphate content of isolated liver cells when acetaldehyde oxidation was inhibited by either disulfiram or prior treatment of rats with cyanamide. Additions of 7.5 mM acetaldehyde alone at 40-min intervals to cell suspensions decreased hepatic pyridoxal phosphate content only slightly because acetaldehyde was rapidly metabolized. However, when acetaldehyde oxidation and reduction were inhibited by cyanamide treatment and by 4-methyl-pyrazole and isobutyramide, respectively, a 40% decrease in hepatic pyridoxal phosphate content was observed in 80 min of incubation. In equilibrium dialysis experiments, acetaldehyde, 7.5 and 15 mM, displaced protein-bound pyridoxal phosphate in undialyzed hepatic cytosol and in hemolysate supernate containing added pyridoxal phosphate. In the presence of alkaline phosphatase, acetaldehyde accelerated the degradation of pyridoxal phosphate in dialyzed hemolysate supernate and hepatic cytosol with added pyridoxal phosphate. Acetaldehyde also inhibits tyrosine aminotransferase. The kinetics of inhibition were mixed competitive-noncompetitive with respect to pyridoxal phosphate. These observations support the hypothesis that the deleterious effect of ethanol oxidation on pyridoxal phosphate metabolism is mediated at least in part by acetaldehyde which displaces this coenzyme from protein binding, thereby enhancing its degradation.

Plasma content of B6 vitamers and its relationship to hepatic vitamin B6 metabolism.

The plasma content of B6 vitamers is governed by, among other factors, dietary supply and metabolic interconversion. This study examines the effect of pyridoxine supplementation on the plasma content of B6 vitamers and pyridoxic acid in man, and the metabolic conversion and release of B6 compounds in isolated rat hepatocytes. Six healthy human subjects were given 100 mg pyridoxine-HCl/d orally for 1--3 wk. Before pyridoxine supplementation, the mean total plasma level of B6 vitamers was 114 +/- 9 nM; and pyridoxal-P, pyridoxamine-P, pyridoxal, pyridoxine, and pyridoxamine accounted for 54, 3, 11, 27, and 5%, respectively. Plasma level of pyridoxic acid was 40 +/- 7 nM. Thus, pyridoxal-P is the principal B6 vitamer in plasma. During pyridoxine supplementation, mean plasma levels of the B6 vitamers and pyridoxic acid increased to 655 +/- 122 and 222 +/- 55 nM, respectively. The plasma content of pyridoxal-P and pyridoxic acid increased 6--7-fold and that of pyridoxal, 12-fold, but the pyridoxine level did not increase. Isolated hepatocytes, 1 g/15 ml, were incubated for 2 h with 3.33 microM [14C]pyridoxine (6 microCi/mumol). At zero time, the cells contained about 35 nmol pyridoxal-P and 25 nmol pyridoxamine-P. After 2 h incubation, the cellular content of pyridoxal-P and pyridoxamine-P did not change significantly, but the medium contained 5.9 nmol pyridoxal-P, 0.3 nmol pyridoxamine-P, 7.2 nmol pyridoxal, 26.6 nmol pyridoxine, 0.3 nmol pyridoxamine, and 7.5 nmol pyridoxic acid. Whereas the specific radioactivity of pyridoxal-P, pyridoxal, and pyridoxic acid in the medium approached that of [14C]pyridoxine, the specific radioactivity of cellular pyridoxal-P and pyridoxamine-P was only 20% of that of pyridoxine. Thus, newly synthesized pyridoxal-P is not freely exchangeable with endogenous pyridoxal-P, but is preferentially released or degraded to pyridoxal and pyridoxic acid. The latter B6 compounds are also released. These results suggest that orally ingested pyridoxine is rapidly metabolized in liver and its products are released into the circulation in the form of pyridoxal-P, pyridoxal, and pyridoxic acid.

