N-methylation of normorphine by rat tissues in vitro

A N-methyl transferase that catalyzes the transfer of methyl groups from S-adenosylmethionine to normorphine has been purified from rat liver and brain. The enzymic activity purified from liver is five times more active than that from brain. However, preparations from hypothalamus and adjacent structures of brain are more active than those from liver. Evidence on the identity of this enzymic activity with nicotinamide methylferase is not conclusive at the present time.     

N-methylation of benzimidazole by catechol-O-methyltransferase

The substrate specificity of catechol-O-methyltransferase (COMT) has been extended in the present study to include benzimidazole, a non-catechol compound. Gas chromatography-mass spectrometry. thin-layer chromatography, and reverse isotope dilution analysis were used to verify the identity of the product of this reaction as 1-methylbenzimidazole. This is the first demonstration of an N-methylation by COMT. The conclusion is reached that benzimidazole and catechol are bioisosteric molecules. The implication of this in the pharmacology of adrenergic systems is discussed.     

Bioavailability of quinidine in congestive heart failure.

1 The oral bioavailability of quinidine was evaluated in eight patients with moderate to severe congestive heart failure. Each patient was given a 400 mg dose of quinidine gluconate by intravenous infusion and orally in solution. Serial plasma samples and total urine for drug analysis were collected for 24 and 48 h after drug administration, respectively. 2 When compared to control cardiac patients, the rate of quinidine absorption was slower in the heart failure patients. The mean value for the apparent absorption half-life and time to achieve peak plasma quinidine concentration was 38 +/- 18 min and 2.4 +/- 1.5 h respectively. The corresponding values observed in the control subjects were 18 +/- 6 min and 1.0 +/- 0.6 h. 3 The extent of quinidine absorption when evaluated by the AUC and urinary excretion methods was about 72% of the administered dose in the congestive heart failure patients. This value was similar to the extent of quinidine absorption (approximately 73%) observed in the control subjects. 5 When compared with non-heart failure cardiac patients, the results of this study suggest that patients with congestive heart failure may require smaller oral quinidine dosages to achieve therapeutic drug concentrations in the plasma or serum.     

Phosphatidylethanolamine N-methylation and insulin release in isolated pancreatic islets of the rat

Rat pancreatic islets methylate phosphatidylethanolamine (PE) lipids to form phosphatidylcholine (PC) with S-adenosyl-L-[methy-3H]methionine as the methyl donor. Islet PE-N-methyltransferase had activity optima at pH 6-7 and 8-9. S-Adenosyl-L-homocysteine, sodium deoxycholate, and Triton X-100 inhibited methylation in islet homogenates. Addition of phosphatidyl-N-monomethylethanolamine and phosphatidyl-N,N-dimethylethanolamine (PDME) enhanced [3H]methyl incorporation into PDME and PC, respectively. Isoproterenol, but not glucose, stimulated phospholipid methylation in islet homogenates. Propranolol inhibited the isoproterenol effect. In intact islets, glucose or isoproterenol stimulated insulin release and incorporation of [3H]methyl groups from [methyl-3H]methionine into phospholipids. Isoproterenol enhanced to a similar extent glucose-stimulated methylation and hormone release. Neither 2-deoxyglucose, tolbutamide, nor 8-bromo-cyclic AMP stimulated islet phospholipid methylation. The methyl-transferase inhibitor 3-deazaadenosine inhibited both glucose and isoproterenol-stimulated methyltransferase activity and insulin release. Propranolol inhibited the beta-adrenergic potentiation of glucose-induced phospholipid methylation and insulin release. These data suggest that PE-N-methyltransferase plays a role in amplification of the islet cell stimulus-secretion coupling response to certain secretagogues.     

Quinine impairs quinidine clearance in rat perfused liver

Abstract— We have examined the disposition of the cinchona alkaloids quinine and quinidine in the rat recirculating isolated perfused liver preparation. When administered as separate 1 mg doses, the hepatic clearances of quinine and quinidine were similar to the hepatic perfusate rate of 10 mL min^?1. When 1 mg of each was administered simultaneously, mean hepatic clearance of quinine was unchanged (9·00 ± 2·20 mL min^?1 separate dosage, n = 7; 6·87 ± 1·77 mL min^?1 simultaneous dosage, n = 7; P > 0·05). By contrast, mean hepatic clearance of quinidine was reduced significantly by concomitant quinine (10·6 ± 1·72 mL min^?1 separate dosage, n = 7; 4·82 ± 1·25 mL min^?1 simultaneous dosage, n = 7; P < 0·05). There was no significant difference in volumes of distribution when each alkaloid was administered separately (131 ± 46 mL quinine, 129 ± 21 mL quinidine; P > 0·05) but concomitant quinine administration increased quinidine volume of distribution to 169 ± 30 mL (P < 0·05). Four further experiments with simultaneous dosages of 0·5 mg of each alkaloid produced similar findings, indicating that the interactions did not derive from nonlinear drug disposition.     

