R.Craig Schnell

Studies on the mechanism of cadmium-induced inhibition of the hepatic microsomal monooxygenase of the male rat.

Cadmium is a potent inhibitor of hepatic drug metabolism but little is known about the biochemical mechanisms involved. Male rats receiving a single dose of cadmium acetate (2.0 mg Cd2+/kg, ip) exhibited significant decreases in hepatic microsomal metabolism of hexobarbital (79%), ethylmorphine (71%), and aniline (47%), as well as decreased amounts of both microsomal cytochrome P-450 (39%) and cytochrome b5 (30%) 72 hr after administration of the metal. In addition, the magnitudes of microsomal hexobarbital, ethylmorphine, and aniline binding spectra were significantly reduced. NADPH-cytochrome c reductase activity was not altered. In vitro addition of Cd2+ (10−6 to 10−3 m) to liver microsomes produced a concentration-dependent inhibition of the metabolism of the substrates tested, a reduction in the level of cytochrome P-450 which was accompanied by an increase in the level of the inactive form of the hemo-protein, P-420, and decreases in the microsomal binding spectra of the three substrates. In kinetic studies, apparent Vmax values were significantly lowered for ethylmorphine N-demethylase (75%) and aniline hydroxylase (57%) by cadmium treatment. The apparent Km value for aniline hydroxylase was not altered, but that for ethylmorphine N-demethylase was significantly decreased following cadmium treatment. After in vitro cadmium addition, concentration-dependent decreases were observed in the apparent Vmax and Km values for both microsomal reactions.

Cadmium potentiation of drug response—Role of the liver☆

Abstract Three days after a single cadmium dose (2 mg/kg), the duration of response to subsequently administered hexobarbital or zoxazolamine was potentiated. Duration of sleep was prolonged (225 per cent) in cadmium-treated rats, but the awakening plasma hexobarbital levels were similar in both the cadmium-treated animals and saline-treated controls. Moreover, cadmium-treated animals exhibited significantly higher plasma hexoharbital levels when sacrificed prior to awakening at a time corresponding to the mean duration of sleep in control rats, thus suggesting that the cadmium effect may be mediated by a decline in the plasma disappearance of hexobarbital. Duration of hexobarbital sleep was not changed in cadmium-treated hepatectomized rats compared to hepatectomized control animals, indicating the necessity of an intact liver for the cadmium effect to be elicited. In addition, pretreatment with cadmium significantly inhibited metabolism of hexobarbital in vitro in the isolated perfused rat liver (65 per cent). After the cadmium treatment zoxazolamine-induced paralysis was prolonged (185 per cent), but the plasma drug levels measured upon recovery were significantly lower. Significantly higher plasma levels of zoxazolamine were found in cadmium-treated rats when they were sacrificed prior to recovery at a time corresponding to the mean duration of paralysis in control rats. These results thus infer that the cadmium interaction with zoxazolamine may involve both an alteration in drug disposition as well as a change in tissue responsiveness.

Selenium antagonism of cadmium-induced inhibition of hepatic drug metabolism in the male rat

Abstract Experiments were undertaken to examine the effect of selenium, an essential trace element, on cadmium-induced inhibition of drug metabolism in male, Sprague-Dawley derived rats. Prior administration of sodium selenite (1.6 mg Se/kg, ip) blocked the cadmium-induced (0.84 mg Cd/kg, ip) prolongation of hexobarbital-induced hypnosis and inhibition of hepatic microsomal biotransformation of ethylmorphine or aniline. Selenium also blocked cadmium-induced reduction in microsomal cytochrome P-450 content and the microsomal binding of both ethylmorphine and aniline. However, pretreatment of rats with selenium did not prevent the inhibitory effect of cadmium (10−6 to 10−3 m ) added in vitro on either ethylmorphine or aniline biotransformation. In addition, the reduction in biotransformation of both substrates following in vivo cadmium administration was not reversed following the in vitro administration of selenium but, in fact, selenium produced further concentration-dependent decreases in drug metabolism. In additional in vitro experiments it was found that the inhibition in drug metabolism induced by in vitro additions of cadmium is not affected by similar additions of selenium when added to the incubation vessel either before or after the cadmium. Thus, for selenium to prevent the cadmium-induced inhibition of hepatic drug metabolism requires in vivo administration of selenium.

Cadmium alteration of hexobarbital action: sex-related differences in the rat.