Vitamin B6 metabolism in chronic alcohol abuse. Pyridoxal phosphate levels in plasma and the effects of acetaldehyde on pyridoxal phosphate synthesis and degradation in human erythrocytes.

The plasma pyridoxal-5'-phosphate (PLP) level of alcoholic subjects has been compared with that of non-alcoholic individuals in order to ascertain the incidence of abnormal vitamin B(6) metabolism in chronic alcohol abuse. 66 alcoholic subjects were selected on the basis that they did not exhibit abnormal liver function tests and hematologic findings. 35 of them had plasma PLP concentrations less than 5 ng/ml, the lowest value encountered in 94 control subjects, indicating a high incidence of deranged PLP metabolism in alcoholic patients even when hepatic and hematologic abnormalities are absent. The biochemical basis for the altered PLP metabolism in chronic alcohol abuse was examined. Low plasma PLP levels in alcoholics were not accompanied by decreased pyridoxal kinase and pyridoxine phosphate oxidase activities in erythrocytes. Further studies with erythrocytes demonstrated that the cellular content of PLP is determined not only by the activities of these PLP-synthesizing enzymes but also by the activity of a phosphate-sensitive, membrane-associated, neutral phosphatase, which hydrolyzes phosphorylated B(6) compounds.Acetaldehyde, but not ethanol, impaired the net formation of PLP from pyridoxal, pyridoxine, and pyridoxine phosphate by erythrocytes. However, when the B(6)-phosphate phosphatase activity was inhibited by 80 mM phosphate, this effect of acetaldehyde was abolished. By the use of broken cell preparations, it was possible to demonstrate directly that the action of acetaldehyde is mediated by the phosphatase, resulting in an acceleration of the degradation of the phosphorylated B(6) compounds in erythrocytes.

Detection of a protein-acetaldehyde adduct in the liver of rats fed alcohol chronically.

We report here the formation in vivo of a protein-acetaldehyde adduct (protein-AA) in liver when rats were fed alcohol chronically. This chemically modified protein was demonstrated by electroimmunotransblot technique and with rabbit polyclonal antibodies that recognize acetaldehyde adduct as an epitope (i.e., both anti-hemocyanin-AA IgG and anti-myoglobin-AA IgG). It has a molecular weight of 37,000. It can be detected in the liver of rats fed the alcohol-containing American Institute of Nutrition 1976 liquid diet for only 1 wk. Since the protein profiles of soluble hepatic proteins from alcohol-fed and control rats were identical on SDS-PAGE, the peroxidase-positive band demonstrated by electroimmunotransblot was most likely not a new protein synthesized de novo. Borohydride reduction was not necessary to stabilize this protein-AA. Intraperitoneal injections of ethanol (2 g/kg body wt) at 8-h intervals to rats over a 24-h period did not produce any detectable protein-AA in the liver. Incubation of the liver homogenate from a control liver with acetaldehyde without sodium cyanoborohydride for 4 h also failed to generate any protein-AA. Therefore, the formation of the 37-kD protein-AA in vivo reported here is dependent on chronic alcohol consumption.

Vitamin B6 metabolism in chronic alcohol abuse The effect of ethanol oxidation on hepatic pyridoxal 5'-phosphate metabolism.

Individuals with chronic alcohol abuse frequently exhibit lowered plasma levels of pyridoxal 5'-phosphate, the coenzyme form of vitamin B6. Because the liver is the primary source of this coenzyme in plasma and also the principal organ that oxidizes ethanol, the effect of ethanol on hepatic pyridoxal phosphate metabolism was studied in the rat. The chronic feeding of ethanol (36 percent of the total dietary calories) for 6 wk significantly decreased the hepatic pyridoxal phosphate content both in animals given a sufficient amount of vitamin B6 in their diet and in those rendered vitamin B6 deficient. In isolated perfused livers, the addition of 18 mM ethanol lowered the pyridoxal phosphate content of livers from vitamin B6-sufficient animals and deceased the net synthesis of pyridoxal phosphate from pyridoxine by the livers of vitamin B6-deficient animals. Ethanol also diminished the rate of release of pyridoxal phosphate into the perfusate by the livers of vitamin B6-deficient rats. These effects of ethanol, in vitro, were abolished by 4-methyl pyrazole, an inhibitor of alcohol dehydrogenase. Thus the derangement of pyridoxal phosphate metabolism produced by ethanol is dependt upon its oxidation. These data support previous findings whic indicate that acetaldehyde is the responsible agent which acts by accelerating the degradation of intracellular pyridoxal phosphate.