Compared efficacy of quinidine, amiodarone and diltiazem on both nomotopic and ectopic automaticity in the rat heart

The effect of quinidine, amiodarone and diltiazem have been studied on both nomotopic (isolated right atria) and ectopic automaticity (induced by a local injury in the isolated right ventricle) in the rat heart. The efficacy of these three drugs, measured by the corresponding ID25, was diltiazem greater than quinidine greater than amiodarone on atrial rate and amiodarone greater than quinidine greater than diltiazem on ectopic ventricular automaticity. A depression of isolated right atria rate (sinus node function) could be regarded as an adverse effect of the drug tested (toxicity-index). On the other hand, the reduction of ectopic ventricular automaticity by the antiarrhythmic agent is considered as desired effect (effectiveness-index). We conclude that this method might be useful to evaluate the effectiveness-toxicity ratio, as a first experimental step in the development of new antiarrhythmic drugs with a direct action on cardiac tissue.     

Alterations by age of calcium handling in rat resistance arteries

This study investigated the alterations by age of Ca handling in rat small mesenteric arteries. The contractile responses to phenylephrine and caffeine in small mesenteric arteries from young and old rats were studied in Ca -containing and Ca -free medium. In Ca -containing medium the contraction to phenylephrine (100 micro M) but not to caffeine (10 mM) was greater in old than in young rats. Concentration-response curve to phenylephrine was affected to the same extent by nifedipine (1 micro M) in both age groups, whereas ryanodine (20 micro M) decreased the maximal response to phenylephrine only in young rats. These results suggest the participation of intracellular Ca handling on the observed differences by aging. In Ca -free medium, phenylephrine (10 micro M) but not caffeine (10 mM) induced a greater contraction in old than in young animals, corroborating the results obtained in Ca -containing solution. The greater response to phenylephrine observed in old rats cannot be explained by an increase in the inositol 1,4,5-trisphosphate (IP ) formation because the accumulation of inositol phosphates by phenylephrine was not affected by aging. Results obtained in Ca -free medium using caffeine after phenylephrine or vice versa suggest a common intracellular Ca pool. Pretreatment with ryanodine in Ca -free medium almost abolished contractile response to phenylephrine and caffeine in young rats but only partially decreased them in old animals, suggesting an impairment in the Ca -induced Ca release (CICR) mechanism leading to an increase in the stored Ca content. The greater amount of stored Ca could explain the higher contractile response to phenylephrine observed in aged rats. As a consequence of all these changes due to aging, an imbalance between the two Ca release mechanisms from sarcoplasmic reticulum was observed with a major role of Ca induced release by IP at the expense of an impairment of CICR mechanism. This observation will also help explain the results obtained in the presence of extracellular Ca, where phenylephrine induced a greater maximum response in old animals in spite of a decrease in the midrange sensitivity to this agonist.     

Concentration-dependent exsorption of quinidine in the rat intestine.

During intravenous infusion, the luminal concentration of quinidine was higher than the plasma concentration. The intestinal clearance (CLi) of the drug was measured by dividing the rate of appearance of the drug in the intestinal luminal perfusate by the plasma concentration. The CLi of quinidine was therefore much higher than the rate of luminal perfusion. Over the infusion dose range of 0.1-2 mg h-1, the CLi of quinidine decreased with increasing plasma concentration of quinidine. Adding quinidine into the luminal perfusate had little effect on the CLi of quinidine. Co-administration of quinidine with other agents intravenously did not alter the CLi of salicylic acid and urea, while the same treatment decreased the CLi of theophylline and S-disopyramide. In-vitro experiments on brush-border membrane vesicles showed that quinidine decreased the rate of Na+ uptake and H+ efflux. The inhibition was significant at quinidine concentrations above 20 microM. Quinidine was a more potent inhibitor than amiloride. At quinidine infusion rates less than 2 mg h-1, quinidine concentration in plasma or in the luminal perfusate was at the lower limit of the inhibitory concentration. Microclimate pH at the intestinal surface was also measured. At mid-jejunum, the microclimate pH increased 0.3 pH units by infusing 2 mg h-1 of quinidine, while the microclimate pH at most other measuring sites was not significantly altered by quinidine infusion. It was concluded that quinidine is exsorbed from blood into the intestinal lumen by a carrier-mediated pathway in addition to the passive diffusion. At high plasma concentration, quinidine exsorption becomes saturated.(ABSTRACT TRUNCATED AT 250 WORDS)     

N-methylation of nicotine enantiomers by human liver cytosol

Incubation of human liver cytosol with either R-(+)-[3H-N'CH3]nicotine or S-(-)-[3H-N'CH3]nicotine results in the formation of the corresponding N-methyl quaternary ammonium metabolite. A substrate stereoselectivity was observed in that the turnover number for the methylation of the S-(-)-isomer was 0.25 pmol mg-1 protein h-1, whereas that for the R-(+)-isomer was 2.11. The latter substrate exhibited an apparent Km value of 20.1 microM. Nicotine N-methylation appears to be species-dependent, since rat liver homogenates contained no 'nicotine N-methyltransferase' activity, whereas with guinea-pig liver homogenates, a substrate specificity for only R-(+)-nicotine was observed.     

