Hepatic Microsomal Ethylmorphine N-demethylase Research Articles

Studies on the acute effects of coumarin and some coumarin derivatives in the rat

The mechanism of acute coumarin-induced hepatotoxicity in the rat has been investigated by comparing the effects of coumarin with those of a number of methyl-substituted coumarin derivatives. Male Sprague-Dawley rats were given single ip doses of corn oil (control), coumarin (0.86 and 1.71 mmol/kg body weight), 3,4-dimethylcoumarin (3,4-DMC, 1.71 and 2.57 mmol/kg), 3−, 4− and 6-methylcoumarins (3-MC, 4-MC and 6-MC, 1.71 mmol/kg) and 3− and 4-methyloctahydrocoumarins (3-MOHC and 4-MOHC, 2.57 mmol/kg) and hepatotoxicity assessed after 24 hr. Coumarin administration produced dose-related hepatic necrosis and a marked elevation of plasma alanine aminotransferase and aspartate aminotransferase activities. In contrast, none of the coumarin derivatives examined produced either hepatic necrosis or elevated plasma transaminase activities. Treatment with coumarin reduced hepatic microsomal ethylmorphine N-demethylase 7-ethoxycoumarin O-deethylase activities, whereas one or both mixed-function oxidases appeared to be induced by treatment with 3,4-DMC, 4-MC, 3-MOHC and 4-MOHC. These results provide further evidence that acute coumarin-induced hepatotoxicity in the rat is due to the formation of a coumarin 3,4-epoxide intermediate. That 3- and/or 4-methyl substitution (i.e. 3-MC, 4-MC and 3,4-DMC) leads to a reduction in coumarin-induced hepatotoxicity, due to diminished formation of 3,4-epoxide intermediates, was confirmed by the results of molecular orbital calculations.

Correlation of hepatic cytosolic androgen binding proteins with androgen induction of hepatic microsomal ethylmorphine N-demethylase in the rat

Previous reports have demonstrated the presence of moderate to high affinity binding for androgens in the cytosol of livers from male rats. This binding was significantly lower in female rats or in immature rats of either sex. The hepatic androgen binding protein, which sedimented at approx. 4 S on sucrose density gradients, has been called a receptor which mediates the actions of androgens in the liver. The experiments in the present study were designed to evaluate the hepatic androgen binding protein for characteristics which have been attributed to receptors in other tissues and to correlate the presence of androgen binding with androgen induction of hepatic drug metabolism. In the current studies, we have shown that cytosol from the livers of male rats bound [ 3H]dihydrotestosterone ([ 3H]DHT) and translocated this steroid ligand to the nucleus in a time and temperature dependent manner. Cytosol prelabeled with [ 3H]DHT, when passed over a column of denatured DNA cellulose, eluted in three radioactive peaks. Two of these peaks were absent when cytosol from livers of female or hypophysectomized males was used. In addition, the presence of high concentrations of hepatic androgen binding correlated well with the ability of androgen to induce ethylmorphine N-demethylase, a marker of microsomal cytochrome P-450-dependent drug metabolism. Values for both parameters were higher in males than in either females or hypophysectomized males. Testosterone treatment induced both parameters in ovariectomized females and 17β-estradiol repressed both in males. However, testosterone treatment failed to induce hepatic androgen binding in hypophy-sectomized males and immature males, both of which are also unresponsive to androgen induction of drug metabolism. The results suggest that one or more hepatic cytosolic androgen binding proteins possess several characteristics associated with steroid receptors in reproductive tract tissue. Furthermore, this binding may be implicated as a mediator for the androgen induction of at least one component of hepatic drug metabolism.

The effect of NADH and low oxygen pressure on the hepatic microsomal ethylmorphine N-demethylase activity of the male rat.

Plots of reciprocal initial rate of formaldehyde formation as a function of reciprocal ethylmorphine concentration provide parallel lines for oxygen concentrations ranging from 5.41 to 211 microM, and the apparent Km values for ethylmorphine at these extremes are 167 and 417 microM, respectively. Similarly, the apparent Km for O2 decreases from 7.69 microM at 2 mM ethylmorphine to 3.64 microM at 0.2 mM ethylmorphine. Reciprocal plots of 1/Km app and 1/Vm for ethylmorphine against 1/[O2] give Km values for O2 of 9.35 microM and 9.09 microM, respectively. Similar plots give Km values for ethylmorphine of 0.28 mM and 0.36 mM. Similarly, NADH causes an uncompetitive stimulation. These data suggest that hepatic microsomal ethylmorphine N-demethylase follows parallel plot sequential kinetics. These results, along with studies of other workers, suggest that cytochrome P-450 forms an active complex with O2 which is stable for at least many milliseconds.