Serum low density lipoprotein of alcoholic patients is chemically modified in vivo and induces apolipoprotein E synthesis by macrophages.

This work was carried out to investigate the effect of alcohol drinking on serum LDL. Agarose gel electrophoresis showed that LDL samples from alcoholic patients without serious liver disease were more negatively charged and moved faster toward the cathode than LDL from nondrinking control subjects. Rabbit antibodies raised by using keyhole limpet hemocyanin modified in vitro by 4-hydroxynonenal or by acetaldehyde as immunogens reacted more strongly with patients' LDL than with control LDL, indicating the presence of oxidatively modified epitopes and acetaldehyde adducts in alcoholic patients' LDL. LDL of alcoholic patients has decreased vitamin E contents. The electromobility of LDL decreased after abstinence from alcohol and returned to normal in 2 wk, but this was not accompanied by a significant increase in its vitamin E contents. When incubated with mouse peritoneal macrophages, patients' LDL induced apolipoprotein E secretion by threefold over control LDL with a concomitant increase in cellular cholesterol. Our results thus demonstrate that LDL of alcoholic patients has lower vitamin E content, is chemically modified in vivo, and exhibits altered biological function. These changes in heavy alcoholic drinkers may render LDL more atherogenic and thereby may counter the antiatherosclerosis effects of moderate alcohol consumption.

Predictors of outcome in patients with unresectable hepatocellular carcinoma receiving transcatheter arterial chemoembolization.

BACKGROUND: Transcatheter arterial chemoembolization (TACE) has been shown to improve survival in patients with unresectable hepatocellular carcinoma (HCC). AIM: To identify pretreatment factors that predicts increased mortality in HCC patients receiving TACE. METHODS: Retrospective review of all patients who underwent TACE for HCC from January 1999 to November 2004. Patient demographics, aetiology of liver disease, laboratory and imaging data regarding tumour characteristics were obtained. RESULTS: Eighty-eight patients (57 +/- 1 years age) received 1-4 sessions of TACE (1.4 +/- 0.1). Tumour size was 3.3 +/- 0.2 cm (range 1-13 cm, median 3 cm) with mean number of lesions 1.9 +/- 0.1 (range 1-7). Mean model for the end stage liver disease score: 11 +/- 0.4; cancer of the liver Italian program score: 1.3 +/- 0.1. Survival post-TACE (excluding those underwent orthotopic liver transplantation) was 12 +/- 0.3 months. By multivariate analysis, tumour size (HR = 1.37, 95% CI: 1.11-1.68, P = 0.003), hypovascularity (HR = 12.62, 95% CI: 1.79-88.92, P = 0.01) and elevated international normalized ratio (HR = 1.46, 95% CI: 1.10-1.92 P = 0.008) are shown to be significant risk factors for increased mortality. CONCLUSION: TACE therapy leads to a mean survival of 12 months in patients not receiving orthotopic liver transplantation. Tumour size, hypovascularity on imaging, and elevated international normalized ratio are predictors of increased mortality after TACE therapy for HCC.

Protein expression changes in the nucleus accumbens and amygdala of inbred alcohol-preferring rats given either continuous or scheduled access to ethanol.