Comparative study of the block of Vmax by aprindine and quinidine in the guinea-pig heart muscle

The depression of Vmax of the action potential in guinea-pig ventricular muscle by aprindine and quanidine was compared. Aprindine caused a more pronounced rate-dependent block (Kd = 10(-6) M at 3.3 Hz) than did quinidine (Kd = 1.6 X 10(-5) M at 3.3 Hz). Aprindine shifted the relationship between Vmax and resting potential to a more negative potential (mean 9.2 mV: 10 microM) than did quinidine (mean 5.7 mV: 10 microM). In addition, aprindine caused a more pronounced resting block (Kd = 1.3 X 10(-5) M) than quinidine (Kd = 8.6 X 10(-5) M). It is concluded that aprindine has a higher affinity for activated and/or inactivated and resting state channels than quinidine, making channels unavailable for conduction upon activation.     

Alterations induced in vitro by ochratoxin A in rat lymphoid cells

Ochratoxin A (OTA) is a nephrotoxic mycotoxin produced by species of the genus Penicillium and Aspergillus that is present in food and feed as a natural contaminant. It modifies the immune function in animals and inhibits the proliferative response of lymphocytes in vitro. The toxic effect of OTA (0.5, 2, 20 microM) in lympho-proliferative response, natural killer (NK) cell activity, cytotoxic T lymphocytes (CTL) activity and macrophages' bacteriolytic capability was studied in vitro after 1 hour of treatment. The proliferative response of lymphocytes to concanavalin A and lipopolysaccharide was not affected by OTA; the cytotoxic activity of NK cells was dose-dependent decreased; the CTL activity was significantly decreased at the lowest concentration; the bacteriolytic activity of macrophages varied only slightly. These in vitro results reproduced, at least in part, some effects detected previously in vivo. The protein synthesis inhibition and the oxidative metabolism of OTA coupled to the prostaglandin synthesis are probably implicated in NK cells' toxicity, because the effects were reverted by the addition of phenylalanine or piroxicam to the culture medium. The induction of apoptosis seems to be the principal mechanism of action in the CTL effect. The intracellular concentration of OTA after 1 hour was analysed by HPLC and was found to be proportional to the quantity of OTA added to the culture medium for the three cell types; the presence of phenylalanine and piroxicam on the culture medium did not change the intracellular OTA concentration.     

Selective in vivo inhibition by quinidine of methoxyphenamine oxidation in rat models of human debrisoquine polymorphism

1. Lewis and Dark Agouti (DA) rat strains (n = 4), models of human extensive and poor metabolizer phenotypes of debrisoquine/sparteine, respectively, were dosed with methoxyphenamine with and without prior administration of quinidine. Methoxyphenamine and its three metabolites, namely N-desmethylmethoxyphenamine, O-desmethylmethoxyphenamine and 5-hydroxymethoxyphenamine were quantified in 0-24 h urine. 2. The oxidative metabolic routes of methoxyphenamine which had been previously shown to involve the debrisoquine/sparteine isozyme, namely O-demethylation and aromatic 5-hydroxylation, were both significantly inhibited by quinidine in the two rat strains. 3. The oxidative metabolic route of methoxyphenamine which had been previously shown to not involve the debrisoquine/sparteine isozyme, namely N-demethylation, was not significantly inhibited by quinidine in either rat strain. 4. The Lewis strain pretreated with quinidine resembled the DA strain without such pretreatment in terms of O-desmethylmethoxyphenamine and 5-hydroxymethoxyphenamine in that the mean percentages of the dose excreted as these two metabolites and the mean O-desmethylmethoxyphenamine/methoxyphenamine and 5-hydroxymethoxyphenamine/methoxyphenamine ratios were similar to one another. 5. Ten days after quinidine administration to the Lewis strain of rat, all parameters of methoxyphenamine and its metabolites returned to normal. 6. A protocol involving substrate administration to Lewis strain rats with and without prior administration of quinidine could be developed as an attractive approach to screen substrates for metabolism in vivo by the debrisoquine/sparteine isozyme. Such an approach obviates interstrain differences.     