Bleomycin Inhibits Hepatic Mixed Function Oxidation

Bleomycin inhibits hepatic microsomal ethylmorphine N-demethylase activity in both the athymic (nude) mouse and the rat four days after a single injection. This inhibition occurred both with subcutaneous and intraperitoneal injections. With multiple dose chronic treatment, s.c. injections resulted in decreases in both enzymatic activity and cytochrome P450 concentration in both species. However, i.p. chronic injection resulted in decreases only in the rat.

Interaction of pargyline with rat hepatic microsomes.

When administered by intraperitoneal injection daily for 3, 7 or 14 days, pargyline (75 mg/kg) significantly reduced rat hepatic microsomal ethylmorphine N-demethylase activity and cytochrome P-450 content. Injection of a lower dose of pargyline (15 mg/kg) failed to alter significantly either ethylmorphine N-demethylase activity or cytochrome P-450 content. Studies performed in vitro reveal that pargyline is metabolized to at least three compounds by rat hepatic microsomes. Thin-layer chromatography and other analyses suggest that one metabolite is an N-demethylated form, norpargyline.

Characteristics of neonatal androgen-induced imprinting of rat hepatic microsomal monooxygenases

Both the apparent K m of hepatic microsomal ethylmorphine N-demethylase and the responsiveness of this enzyme to testosterone treatment in adult rats are imprinted neonatally by androgen. The maintenance of these imprinted male-specific characteristics of this enzyme activity depends on the presence of an intact pituitary. Hypophysectomy alters the apparent K m of this enzyme activity from male into female type, whereas castration, thyroparathyroidectomy or adrenalectomy does not. Androgen or estrogen administration has opposite effects on the apparent K m , but similar effects on plasma FSH and LH concentrations. These results tend to mitigate against mediator roles for ACTH, TSH, LH and FSH. The sensitive periods for androgen-induced imprinting of the apparent K m of ethylmorphine N-demethylase differs from that of the responsiveness of this enzyme. With the former, imprinting occurs at days 2 and 4 of age, whereas the latter extended beyond days 2 and 4 to days 12 and 14 of age. Unlike imprinting in the CNS, neonatally administered estradiol-17β is ineffective in inducing the male characteristics of this enzyme activity. Although hypophysectomy alters the imprinted male characteristics of the ethylmorphine N-demethylase (apparent K m and responsiveness) to the female type, it shifts the neonatal androgen-imprinted differences in the turnover of cytochrome P-450 from female toward that of the male type. These results suggest that different hypophyseal factors may be responsible for the regulation of these neonatal androgen-imprinted male characteristics in hepatic microsomal monooxygenases.

Androgen receptor dependent and independent activities of testosterone on hepatic microsomal drug metabolism.

Administration of testosterone for 6 days to intact female and castrate male BALB/cJ mice stimulated hepatic microsomal ethylmorphine N-demethylase activity and cytochrome P-450 content by 50-75%. Testosterone also stimulated hepatic microsomal NADPH-oxidase activity, but to a lesser degree. To probe the mechanism of this effect of androgens, two antiandrogens (cyproterone acetate and flutamide) were employed. Since cyproterone acetate was a potent stimulator of hepatic microsomal ethylmorphine N-demethylase activity and cytochrome P-450 content, no antiandrogenic activity of this steroid could be detected. By contrast, flutamide alone had little effect on either ethylmorphine N-demethylase activity or cytochrome P-450 content. However, this drug effectively blocked the stimulatory effects of testosterone on ethylmorphine N-demethylase activity and cytochrome P-450 content but not on NADPH-oxidase activity. This effect was not species specific, since flutamide also prevented androgen stimulation of ethylmorphine metabolism in adult castrate and prepubertal male Fisher rats. The testosterone-induced increase of hepatic weight and microsomal protein content was not affected by the administration of flutamide. The observations are consistent with the hypothesis that androgens have two distinct effects on the liver. First, testosterone may act as a general, nonspecific stimulant of liver weight and microsomal protein content which is independent of the androgen receptor. Secondly, testosterone action in the liver may be expressed via an androgen-specific or androgen receptor-dependent mechanism which controls, in part, the cytochrome P-450-dependent demethylase system.

Phenobarbital induced effect on pulmonary and hepatic microsomal ethylmorphine N-demethylase and lipid peroxidation during oral intoxication of organic solvents in rats.

Phenobarbital induced effect on pulmonary and hepatic microsomal ethylmorphine N-demethylase and lipid peroxidation during oral intoxication of organic solvents in rats.

Dependence of Km and Vmax on substrate concentration for rat hepatic microsomal ethylmorphine N-demethylase

Dependence of Km and Vmax on substrate concentration for rat hepatic microsomal ethylmorphine N-demethylase