Chronic ethanol (EtOH) drinking produces neuronal alterations within the limbic system. To investigate changes in protein expression levels associated with EtOH drinking, inbred alcohol-preferring (iP) rats were given one of three EtOH access conditions in their home-cages: continuous ethanol (CE: 24h/day, 7days/week access to EtOH), multiple scheduled access (MSA: four 1-h sessions during the dark cycle/day, 5 days/week) to EtOH, or remained EtOH-naïve. Both MSA and CE groups consumed between 6 and 6.5g of EtOH/kg/day after the 3rd week of access. On the first day of EtOH access for the seventh week, access was terminated at the end of the fourth MSA session for MSA rats and the corresponding time point (2300h) for CE rats. Ten h later, the rats were decapitated, brains extracted, the nucleus accumbens (NAcc) and amygdala (AMYG) microdissected, and protein isolated for 2-dimensional gel electrophoretic analyses. In the NAcc, MSA altered expression levels for 12 of the 14 identified proteins, compared with controls, with six of these proteins altered by CE access, as well. In the AMYG, CE access changed expression levels for 22 of the 27 identified proteins, compared with controls, with 8 of these proteins altered by MSA, as well. The proteins could be grouped into functional categories of chaperones, cytoskeleton, intracellular communication, membrane transport, metabolism, energy production, or neurotransmission. Overall, it appears that EtOH drinking and the conditions under which EtOH is consumed, differentially affect protein expression levels between the NAcc and AMYG. This may reflect differences in neuroanatomical and/or functional characteristics associated with EtOH self-administration and possibly withdrawal, between these two brain structures.

Hormonal control of ornithine decarboxylase in isolated liver cells and the effect of ethanol oxidation.

The regulation of ornithine decarboxylase activity was studied in freshly isolated rat hepatocytes incubated in a chemically defined medium for 5 h. Glucagon, dibutyryl cyclic AMP, insulin and dexamethasone produced dramatic increases in ornithine decarboxylase activity, 6--100-times the basal activity. Actinomycin D inhibited completely the stimulatory action of these substances. With glucagon, dibutyryl cyclic AMP and insulin, the rise in ornithine decarboxylase activity was rapid but transient, peaking at 200 min and then declining rapidly. By contrast, the response to dexamethasone was gradual and sustained in the 5 h incubation. The transient nature of the response to glucagon was unaltered by repeated additions of optimally effective doses of glucagon suggesting the development of 'refractoriness' to the actions of this hormone. Ethanol oxidation inhibited by 50% the stimulation of ornithine decarboxylase by glucagon and dexamethasone and this effect was blocked by 4-methylpyrazole, an inhibitor of alcohol dehydrogenase. Acetate (2.5--20 mM), the metabolic product of hepatic ethanol oxidation, was also effective. The data indicate that glucagon, insulin and glucocorticoids are all effective in stimulating the activity of ornithine decarboxylase in isolated hepatocytes but they differ in their duration and time of peak of action. Additionally, the inhibitory effect of ethanol on the hormonal stimulation of ornithine decarboxylase is dependent on its oxidation and may be mediated by acetate.

Gene expression in the hippocampus of inbred alcohol-preferring and -nonpreferring rats.

The hippocampus is sensitive to the effects of ethanol and appears to have a role in the development of alcohol tolerance. The objective of this study was to test the hypothesis that there are innate differences in gene expression in the hippocampus of inbred alcohol-preferring (iP) and -nonpreferring (iNP) rats that may contribute to differences in sensitivity to ethanol and/or in the development of tolerance. Affymetrix microarrays were used to measure gene expression in the hippocampus of alcohol-naive male iP and iNP rats in two experiments (n=4 and 6 per strain in the two experiments). Combining data from the two experiments, there were 137 probesets representing 129 genes that significantly differed (P < or = 0.01); 62 probesets differed at P < or = 0.001. Among the 36% of the genes that were expressed more in the iP than iNP rat at this level of significance, many were involved in cell growth and adhesion, cellular stress reduction and anti-oxidation, protein trafficking, regulation of gene expression, synaptic function and metabolism. Among the 64% of the genes that had lower expression in the hippocampus of iP than iNP rats were genes involved in metabolic pathways, cellular signaling systems, protein trafficking, cell death and neurotransmission. Overall, the data indicate that there are significant innate differences in gene expression in the hippocampus between iP and iNP rats, some of which might contribute to the differences observed in the development of alcohol tolerance between the selectively bred P and NP lines.