Pharmacokinetics of dihydroquinidine in congestive heart failure patients after intravenous quinidine administration

Summary The pharmacokinetics of dihydroquinidine were studied in 8 patients with congestive heart failure following a 22 min intravenous infusion of a quinidine preparation that contained 5.9% dihydroquinidine as an impurity. Using a thin layer chromatography-fluorometric assay procedure for dihydroquinidine, the post-infusion plasma dihydroquinidine concentrations declined biexponentially. The half-life of the fast and slow dispositional processes was 4.42±1.81 min and 6.52±2.40 h, respectively. The central compartment volume for dihydroquinidine in these patients was 0.44±0.11 l/kg with an overall apparent volume of distribution of 1.14±0.38 l/kg. The computed values of total body plasma clearance of dihydroquinidine ranged from 1.29 to 2.69 ml/min/kg with a mean value of 1.94±0.60 ml/min/kg. In these patients, approximately 16% of the administered dihydroquinidine dose was excreted intact into the urine in 48 h. The estimated value of renal clearance was 0.314±0.129 ml/min/kg. When compared to control cardiac patients, the data showed that the apparent volume of distribution for dihydroquinidine is smaller in patients with congestive heart failure and as a result of this diminished volume, the clearance rate of dihydroquinidine was slower. The net effect of these differences was the production of higher plasma concentrations of dihydroquinidine in the heart failure group.     

Alterations in renal function induced by aflatoxin B1 in the rat

The effects of aflatoxin B1 (AFB1) on renal function were determined in adult male Wistar rats by in vivo and in vitro studies. In vivo studies demonstrated that AFB1 decreased glomerular filtration rate, tubular reabsorption of glucose, and tubular transport for p-amino-hippurate. Furthermore, AFB1 pretreatment increased urinary excretion of sodium and potassium and urinary gamma-glutamyl transferase content. In vitro studies showed that slices of renal cortical tissue obtained from AFB1-treated rats (100 microgram/kg) exhibited a diminished capacity to accumulate p-aminohippurate. On the other hand, total water content of tissue, amount of extracellular water, and intracellular sodium were increased. Intracellular potassium was diminished by treatment with AFB1. Although the molecular species responsible for the effects is not known, these results indicated that AFB1 is nephrotoxic in the rat following a single low dose (100 microgram/kg body wt, ip).     

Alterations to the electrical activity of atrial muscle isolated from the rat heart, produced by exposure in vitro to amiodarone.

Glass microelectrodes were used to record transmembrane electrical activity from cells located just beneath the endocardial surface of the left atrial free wall of rat hearts during superfusion and electrical stimulation in vitro at 37 degrees C. Availability of the fast sodium channel for current flow was inferred from the maximum rate of rise of membrane potential during phase O of the action potential (Vmax). Muscle exposed to polysorbate 80 (10 to 80 micrograms ml-1) showed a concentration-dependent lengthening of action potential duration (APD) but no detectable change in Vmax. Amiodarone (1 to 20 micrograms ml-1) was dissolved in physiological salt solution with the aid of polysorbate 80 (50 micrograms ml-1) and caused a concentration-dependent prolongation of APD and a decrease in Vmax, both of which were slow to develop and extremely slow to wash-out. The speed of onset of action of amiodarone varied with drug concentration and ranged from a few minutes with high concentrations to many hours with low concentrations.     

Block of NA+ and K+ currents in rat ventricular myocytes by quinacainol and quinidine

1. The electrophysiological actions of quinacainol were investigated on sodium (I(Na)), transient outward (i(to)) and sustained-outward plateau (iKsus) potassium currents in rat isolated cardiac myocytes using the whole-cell patch-clamp technique and compared with quinidine. 2. Quinacainol blocked sodium currents in a concentration-dependent manner and with a potency similar to that of quinidine (mean (+/-SEM) EC50 50+/-12 vs 95+/-25 micromol/L for quinidine and quinacainol, respectively). However, quinacainol had a considerably prolonged onset and recovery from block compared with quinidine. 3. Neither quinacainol nor quinidine significantly changed the steady state voltage dependence of activation of sodium currents. Quinidine produced a hyperpolarizing shift in the voltage dependence for sodium current inactivation, but no such shift was observed with quinacainol at doses that produced a substantial current block. 4. Although quinacainol did not effectively block voltage-dependent potassium currents, even at concentrations as high as 1.5 mmol/L, quinidine, at a half-maximal sodium channel-blocking concentration, reduced peak i(to) current amplitude, increased the rate of inactivation of i(to) and blocked iKsus. 5. These results indicate that quinacainol, a quinidine analogue, blocks sodium currents in cardiac myocytes with little effect on i(to) or iKsus potassium currents, which suggests that quinacainol may be exerting class 1c anti-arrhythmic actions.