Sampling of orbital sinus blood closely reflects brain ethanol content in rats

It was demonstrated that the aerial righting reflex can be used as an index of acute ethanol-induced impairment of motor coordination in rats, and was found to directly correlate with blood ethanol content from the infraorbital plexus. A study of ethanol within the blood and its distribution in brain regions showed that the ethanol content of orbital sinus blood closely reflected that in the cerebral cortex, midbrain, and cerebellum. Ethanol administration by intraperitoneal (IP) injection (2, 3 or 4 g/kg) produced the same distribution as 24 hr ethanol vapor inhalation (28 mg/l). Blood ethanol concentrations were slightly higher than brain ethanol concentrations when measured at 10, 30, and 60 min after IP injection and immediately following ethanol vapor administration. Also, in rats 48 hr following ethanol vapor inhalation when tolerance to ethanol is exhibited, the distribution and concentrations of ethanol in blood and brain from acute ethanol (2 g/kg, IP) were unaltered when compared with controls. These data suggest that ethanol distribution within the brain does not play a role in the phenomenon of tolerance to ethanol.     

Impaired trace fear conditioning following neonatal ethanol: reversal by choline

Neonatal ethanol exposure in animals results in performance deficits on tests of hippocampus-dependent spatial memory, and recent studies have shown that extra dietary choline can ameliorate some of these impairments. In this experiment, rats were administered 5.25 g/kg ig ethanol per day or sham intubations on Postnatal Days (PD) 4-9 and choline (0.1 ml of an 18.8 mg/ml solution) or saline subcutaneously on PD 4-20. On PD 30, rats were given delay or trace fear conditioning trials and were tested for conditioned stimulus-elicited freezing 24 hr later. Neonatal ethanol produced a profound impairment in trace conditioning that was reversed by choline. Groups did not differ in delay conditioned responding, indicating that neonatal ethanol produces a relatively selective cognitive deficit that can be alleviated with supplemental choline.     

The effect of repeated withdrawal episodes on acquisition and loss of tolerance to ethanol in ethanol-treated rats

Male rats were administered ethanol via an intragastric catheter (8.0-12.0 g/kg/day) either continuously for 8 weeks or on a binge schedule with four 2 week cycles of drug administration separated from each successive cycle by a 2 week period of no drug treatment. Older rats were administered ethanol for 2 weeks, to provide an age control for the binge-treated animals as age can alter an animal's sensitivity to ethanol. Acquisition and loss of tolerance to ethanol-induced motor impairment were measured on a dowel task while acquisition and loss of tolerance to ethanol-induced hypothermia were assessed by measuring rectal temperature. Acceleration of tolerance development to both ethanol-induced motor impairment and hypothermia was observed in animals subjected to repeated withdrawal episodes (binge-Study 1) but not in the controls for total dose and duration of drug treatment who experienced withdrawal only once (continuous-Study 2). Persistence of tolerance to ethanol-induced motor impairment occurred in both binge and continuously treated animals while persistence of tolerance to ethanol-induced hypothermia was seen only in the binge treated animals. Age (3 to 7 months) did not affect tolerance development or decay. After three cycles of drug treatment (three withdrawal episodes), binge treated animals showed an impairment in motor ability when blood ethanol levels were near zero. This impairment disappeared when the animals were administered ethanol, indicating a normalizing effect of ethanol on motor behavior in animals subjected to repeated episodes of withdrawal. A similar, but not significant, effect was seen in continuously treated animals. Thus, in an animal exposed to prolonged ethanol treatment, persistent changes in responding to the drug were found. The persistence of these changes was enhanced by the experience of withdrawal from ethanol.     

Maternal care alterations induced by repeated ethanol leads to heightened consumption of the drug and motor impairment during adolescence: a dose-response analysis

Maternal ethanol exposure during lactation induces behavioral alterations in offspring, including disruptions in motor skills and heightened ethanol ingestion during adolescence. These behavioral outcomes appear to partially depend on ethanol-induced disruptions in maternal care. The present study assessed motor skills and ethanol intake in adolescent rats raised by dams that had been repeatedly given ethanol during lactation. Female rats (postpartum days [PDs] 3-13) were administered ethanol (0.5, 1.5, or 2.5 g/kg) or vehicle every other day and allowed to freely interact with their pups. During adolescence, the offspring were evaluated for motor coordination (accelerating rotarod test) and oral ethanol self administration. The lowest maternal ethanol dose (0.5 g/kg) mildly affected motor performance, whereas the higher doses (1.5 and 2.5 g/kg) resulted in motor coordination impairment and greater ethanol intake. Maternal care behavior was affected by ethanol in a dose-dependent fashion. These results indicate that early experience with ethanol during lactation, even when the drug dosage is kept relatively low, leads to long-term consequences in offspring. Dose-response effects on maternal care behavior (i.e., nest building, crouching) may underlie disruptions in motor development and greater ethanol intake resulting from these early ethanol experiences.     