Genetic polymorphism of enzymes of alcohol metabolism and susceptibility to alcoholic liver disease.

Differences in the pharmacokinetics of alcohol absorption and elimination are, in part, genetically determined. There are polymorphic variants of the two main enzymes responsible for ethanol oxidation in liver, alcohol dehydrogenase and aldehyde dehydrogenase. The frequency of occurrence of these variants, which have been shown to display strikingly different catalytic properties, differs among different racial populations. Since the activity of alcohol dehydrogenase in liver is a rate-limiting factor for ethanol metabolism in experimental animals, it is likely that the type and content of the polymorphic isoenzyme subunit encoded at ADH2, beta-subunit, and at ADH3, the gamma-subunit, are contributing factors to the genetic variability in ethanol elimination rate. The recent development of methods for genotyping individuals at these loci using white cell DNA will allow us to test this hypothesis as well as any relationship between ADH genotype and the susceptibility to alcoholism or alcohol-related pathology. A polymorphic variant of human liver mitochondrial aldehyde dehydrogenase, ADLH2, which has little or no acetaldehyde oxidizing activity has been identified. Individuals with the deficient ALDH2 phenotype do not have altered ethanol elimination rates but they do exhibit high blood acetaldehyde levels and dysphoric symptoms such as facial flushing, nausea and tachycardia, after drinking alcohol. Because acetaldehyde is so reactive, it binds to free amino groups of proteins including a 37 kilodalton hepatic protein-acetaldehyde adduct and may elicit an antibody response. We would predict that individuals who have low ALDH2 activity because of liver disease or because they have the inactive ALDH2 variant isoenzyme might form more protein-acetaldehyde adducts and elicit a greater immune response. These adducts may represent good biological markers of alcohol abuse and may also play a role in liver injury due to chronic alcohol consumption.

Adequacy of vitamin B6 supplementation during pregnancy: a prospective study.

This prospective study assesses the effect of 2.5, 4, and 10 mg of pyridoxine supplementation during pregnancy on maternal and fetal plasma levels of pyridoxal 5'-phosphate (PLP) and on the degree of coenzyme saturation (activation factor) of aspartate aminotransferase and alanine aminotransferase (alphaEGOT and alphaEGPT) in maternal erythrocytes. More than 4 mg of pyridoxine supplementation daily was required for most pregnancies to maintain maternal plasma PLP levels within the range observed during the first trimester and in the nonpregnant state. The plasma PLP concentrations in maternal and cord blood were highly correlated and indicated a dependence of fetal vitamin B6 nutrition on maternal circulating PLP. Measurements of alphaEGOT and alphaEGPT were not as reproducible as plasma PLP assays and were less sensitive and quantitative indicators. In the majority of subjects, the changes in alphaEGOT and alphaEGPT with time correlated poorly with the changes in plasma PLP. However, when the data were analyzed without regard for their dependence on time, they demonstrated a negative, linear correlation between alphaEGOT and log plasma PLP and between alphaEGPT and log plasma PLP for the group on 2.5 mg of pyridoxine and for all the subjects combined. Finally, the dietary records showed that most of the subjects consumed less than 2 mg of vitamin B6 daily from their food. The results indicate that the current Recommended Dietary Allowance for vitamin B6 during pregnancy (2.5 mg) is too low and that supplementation of this vitamin in an amount more than 4 mg daily is recommended.

