On gestation day 7, osmotic pumps containing morphine base, 66.5 mg/ml, or 0.9% sodium chloride vehicle were implanted into anesthetized pregnant female rats to determine the effects of chronic perinatal exposure to morphine on the neurochemical and behavioral development of 10-day-old rat offspring. There was a significant decrease in the density of D1 receptors in the nucleus accumbens (NAc) of offspring exposed to morphine. In contrast, D2 receptor density was significantly increased in the NAc at this age compared with controls. Perinatal exposure to morphine significantly decreased the density of D2 receptors in the striatum, but had no significant effect on the K-d of D1 or D2 receptors in this brain area. Exposure to morphine also had no significant effect on the locomotor response of 10-day-old offspring to SKF 38393 compared with pair-fed controls, nor was there a significant difference in the response of experimental offspring to quinpirole compared with controls. The results indicate that chronic perinatal exposure to morphine alters the density, but not the sensitivity, of DA receptors in the 10-day-old rat brain.
To study the effects of caffeine on carcinogen biotransformation, the effects of caffeine on N-nitrosodimethylamine demethylase were studied in liver microsomes from control, phenobarbital (PB)-, and 3-methylcholanthrene (3-MC)-pretreated rats. Caffeine significantly decreased N-nitrosodimethylamine demethylase in control microsomes whereas it significantly enhanced the enzyme activity in 3-MG induced microsomes. No apparent effect was observed in PB-induced microsomes. Additionally, caffeine did not show any significant effects on aminopyrine N-demethylase and aniline hydroxylase in control microsomes but significantly inhibited those reactions in 3-MC-induced microsomes. The effects of caffeine on the biotransformation of xenobiotics including carcinogens are different depending on induction states. Bioactivation of carcinogens appears to be enhanced in the presence of caffeine under certain circumstances.
The threshold of the Abbott TDx Cocaine Metabolite Assay (TDx) was lowered from 300 ng/mL benzoylecgonine (BE) to 50 ng/mL by dilution of calibrators and controls with drug free urine. The between-run precision of the modified assay was evaluated by analyzing the urine control samples containing 50, 100, and 500 ng/mL BE for over a 12-week period which yielded CVs of < 8.2%. Five hundred consecutive urine samples from pregnant women and neonates tested for BE by EMIT-d.a.u. with the cut-off value of 300 ng/mL were also tested by the modified TDx assay with the new cutoff value of 50 ng/mL. All positive immunoassay results were confirmed by Gas Chromotography/Mass Spectrometry (GC/MS). The modified TDx assay resulted in 27 more positive samples than the EMIT-d.a.u. Screening urine samples obtained from the Prenatal Clinic and OB-ER by the modified TDx assay yielded a 70% (51/30) increase in positive BE results compared with EMIT-d.a.u. Lowering the threshold of TDx assay to 50 ng/mL enhances the detection of BE in this population at high risk for the tragic consequences of cocaine abuse.
The neuronal membrane fluidity in six different regions of the rat brain was measured by electron spin resonance spectrometry using the spin labels 5- and 16-doxylstearic acid (5- and 16-DS) after administration of ethanol solution (7%) for 3 weeks. The order parameter, calculated from 5-DS spectra, which is utilized to assess the fluidity of the lipid bilayer near the surface of the membrane, increased in the cerebral cortex and hippocampus, and the motion parameter, calculated from 16-DS spectra, for the core of the lipid bilayer, increased in the cerebral cortex. On the other hand, almost no membrane fluidity change was observed in the striatum, midbrain, pons-medulla oblongata and cerebellum. These results indicate that chronic ethanol administration does not decrease membrane fluidity evenly in all the regions of the brain.
Focal glomerulosclerosis is the predominant glomerular lesion in patients with drug abuse. Since mesangial expansion is a precursor of glomerulosclerosis, we studied whether illicit drugs can alter mesangial cell handling of deposited macromolecules. We studied the effects of morphine, cocaine, and alcohol on the aging of mesangial cells as judged by the distribution pattern of mitochondria. After 48 hours of incubation, mesangial cells treated with morphine, alcohol or cocaine showed a higher percentage of aged cells compared with control cells. When growth-arrested mesangial cells were treated with morphine and alcohol, they also had an increased percentage of aged cells compared with control cells. These results suggest that morphine and cocaine accelerate the aging process in mesangial cells. This effect of morphine and cocaine may alter the handling of phlogogenic macromolecules by mesangial cells and may, thus, contribute to the expansion of the mesangium.