Ethanol and caffeine effects on daytime sleepiness/alertness

Eighteen normal-sleeping young (mean age 25.6 years) volunteers received either ethanol (0.75 g/kg producing blood ethanol concentrations of 71.1 +/- 24.3 mg/100 ml on average) or caffeine (4.0 mg/kg dissolved in 300 ml of 97% caffeine-free instant coffee) at 0920-0950 h after spending 5, 8, or 11 h time in bed (TIB) the previous night. Latency to sleep onset was tested at 1000, 1200, 1400, and 1600 h. Mean sleep latency differed significantly between drugs on each day of testing, with subjects being sleepier after ethanol than caffeine. On day 2 the TIB manipulation produced significant differences in latency, with the 11-h condition differing from both the 8- and 5-h conditions. The significant interaction revealed that in fully rested subjects (11-h TIB), ethanol did not produce sleepiness to the degree it did after 5 or 8 h in bed. In this condition latencies were similar to those of the caffeine and 5- or 8-h TIBs.     

Differential effects of ethanol concentration on blood pH, PCO2 and PO2 in LS and SS mice

Concentration-dependent effects of ethanol upon behavior and upon physiological regulatory mechanisms have been suggested. In a previous study, we found that the concentration of an acute ethanol injection confounded dose-response relationships for measures of blood pH, PCO2, and PO2. Two lines of mice that differ in CNS sensitivity to the hypnotic effects of ethanol (long sleep, LS; short sleep, SS) have also been found to differ in sensitivity to the physiological depressant effects of this drug. Therefore, we designed the present study to examine how intraperitoneal (IP) injections of varying ethanol concentrations differentially affect blood parameters (pH, PCO2, and PO2) and respiration rate in the LS and SS mouse lines. Different groups of LS female mice were injected IP with 165.0, 198.8, 248.1, or 330.0 mg/ml ethanol at a constant dose of 3.3 g/kg. Groups of SS female mice received 205.0, 247.0, 308.3, or 410.0 mg/ml ethanol at a dose of 4.1 g/kg. Blood parameters and respiration rate were measured at 60 min post-injection. In both the LS and SS mice, increasing concentrations of ethanol caused a progressive decline in respiration rate and blood pH. Blood PCO2 values were greater than control only at the highest ethanol concentration. Concentration-dependent effects of ethanol on blood PO2 values were not found in either line. However, LS PO2 was significantly elevated from the control value at all ethanol concentrations. These results suggest that a dose-response relationship may be obtained by varying ethanol concentration for some physiological measures, but not for others. Thus, attention should be paid to differences in concentration as well as amount of ethanol when dose-response curves are to be constructed.     

Effect of chronic acamprosate treatment on voluntary alcohol intake and beta-endorphin plasma levels in rats selectively bred for high alcohol preference

Our previous studies have shown that repeated acamprosate administration to ethanol-naive Warsaw high preferring (WHP) rats resulted in increased plasma beta-endorphin levels and at least partially prevents increases in levels of this peptide after a single administration of ethanol compared with untreated control rats. The objective of the present study, which included 45 WHP rats, was to continue the past research and investigate the effect of 10-day acamprosate treatment (200 mg/kg p.o.) on alcohol intake using a free-choice procedure and on changes in plasma beta-endorphin levels while alcohol is available, and 10 days after alcohol withdrawal. Voluntary alcohol consumption increases plasma levels of beta-endorphin from 440+/-25 pg/ml to 711+/-57 pg/ml (p=0.0002). After a 10-day of alcohol withdrawal, the levels of this peptide were significantly reduced compared with levels in rats with free access to ethanol (711+/-57 pg/ml vs. 294+/-38 pg/ml, p=0.000001) and in control naive rats (440+/-25pg/ml vs. 294+/-38pg/ml, p=0.044). Chronic treatment with acamprosate increased plasma beta-endorphin levels both in WHP rats with free access to ethanol (440+/-25 pg/ml vs. 616+/-49 pg/ml, p=0.008) and in rats after ethanol withdrawal (440+/-25 pg/ml vs. 620+/-56 pg/ml, p=0.007). In the group with free access to ethanol, there was a significant reduction in mean ethanol intake, from 6.75+/-0.20 g/kg body weight/day to 4.68+/-0.25 g/kg/day. Our results indicate that chronic acamprosate treatment may have beneficial effects, as it increases the beta-endorphin concentration thereby compensating for beta-endorphin deficiency during ethanol withdrawal. As the endogenous opioid system has an important role in the development of craving for alcohol, restoring the alcohol-induced deficits in beta-endorphin levels may be an important factor to prevent craving and maintaining abstinence. We suppose that the anti-craving mechanism of acamprosate that has been reported to abolish excessive glutamate release during alcohol withdrawal may be accompanied by compensation for the beta-endorphin deficiency.     