The measurement of plasma vitamin B6 compounds: comparison of a cation-exchange HPLC method with the open-column chromatographic method and the L-tyrosine apodecarboxylase assay.

A cation-exchange high-performance liquid chromatographic (HPLC) method was found to be comparable to the open-column (OCC) method for measuring six different B6 compounds in human plasma and the L-tyrosine apodecarboxylase (LTD) assay for pyridoxal-P (PLP). Plasma samples were obtained from 9 subjects before and after 7 days of pyridoxine (PN) supplementation. PLP, pyridoxal (PL) and 4-pyridoxic acid (4-PA) were the major B6 compounds in plasma and the only compounds which increased after supplementation. The coefficients of correlation between any 2 of the 3 methods in measuring plasma PLP were greater than 0.93, and between HPLC and OCC in quantifying PL and 4-PA were 0.82 and 0.63, respectively. With the low plasma levels of pyridoxamine-P, PN and pyridoxamine, the results from OCC were consistently higher than those from HPLC. However, recoveries of spiked B6 compounds in plasma by these methods were between 84 to 105 percent for all the 5 vitamers and 4-PA.

Benzodiazepine receptor ligands with different intrinsic efficacies alter ethanol intake in alcohol-nonpreferring (NP) rats.

Benzodiazepine (BDZ) receptor ligands with varying intrinsic efficacies [RO19-4603, 0.02-0.15 mg/kg; FG 7142 1-16 mg/kg; DMCM, 1-8 mg/kg; RO16-6028 (bretazenil), 8-32 mg/kg] in modulating GABAergic activity were examined for the ability to alter palatability-induced ethanol (EtOH) intake in the alcohol-nonpreferring (NP) line of rats. NP rats on a 22-hour fluid-deprivation schedule were given 2-hour daily access to a 10% (v/v) EtOH/3% (g/v) polycose solution and water. Average EtOH intake was 2.1 +/- 0.2 g/kg/2 hours, and water intake was 17.1 +/- 0.9 ml/2 hours. During the initial 15 minutes of the 2-hour session, RO19-4603, the imidazothienodiazepine partial inverse agonist reduced EtOH intake to 19% of control values at 0.04 mg/kg and completely suppressed drinking of the EtOH solution at 0.15 mg/kg. Twenty-four-hour postdrug administration, the 0.08-mg/kg dose of RO19-4603 completely suppressed drinking of the EtOH solution at the 60-minute interval, and the 0.15-mg/kg dose reduced intake to 20% of control levels at the 15-minute interval. FG 7142, the partial beta-carboline inverse agonist reduced EtOH drinking at the 60-minute interval with the 1-mg/kg dose, and the 16-mg/kg dose reduced water intake at the 15-minute interval. DMCM, the full beta-carboline inverse agonist, significantly reduced water intake at 15 minutes (4 and 8 mg/kg), and the same doses caused a substantial increase in EtOH drinking at the 120-minute interval. The anxiolytic agent bretazenil (16 and 32 mg/kg) increased EtOH consumption during the initial 15 minutes to 270% to 425% of control levels, and water intake increased by the end of the 2-hour session to as much as 210% of control following administration of the 32-mg/kg dose. These findings support existing evidence suggesting that BDZ receptor ligands may modify neuronal processes that mediate some reinforcing and/or aversive properties of alcohol. They further demonstrate a potential importance of the GABAA-BDZ receptor complex in mediating palatability- (environmentally) induced EtOH drinking even in rats selectively bred for low alcohol preference.

Persistence of cyclosporine after withdrawal of the drug in a patient with chronic liver transplant rejection. Role of the monoethylglycinexylidine test.

Patients with chronic rejection of liver allografts may show persistently high cyclosporine levels. This phenomenon may be due to a down-regulation of the P450 cytochrome system. The monoethylglycinexylidine test was useful in confirming this hypothesis.

Intraoperative placement of a Wallstent for portal vein stenosis and thrombosis after liver transplantation.