The combination of decreased antioxidant vitamins, oxidation of sulfhydryl groups and increased oxygen radical production suggest an oxidative mechanism of cigarette smoke induced damage. Cigarette smoke exposed rats show an increased myocardial sensitivity towards ischaemia/reperfusion. The aim of this study was to determine whether smoke exposure of rats resulted in altered antioxidant concentrations (vitamin E, vitamin C, GSH, cysteine) in the heart, lung, liver, erythrocyte and plasma. The effect of myocardial ischaemia/reperfusion on smoke exposed rats was also investigated. Protein modification (measured by sulfhydryls and carbonyls) of the plasma and isolated hearts was determined. Results showed that there was redistribution of antioxidants between the compartments investigated and oxidative protein modification was observed in the plasma after smoke exposure. Protein modification occurred in the heart but was not due to oxidative injury. Myocardial mitochondrial function was similar to that of rats not subjected to smoke.
Different dose levels of ethanol (2,3,4,5 g/kg) were administered to rat pups between postnatal days 4 and 10. Ethanol caused a dose - dependent decrease in brain weight (measured on postnatal day 12) and inhibition of carbachol - stimulated phosphoinositide metabolism (measured in cerebral cortex slices on postnatal day 7). The 2 g/kg dose, which gave blood alcohol levels of 128 mg/dl, was a no-effect-level for both endpoints. Ethanol administration did not alter the relative distribution of phosphoinositides in the cerebral cortex from 7 day - old rats. These results show a dose - dependent correlation between ethanol - induced microencephaly and inhibition of muscarinic receptor - stimulated phosphoinositide metabolism and add support the hypothesis that this second messenger system may be involved in the developmental neurotoxicity of ethanol.
Pregnenolone-16 alpha-carbonitrile and triamcinolone increase the body's resistance to numerous xenobiotics. Since ethanol, taken in considerable amounts, is metabolized by both alcohol dehydrogenase and the microsomal ethanol oxidizing system, we examined the effect of the mentioned two steroids on the body's response to ethanol and upon the concentrations of this alcohol in plasma. It was found that treatment with the two steroids or with their combination did not substantially alter the body's resistance (dyskinesia, mortality) to ethanol or its concentrations in the plasma. The results are discussed in reference to the role of CYP3A induced by pregnenolone-16 alpha-carbonitrile and CYP2E1 involved in ethanol metabolism.
To evaluate the effects of ascorbic acid on ethanol-induced hypothermia, male rats of the Sprague-Dawley strain (200-250 g) were treated with a hypothermic (2.0 g/kg ip) dose of ethanol, with and without pretreatments with graded doses of ascorbic acid (pH 2.3, 4.0 or 6.8, 500-1000 mg/kg ip) or sodium ascorbate (apparent pH 6.8, 562-1125 mg/kg ip) solutions, and the body temperatures were monitored for 3 hr. Ethanol caused a significant drop in body temperature, which was maximal at 60 min postdosing, and which did not return to preethanol values for the remaining of the observation period. A solution of ascorbic acid at a pK 2.3 was hypothermic in itself, and when given as a pretreatment it accentuated the hypothermic response to ethanol. In doses > 500 mg/kg, both ascorbic acid (pH 4.0 or 6.8) and sodium ascorbate attenuated the thermolytic effect of ethanol and accelerated the return of the body temperatures to pretreatment values in a dose-related manner. Sodium ascorbate caused a significant (p < 0.05) reduction in the blood ethanol levels and an enhancement of the blood and liver acetaldehyde levels that followed a hypothermic dose of ethanol. The same pretreatment did not influence the blood ethanol levels of rats on a nonhypothermic (1 g/kg ip) dose of ethanol to a significant extent. Furthermore, ascorbate offered a marked protection against the hypothermia due to the cyanamide-ethanol reaction. These results suggest that the protective effects of ascorbate are related to its ability to stimulate blood ethanol clearance through increased metabolism.