Adrenalectomy and the anorectic effects of benzodiazepine inverse agonists and opiate antagonists in rats fed a palatable diet

The benzodiazepine (BZ) receptor inverse agonists, FG 7142 (1.25-10.0 mg/kg, IP) and CGS 8216 (2.5-20.0 mg/kg, IP), significantly attenuated the consumption of a palatable sweetened diet by non-deprived male rats in a 30 min test. Adrenalectomy failed to affect the reduction in food intake produced by these two drugs. Similarly, the anorectic effects of the opiate antagonists, naltrexone (0.3-3.0 mg/kg, SC) and diprenorphine (0.3-3.0 mg/kg, SC) in the same feeding paradigm were unaffected by adrenalectomy. So far as palatability-induced feeding in concerned, anorectic effects of BZ inverse agonists and opiate-antagonists appear to be adrenal-independent in the rat. The benzodiazepines, clonazepam (0.3 mg/kg, IP) and diazepam (1.0 mg/kg, IP), stimulated food consumption in both adrenalectomized and sham-operated animals.     

Hepatic vagotomy (partial hepatic denervation) does not alter ingestive responses to metabolic challenges

The effects of transecting the hepatic branch of the vagus nerve were studied in male and female Long Evans rats. No differences between same-sex hepatic vagotomized (HV) and sham operated (SHM) animals were found in postoperative body weight, diurnal patterns of ingestion, or the response to insulin (4, 8, 12 U/kg, IP), 2-deoxy-D-glucose (125, 250, 500 mg/kg, IP), sodium chloride (10 ml/kg: 0.5, 1.0, 2.0 M, IP), or polyethylene glycol (10 ml/kg: 30% w/v, SC). These results can be interpreted as evidence for a minor role of the hepatic vagus in the total complement of hepatic afferent innervation.     

Ethanol prevents stress-induced increase in cortical DOPAC: reversal by RO 15-4513

Electric foot-shock increased DOPAC and decreased DA levels by about 70 and 20% respectively in the medial prefrontal cortex in rats. Pretreatment with diazepam (5 mg/kg IP) or ethanol (1.2 g/kg orally) prevented these stress-induced changes. The protective effect of diazepam and ethanol was eliminated by RO 15-4513 (5 mg/kg IP) a partial inverse benzodiazepine agonist.     

Impairment of the intestinal barrier by ethanol involves enteric microflora and mast cell activation in rodents

Alcohol hepatic toxicity in heavy drinkers is associated with high endotoxin blood levels and increased intestinal permeability. Because endotoxins can cross damaged mucosa, we investigated the mechanisms through which ethanol impairs the colonic epithelium of rats submitted to acute alcohol intake. Colonic permeability to (51)Cr-ethylenediamintetraacetic acid was increased 24 hours after 3.0 g/kg ethanol intake (3.2 +/- 0.2% versus 2.2 +/- 0.2%) and was associated with significant endotoxemia. Antibiotics and doxantrazole (a mast cell membrane stabilizer) significantly inhibited the effect of ethanol. Two hours after intake, plasma concentrations of ethanol were twofold higher in antibiotic-treated rats than in controls (155.8 +/- 9.3 mg/dl versus 75.7 +/- 7.6 mg/dl, P < 0.001). Lumenal concentrations of acetaldehyde were markedly increased after ethanol intake (132.6 +/- 31.6 micromol/L versus 20.8 +/- 1.4 micromol/L, P < 0.05) and antibiotics diminished this increase (86.2 +/- 10.9 micromol/L). In colonic samples mounted in Ussing chambers, acetaldehyde but not ethanol increased dextran flux across the mucosa by 54%. Doxantrazole inhibited the effect of acetaldehyde. This study demonstrates that an acute and moderate ethanol intake alters the epithelial barrier through ethanol oxidation into acetaldehyde by the colonic microflora and downstream mast cell activation. Such alterations that remain for longer periods could result in excessive endotoxin passage, which could explain the subsequent endotoxemia frequently observed in patients with alcoholic liver disease.     