We report a case of orthotopic liver transplantation, in which portal vein thrombosis developed in the immediate postoperative period. Surgical thrombectomy and intraoperative placement of a large caliber Wallstent resulted in long-term patency. The unique feature of this case is the intraoperative placement of the stent via the inferior mesenteric vein under fluoroscopic guidance. The use of a large caliber (16 mm) stent obviated the need for postoperative anticoagulation.

Neurontensin studies in alcohol naive, preferring and non-preferring rats.

Neurotensin is a tridecapeptide, present in the central nervous system and the gastrointestinal tract in man and animals. Previous studies in mice selectively bred for differences in hypnotic sensitivity to ethanol have provided data to suggest that neurotensinergic systems may mediate differences in ethanol's actions in these animals. The present study sought to determine if brain neurotensin levels differed between two lines of rats which have been selectively bred for alcohol preferring or non-preferring behaviors. In addition, electroencephalographic and event-related potential responses to intracerebroventricular saline and neurotensin (10 or 30 microg) were evaluated between the rat lines. Similar to human subjects at high genetic risk for alcoholism, preferring rats were found to have more electroencephalographic fast frequency activity and lowered amplitude of the P3 component of the event-related potential in cortical sites under the saline condition. Overall, electrophysiological response to neurotensin, in the two rats lines, was substantially similar to what has been reported previously in outbred Wistar rats, and consisted of dose-related decreases in overall electroencephalographic spectral power concomitant with increases in amplitude and decreases in the latency of the N1 component of the event-related potential. However, differences in neurotensin responses between the preferring and non-preferring rat lines were also found. The differences in electroencephalographic high-frequency activity and in P3 amplitude seen between the rat lines under control conditions were eliminated by administration of neurotensin. In addition, preferring rats appeared to be more sensitive to neurotensin-induced increases in N1 amplitude. Brain neurotensin concentrations were also found to differ between the lines. Significantly lower concentrations of neurotensin were found in the frontal cortex of preferring rats when compared to non-preferring rats or outbred Wistars. Taken together, these studies suggest that differences in the regulation of neurotensin neurons may contribute to the expression of behavioral preference for ethanol consumption in selective rat lines. Additionally, drugs targeting the neurotensinergic system may plausibly be of utility in the treatment of alcoholism.

Rate-determining factors for ethanol metabolism in fasted and castrated male rats.

The effects of castration and fasting upon the alcohol elimination rate, liver alcohol dehydrogenase (LADH) maximum activity (Vmax), and hepatic concentrations of ethanol, acetaldehyde, and free NADH during ethanol oxidation were examined in male Wistar rats. Castration increased the Vmax of LADH and, to a lesser extent, the alcohol elimination rate in vivo. On the other hand, fasting reduced the Vmax of LADH and the alcohol elimination rate in sham-operated and castrated rats but it did not nullify the effect of castration. Castration produced small but significant changes in the hepatic concentrations of ethanol, acetaldehyde and free NADH in fed rats during ethanol oxidation. Fasting also caused significant increases in the concentration of free NADH during alcohol oxidation in both the sham-operated and castrated groups. The ratio of the steady-state velocities of LADH in situ to the maximum velocities of LADH (v/Vmax) under the different experimental conditions was calculated by using the steady-state rate equation for the enzyme mechanism of rat LADH and its kinetic constants. The calculated v/Vmax ratios were 50-62%, indicating that LADH activity was limited to about the same extent by its substrates and products under these conditions and that the changes in alcohol elimination rates produced by fasting and castration mainly reflected changes in the Vmax of LADH. The calculated steady-state velocities in situ (v) were 14-28% lower than the measured rates of alcohol elimination in vivo. The extent of agreement is probably acceptable in view of the assumptions needed to determine the free NADH concentration in liver and the existence of non-LADH-related processes for alcohol elimination in vivo.