Antibodies for cocaine and benzoylecgonine were prepared by established methods using diazotized 4-aminococaine or 4-aminobenzoylecgonine conjugated to bovine serum albumin as immunogens. Enzyme immunoassay was first developed using diazotized 4-aminococaine or 4-aminobenzoylecogonine conjugated to horseradish peroxidase as the enzyme labels. The IC(50)s of cocaine and benzoylecgonine were 4 ng/ml and 2 ng/ml for their respective antibodies. However, the IC(50)s Of cocaine and benzoylecgonine decreased to 0.4 ng/ml and 0.1 ng/ml respectively when two heterologous haptens, 4-formylcocaine and 4-formyl benzoylecgonine, were synthesized and used for enzyme labeling. The sensitivity of the assays was considerably improved using the heterologous bridge strategy.
Experiments were conducted to evaluate the effects of cocaine (0.5 mg/kg) on the development of autochthonous tumors induced by the Moloney sarcoma virus (MSV) in adult BALB/c female mice. Enhancement of MSV-induced tumor incidence and tumor growth together with a delay in the usual prompt regression of the tumors were observed when mice were daily injected with cocaine for 3 days after MSV-inoculation. However, no significant effects of cocaine on tumor development were found when it was administered during the 3 days before tumor was inoculated.
The experiments reported here continue the development of the neurons of the Muscle Receptor Organ (MRO) of the crayfish as a pharmacological model for studying the effects of ethanol on the electrical activity of isolated neurons. Previous work showed that ethanol caused a reduced firing rate in statically stretched MRO neurons (Macmillan et al., 1991) whereas it increased the firing during stretch episodes (Walsh et al., 1994). Its effect on the activity of fully adapted MRO neurons between stretch episodes is unknown. We report here results showing that when ethanol (120mM) was applied to MRO neurons subjected tb a staircase extension regime, the firing rate increased in the interpull intervals, even though the MRO neuron was allowed to adapt fully. The implications of this for interpretations of the effect of ethanol on the neuronal membrane and for further identification of the critical elements are discussed.
This report describes the acute effects of ethanol on the electrical output of a stretch-sensitive neuron during periods of intermittent, non-synaptic, stretch stimulation. The firing rate of the neuron of the tonic muscle receptor organ (MRO) of the crayfish, Cherax destructor, was monitored during intermittent stretch before and after the application of 100mM or 140mM ethanol, concentrations that cause reversible behavioural intoxication in the crayfish.Application of 100mM ethanol resulted in a trend towards an increased rate of firing during subsequent pull episodes, a trend which became statistically significant after 25 minutes. At an ethanol concentration of 140mM the effect was more marked and occurred sooner, suggesting that it is concentration dependent. The same concentrations of ethanol applied to the statically stretched neuron have been shown previously to cause a decrease in firing rate. The present result provides evidence that the effect of ethanol on the tonic neuron of the MRO depends on its level of activity.
The activation energies for the activity of ethylmorphine N-demethylase under the condition of higher temperature than break point were not different between male and female rats. In contrast, the value under the condition of lower temperature than break point in female rats was higher than that in male rats. Tn Arrhenius plot analysis, the phase transition point for ethylmorphine N-demethylase and aniline p-hydroxylase was not different in male and female rats. From these results it seems likely that the difference in the distribution of P-450 involved in the ethylmorphine N-demethylation might play, in part, an important role in the appearance of the sex-dependent difference in the activity of ethylmorphine N-demethylation other than the differential expression of the sex-specific P-450 enzyme(s) such as the P-450 3A subfamily acting as ethylmorphine N-demethylase in rat liver microsomes.
The effect of gender on the possible hepatotoxicity from morphine (100mg/kg), cocaine (30 mg/kg) or a combination of morphine (100mg/kg) and cocaine (30mg/kg) administered intraperitoneally were investigated in 24 male and 24 female guinea pigs. The time course of liver toxicity after single dose administration of test substances was also assessed by measuring plasma alanine aminotransferase (ALT) and isocitrate dehydrogenase (ICDH) levels over a period of 96 hours. ICDH values were measured in addition to the more usual ALT values to determine the suitability of using the former for assessing drug induced hepatotoxicity. Cocaine and morphine combined was shown to be hepatotoxic after single dose administration. Male guinea pigs were more susceptible than females as measured by higher plasma ALT and ICDH levels 4 to 24 hours after drug administration. In both sexes this combination provoked elevated liver enzyme activity different from control (saline) evident till 48 hours (ICDH) and 72 hours (ALT) after drug administration. No animals showed evidence of hepatotoxicity after 96 hours. Morphine alone resulted in higher levels of ALT (than control) in males, at 48 and 72 hours and in females, at 24 hours after administration. There was a significant correlation (r=0.74) between plasma ICDH and ALT levels. This study demonstrates that the combined administration of cocaine and morphine is hepatotoxic in the guinea pig and that the male is more susceptible. A theoretical explanation for this hepatotoxicity may be due to an additive effect of these drugs in depleting liver glutathione levels. Plasma ICDH can be considered a suitable indicator of drug induced hepatotoxicity but was no more sensitive than ALT.