Characterization of a rat lymphocyte culture system for assessing sister chromatid exchange after in vivo exposure to genotoxic agents

Lymphocyte culture systems have the major advantage of permitting the analyssis of in vivo cytogenetic damage with minimal injury to the animal under study. This paper describes a rat lymphocyte culture system designed for the study of sister chromatid exchange (SCE) induced by in vivo exposure to genotoxic agents. A standard protocol was established in which 1 to 2 ml of blood are removed from rats by cardiac puncture, washed three times with phosphate-buffered saline (pH 7.4), and grown in RPMI 1640. 5-Bromodeoxyuridine (BrdU) (1.0 μM) is added after 24 hr, and cells are harvested after 56 hr of culture. Critical steps for successful blast transformation include washing the blood in buffered saline and the adding of 2.0 to 4.0 μg phytohemagglutinin/ml to the culture medium. The use of low concentrations of BrdU (≤5.0 μM) is recommended to maintain low baseline SCE levels and to avoid cytotoxicity. The mutagenic carcinogen, ethyl methanesulfonate (EMS), was used as a positive control agent and at a dose of 300 mg/kg caused a fourfold increase in SCE frequency. Twenty-eight days after EMS administration (30, 100, 300 mg/kg), lymphocytes from treated animals still displayed SCE levels at least 50% above baseline. This system provides a reliable means of investigating chemically induced SCE in the rat.     

Pharmacokinetics of 125I-radiolabelled PEG-hemoglobin SB1

PEG-hemoglobin SB1 (SB1) is a polyethylene glycol (PEG)-modified hemoglobin-based oxygen carrier, intended for use as resuscitation fluid for brain stroke and as a blood substitute. An intravenous pharmacokinetics (PK) studies with SB1 was investigated in male albino Sprague-Dawley (SD) rats and male beagle dogs at doses of 5 and 12.5 ml/kg for rats and 10 ml/kg for dogs. Total hemoglobin in plasma and whole blood was determined by gamma scintillation counter-detecting 125I-radiolabelled SB1. In the 5 ml/kg rats (n = 9), the Cmax, t1/2, AUCt and Tmax were 9.055 mg equivalents/ml, 9.6 hr, 79.6 mg equivalents.hr/ml and 0.20 hr in the plasma and 4.954 mg equivalents/ml, 9.7 hr, 37.6 mg equivalents.hr/ml and 0.11 hr in the whole blood, respectively. Those parameters in the 12.5 ml/Kg of rats (n = 9) were 19.00 mg equivalents/ml, 10.6 hr, 223.5 mg equivalents.hr/ml and 0.33 hr in the plasma and 10.58 mg equivalents/ml, 16.1 hr, 99.0 mg equivalents.hr/ml and 0.33 hr in the whole blood, respectively. An increase in the dose level from 5 to 12.5 ml/kg resulted in the increase in both Cmax and AUC24, and the increases in these parameters appeared to be in proportion to the dose increment. Thus, following the 2.5-fold increase in administered dose, Cmax was increased by a factor of 2.1 in both plasma and whole blood, while AUC24 was increased by a factor of 2.8 for plasma and 2.6 for whole blood. In the dogs receiving 10 ml/kg (n = 3), the Cmax, t1/2, AUC168 and Tmax were 12.70 mg equivalents/ml, 47.2 hr, 425.7 mg equivalents.hr/ml and 0.083 hr in the plasma and 8.372 mg equivalents/ml, 50.3 hr, 241.3 mg equivalents.hr/ml and 1.003 hr in the whole blood, respectively. The present work provides an insight into the pharmacological behavior of a PEG-modified hemoglobin.     

Kindling modifies morphine, cocaine and ethanol place preference

Brailowsky and Garcia (1999) proposed the existence of a relationship between epilepsy and addiction. To prove this hypothesis, pentylenetetrazol kindled rats were tested in the conditioned place preference (CPP) paradigm for their reaction to various addictive drugs with different modes of action (morphine, cocaine and ethanol). In separate experiments, locomotor activity and body temperature after application of the same drugs were tested in kindled and non-kindled rats. In the CPP experiment there were significant differences between both groups of rats. Non-kindled animals showed place preference to morphine (5.0 mg/kg) or cocaine (20.0 mg/kg). This reaction was abolished in the kindled rats. Moreover, control rats demonstrated aversion to 2.0 g/kg ethanol. However, ethanol aversion was not detectable in kindled rats. Moreover, there was no difference between non-kindled and kindled rats in locomotor activity and body temperature after morphine (1.0 and 5.0 mg/kg), cocaine (10.0 and 20.0 mg/kg), or ethanol (0.5 and 2.0 g/kg) application. This suggests alterations in reward systems as a consequence of kindling. It is hypothesised that GABAergic neurones in the ventral tegmental area might play a major role in the alterations found.     

Effects of LH kainic acid infusions on ingestion and autonomic activity

The effects of lateral hypothalamic (LH) infusions of kainic acid (KA) were determined on ingestive behavior, body weight, and motor and autonomic activity. In Experiment 1 male hooded rats received bilateral LH infusions of isotonic saline in volumes of 0.5 or 1.0 μl, 3.0 μg KA in volumes of 0.5 or 1.0 μl, or 6.0 μg of KA in a volume of 1.0 μl. All animals receiving 6.0 μg/1.0 μl of KA died. The 3.0 μg/0.5 μl dose resulted in transient decreases in food and water consumption and body weight. Animals receiving this dose no longer drank in response to 2 cc/kg 15% NaCl injections, exhibited attenuated drinking in response to 24 hr water and food deprivation, exhibited a transient decrease in eating following food deprivation and decreased eating following 750 mg/kg injections of 2-deoxy-D-glucose. Minimal effects on these measures were observed following the 3.0 μg/1.0 μl dose. In Experiment 2 rats received unilateral infusions of KA and the effects on motor and autonomic activity and ingestive behavior and body weight were compared to unilateral saline infused animals and animals with radiofrequency lesions. Only transient decreases in food consumption lasting 1–2 days were observed for both unilateral KA LH infused and lesioned animals. In KA infused rats contralateral exophthalamus, rapid shallow breathing, bilateral mydriasis, no contralateral pupillary constriction response, excessive salivation, body tremors, seizures, convulsions, teeth chattering, contralateral tail suspension induced spinning and turning, and elevated body temperature were observed for up to 6 hr following the infusion. Results are discussed in terms of lateral ventral diencephalic neurons involved in the more permanent deficits associated with bilateral LH damage.     

Physiologic responses of cross-linked hemoglobin in endotoxin-treated rats

Purified human cross-linked hemoglobin (alpha alpha Hb) as well as recombinant human hemoglobin is undergoing clinical trials in the setting of acute blood loss and perioperative hemodilution. We have previously demonstrated that in rabbits with circulating plasma Hb, such as alpha alpha Hb, infusion of endotoxin (LPS) impairs myocardial contractility which results in hypotension, tissue hypoperfusion and increased mortality. The untoward cardiovascular effects occurring after the combined infusion of LPS and alpha alpha Hb in this model are similar to those reported for other agents that inhibit nitric oxide (NO) availability. To determine if the deleterious effects of alpha alpha Hb and LPS were species specific, we performed similar studies in rats. Anesthetized Sprague-Dawley rats received LPS (4 mg/kg or 40 mg/kg) alone or in combination with alpha alpha Hb (0.7 g/kg). Mean arterial blood pressures (MAP) increased in the group that received alpha alpha Hb alone (105 +/- 8 to 120 +/- 7 mm Hg, p = 0.2) and a decrease was noted in the groups that received low dose LPS (4 mg/kg, p = 0.5) and high dose LPS (40 mg/kg, p = 0.016). MAP in rats treated with the LPS at either dose combined with alpha alpha Hb remained unchanged. Levels of urine nitrite, which was measured as a surrogate marker for plasma NO, were significantly decreased at 2 hr in groups that received the combination of alpha alpha Hb and LPS at 4 mg/kg (p = 0.022) and 40 mg/kg (p = 0.003). No significant decrease was observed in animals treated only with alpha alpha Hb (p = 0.21) or LPS (4 mg/kg; p = 0.78 and 40 mg/kg; p = 0.65). Survival was evaluated during 72 hr in animals that were infused with high dose LPS (40 mg/kg) alone or in combination with alpha alpha Hb and then allowed to recover. The survival of rats treated with LPS alone or the combination was 29% at the end of 24 hr and was 100% for rats receiving only alpha alpha Hb. The data suggest that the toxicity of alpha alpha Hb appears to be a species specific phenomenon.     