Cholinergic regulation of ethanol consumption was studied in rats using subcutaneous doses of saline, atropine (5 mg/kg), or methylatropine (5 mg/kg) twice daily for two days. Utilizing a two-bottle free-choice paradigm, rats were offered a choice between a 0.2% saccharin solution and a 10% ethanol/0.2% saccharin solution. Methylatropine significantly decreased both ethanol and total fluid consumption on the first day of treatment and depressed only ethanol consumption for the next four days. Atropine significantly decreased only ethanol consumption on both treatment days. These results suggest a peripheral site of action for anticholinergic effects on ethanol consumption
Zinc is a co-factor for alcohol dehydrogenase, the ethanol metabolizing enzyme. Ethanol-induced abnormality in zinc metabolism could decrease ethanol metabolism and thus increase circulating and tissue ethanol levels. The influence of zinc on ethanol induced fetal anomalies was investigated by the simultaneous administration of ethanol and zinc to pregnant Sprague-Dawley rats from gestational day 6 through 12. Fetuses were recovered on day 20 of gestation. The offspring of animals treated with ethanol and zinc exhibited a significantly higher litter weight and crown-rump length compared with those of offspring from dams treated with ethanol alone. The mean placental weight was, however, higher in fetuses of ethanol-treated animals. The incidence of resorptions and the occurrence of external and internal anomalies were significantly lower in litters of dams treated with ethanol and zinc compared with those of litters which received a chronic ethanol diet. Circulating levels of ethanol and copper were higher and serum zinc concentration was lower in dams treated with ethanol only.
Nutritional deficiencies involving methionine and zinc have been suggested as a possible mechanism of alcohol embryopathy. These nutrients failed to protect chick embryos from the deleterious effects of ethanol.
Selective ion monitoring techniques with gas chromatography-mass spectrometry (GC-MS) were used for the identification and quantitation of thermal decomposition products of cocaine. Cocaine was heated in glass ampules at different temperatures (180-200-degrees-C). After appropriate time periods the ampules were analyzed for cocaine and its pyrolysates using mass spectrometry. Methylecgonidine (MECD) was the only thermal decomposition product found in this study. Obviously, vapors of cocaine under free basing conditions, must be rich in MECD. Methylecgonidine is an exclusive pyrolysate of cocaine; it is also a major metabolite of cocaine in animals and man and could significantly contribute to the overall toxic effects of cocaine.
Sixteen male pigs were randomly assigned to 4 equal groups to investigate the effects of volatile anesthetics (VA) on cocaine induced systemic toxicity. All pigs were anesthetized with sodium pentothal 30 mg/kg iv and 60 mg/kg im and after tracheal intubation for mechanical ventilation anesthesia was maintained with nitrous oxide [4 L/min (4L)] in oxygen [2 L/min (2L)]. Carotid and pulmonary artery catheters were placed for hemodynamic monitoring. No additional anesthetic was administered in Group O (control group). Minimum Alveolar Concentration (MAC)-equivalent concentrations of isoflurane (1.1%), halothane (0.7%), and enflurane (1.7%), were added to the gas mixtures inhaled by pigs in groups I, H and E respectively. Cocaine hydrochloride (0.8 mg/kg/min) was administered iv to all pigs until cardiac arrest. After the start of cocaine infusion there was a significant decrease in cardiac output, mean arterial pressure and heart rate in all groups of pigs as well as a significant increase in systemic and pulmonary vascular resistance, central venous and pulmonary capillary wedge pressure, and blood K+ levels (p < 0.05). Convulsions (focal and general) present in Group O were absent in groups I, H and E. Cardiac arrest (CA) occurred 2 to 3 times faster in pigs receiving VA agents. In all groups studied the harbinger of serious cardiac decompensation was the development of complete heart block (CHB). The time to CA after the onset of CHB was no different between groups. It is proposed that the hastened onset of CA in animals receiving VA is caused by a rapid onset of CHB due to an additive effect of cocaine and VA agents in prolonging atrioventricular conduction.