Preservation of renal structure and function in the rat remnant kidney model of chronic renal failure by enalapril treatment

The remnant kidney model of chronic renal failure was established in rats subject to subtotal (1 7/8) nephrectomy and the evolution of renal injury studied over a period of 6 wk. One wk after subtotal nephrectomy, rats had a mean conscious systolic blood pressure of 158 +/- 5 mm Hg and serum creatinine of 128 +/- 9 mumol/l. Both systolic blood pressure and serum creatinine rose over the next 5 wk in concert with progressive glomerulosclerosis and proteinuria. Enalapril, an angiotensin converting enzyme inhibitor, was administered (5 mg/kg/day) to rats (n = 11) from 1 wk after subtotal nephrectomy. Enalapril lowered systolic blood pressure over the treatment period. Systolic blood pressure was 122 +/- 5 mm Hg compared with 176 +/- 7 mm Hg in untreated rats (p less than 0.001) at 6 wk. Serum creatinine 6 wk after subtotal nephrectomy was 110 +/- 9 mumol/l with enalapril treatment, compared with 159 +/- 21 mumol/l (p less than 0.025) in control animals. Enalapril treated rats had lower urinary protein excretion than controls (15 +/- 3 mg/24 hr vs 85 +/- 22 mg/24 hr, p less than 0.0001) at 6 weeks. Glomerulosclerosis, assessed by blinded histological score, was also reduced in the enalapril treated group (1.79 +/- 0.08 vs 2.36 +/- 0.16, p less than 0.01). Enalapril treatment was associated with a reduction in filtration fraction (51Cr-EDTA/125I-hippurate clearance). At 6 wk, filtration fraction was 0.30 +/- 0.03 in enalapril treated and 0.48 +/- 0.03 in control rats (p less than 0.001). Enalapril treatment in the subtotal nephrectomy model of renal failure preserved renal structure and function.(ABSTRACT TRUNCATED AT 250 WORDS)     

Heme overload inhibits the growth of normal rats

We have developed a model with which to evaluate the direct metabolic effect of circulating heme in chronic hemolytic diseases. Normal rats were injected with heme intraperitoneally (IP) using different treatment schedules and then killed so their growth parameters could be measured; heme injections were well tolerated, but final body weight was decreased in the heme treated groups. In one experiment, we used heme 40 mg/kg IP 2x/d for 8 d and found that overall growth was significantly inhibited (weight increment in the heme-treated group +45 +/- 3g vs +65 +/- 3g in the controls, P less than .05); although serum somatomedin activity was significantly diminished, the liver extract somatomedin activity was not decreased. In another experiment, we compared the effect of heme 40 mg/kg versus heme 80 mg/kg IP 2x/d for 5 d: growth inhibition was dose-dependent (weight change +31 +/- 2g for vehicle-treated group versus +12 +/- 3 grams for heme 40 mg/kg group versus -6 +/- 5 grams for heme 80 mg/kg group) and related to serum heme levels (3.53 +/- 0.47 mcM for vehicle group versus 5.42 +/- 0.71 mcM for heme 40 mg/kg versus 10.91 +/- 0.84 mcM for heme 80 mg/kg group). Our data demonstrate that IP heme administration is feasible and provides a valid approach with which to identify the potential direct metabolic consequences of abnormally circulating heme and its contribution to the growth retardation associated with chronic hemolytic diseases.     

Similar cytokine but different coagulation responses to lipopolysaccharide injection in D-galactosamine-sensitized versus nonsensitized rats.

To compare cytokine release and coagulation disturbances induced by administration of high versus low doses of endotoxin (lipopolysaccharide [LPS]), we used two endotoxin test systems similar in mortality but different in the degree of endotoxemia. One group of rats (n = 11) randomly received endotoxin (15.0 mg/kg of body weight intraperitoneally [i.p.]) and 1 ml of Ringer's solution (nonsensitized animals). The second group (n = 11) received 1 ml of D-galactosamine (500 mg/kg i.p.) and endotoxin (100 micrograms/kg i.p.) simultaneously (sensitized animals). Endotoxin levels in the plasma of nonsensitized rats were 1,000-fold higher than those in the plasma of sensitized rats (69.33 x 10(3) +/- 22.42 x 10(3) versus 75.8 +/- 27.08 ng of LPS per ml), leading to a mortality of 91% in nonsensitized rats versus 82% in the sensitized-rat model within 48 h postendotoxemia. Serum transaminase activity increased up to 100-fold in sensitized rats as a sign of hepatocyte damage. Despite the large difference in LPS levels in plasma, the time courses of the plasma tumor necrosis factor (TNF) increase were similar in the two groups, with a peak at 2 h (54 +/- 12 ng/ml in nonsensitized rats versus 43 +/- 12 ng/ml in sensitized rats), and also similar to that of a group of nonsensitized rats (n = 5) that received a low dose of LPS (100 micrograms/kg) only (52 +/- 21 ng/ml), while D-galactosamine alone did not induce TNF release. Despite similar TNF levels, a more pronounced coagulation disorder was observed at 4 h in nonsensitized rats (with the high LPS dose) as measured by platelet counts, plasma fibrinogen levels, and activated partial thromboplastin time prolongation (191 x 10(3) +/- 107 x 10(3) cells per microliter, 40 +/- 24 mg/dl, and 53 +/- 15 s, respectively) than in rats with the low LPS dose either sensitized (495 x 10(3) +/- 153 x 10(3), 95 +/- 49, and 38 +/- 16, respectively) or nonsensitized (439 x 10(3) +/- 62 x 10(3), 170 +/- 18, and 35 +/- 11, respectively).(ABSTRACT TRUNCATED AT 400 WORDS)     

The phytoestrogen alpha-zearalenol reverses endothelial dysfunction induced by oophorectomy in rats

It has been shown recently that alpha-zearalenol, a resorcyclic acid lactone, prevents bone loss in a rat model of postmenopausal bone loss. We have therefore investigated the effects of this phytoestrogen on endothelial dysfunction induced by estrogen deficiency in rats. Female mature Sprague-Dawley rats underwent a bilateral oophorectomy (OVX rats). Sham-operated animals (sham OVX rats) were used as controls. Three weeks after surgery, animals were randomized to the following treatments: alpha-zearalenol (1 mg/kg/day, i.m., for 4 weeks), 17beta-estradiol (20 microg/kg/day, i.m., for 4 weeks), or their vehicle (100 microl, i.m., of cottonseed oil). Two other groups of rats were treated with alpha-zearalenol or 17beta-estradiol plus the pure estrogen receptor antagonist ICI 182780 (2.5 mg/kg/day, i.m., for 4 weeks). Mean arterial blood pressure (MAP), heart rate (HR), total plasma cholesterol, plasma estradiol, and plasma alpha-zearalenol were studied. We also investigated endothelial-dependent (acetylcholine, 10 nM to 10 microM) and endothelial-independent (sodium nitroprusside, 15 nM to 30 nM) relaxation of aortic rings, as well as N(G)-methyl-L-arginine (L-NMA: 10 to 100 microM)-induced vasoconstriction and calcium-dependent nitric oxide synthase (cNOS) activity in homogenates of lungs taken from both sham OVX rats and OVX rats. Untreated OVX rats had, compared with sham OVX animals, unchanged body weight, MAP, HR, and plasma cholesterol. In contrast oophorectomy reduced plasma estradiol levels (OVX, 2 +/- 0.5 pg/ml; sham OVX, 35 +/- 6 pg/ml), impaired endothelial-dependent relaxation and blunted L-NMA-induced contraction (L-NMA 100 microM: sham OVX, 2.7 +/- 0.3 g/mg tissue; OVX, 1.3 +/- 0.1 g/mg tissue). Moreover OVX rats showed a reduced calcium-dependent NO synthase (cNOS) activity. Treatment with alpha-zearalenol or with 17beta-estradiol reverted the endothelial dysfunction and increased cNOS activity in lung homogenates. These effects were abolished by the pure estrogen receptor antagonist ICI 182780. Our data suggest that alpha-zearalenol improves endothelial-dependent relaxation in OVX rats through an estrogen receptor-mediated effect.