Abstract Three days after ip administration of a single cadmium dose (0.84 mg of Cd/kg), duration of hexobarbital hypnosis was significantly potentiated in male but not in female rats. Cadmium in larger doses of 2, 4, and 6 mg/kg, ip, did not alter duration of hexobarbital hypnosis in female rats, although lethality was observed 24 hr after administration of the two higher cadmium doses. Hepatic microsomal metabolism of hexobarbital was significantly decreased in male rats but not in females when measured 72 hr after cadmium administration (2 mg/kg, ip). Addition of cadmium in concentrations ranging from 5 × 10−6 m to 5 × 10−3 m to microsomes isolated from untreated male and female rats produced a dose-dependent inhibition of hexobarbital metabolism at all Cd concentrations in males. In female-derived microsomes a significant inhibition of hexobarbital metabolism was observed only at higher cadmium concentrations (5 × 10−4 m and 5 × 10−3 m ). These data suggest that a sex-related difference exists in the ability of cadmium to alter response to hexobarbital in the rat which appears to be mediated through differences in hepatic microsomal metabolism.

Cadmium-induced inhibition of hepatic drug oxidation in the rat: time dependency of tolerance development and metallothionein synthesis.

Abstract Experiments were undertaken to determine if a correlation existed between the time-dependent synthesis of hepatic metallothionein and the development of tolerance to the inhibition of hepatic drug oxidation elicited by cadmium in the male rat. Maximal rates of metallothionein synthesis were achieved within 2 hr after administering cadmium in a 0.21 mg Cd/kg (ip) dose. The total hepatic concentration of metallothionein, as well as the cadmium binding capacity of this protein, also increased rapidly with maximal values being observed from 8 to 67 hr after cadmium administration. Despite this rapid increase in hepatic metallothionein levels, the tolerance to the inhibition of in vivo drug oxidation induced by a challenge cadmium (0.84 mg Cd/kg) dose did not develop until 16 hr after treatment with the 0.21 mg/kg cadmium dose. Furthermore, hepatic metallothionein levels decreased from a maximum at 67 hr to approximately control levels at 336 hr. Although the tolerance to the inhibition of drug oxidation also decreased from a maximum at 72 hr a modest degree of protection was maintained even at 336 hr. Pretreatment with the 0.21 mg Cd/kg dose increased the hepatic uptake of a 109 Cd challenge dose (0.84 mg Cd/kg). This increase was associated with an increased 109 Cd binding to metallothionein in the cadmium-pretreated animals. While these data are suggestive of a role for metallothionein in tolerance development, the lack of correlation of the time course of metallothionein synthesis with the development of the tolerance would suggest that factors in addition to metallothionein may also participate.

Studies relating morphine hypothermia with serotonin reuptake in the rat hypothalamus

Experiments were conducted to determine if the acute hypothermia elicited by morphine in the rat resulted from morphine-induced inhibition of serotonin reuptake into hypothalamic nerve-endings. The acute hypothermia elicited by morphine (30 mg/kg, s.c.) was reversed by pretreatment with naloxone (10 mg/kg, s.c.), a narcotic antagonist, which alone did not alter body temperature. In vitro uptake of serotonin into synaptosomes prepared from rat hypothalamus was inhibited 53% by the addition of morphine (7 X 10(-4) M) and 53.9% by naloxone (1 X 10(-3) M). Simultaneous addition of both drugs in these same concentrations further inhibited synaptosomal serotonin uptake 77.8%. Uptake of serotonin uptake into synaptosomes isolated from rats injected with morphine (30 mg/kg, s.c.) was not altered as compared to controls. These data suggest that the acute hypothermic acition in morphine in the rat is not related to an inhibition of the serotonin reuptake mechanism in hypothalamic nerve-endings.

Sex-related differences in cadmium-induced alteration of drug action in the rat

3 days after pretreatment of rats of both sexes with cadmium (2 mg/kg, 1.p.) the duration of hypnosis induced by hexobarbital (75 mg/kg, 1.p.) was potentiated in males but not females. Likewise similar treatment with cadmium leads to significant inhibition of the metabolism of hexobarbital by hepatic microsomal enzymes obtained from male but not female animals. These data suggest that there is a sex-related difference in the ability of cadmium to alter drug action in rats.

Effect of acute and repeated selenium treatment on hepatic monooxygenase enzyme activity in male rats

The effect of sodium selenite administered acutely or repeatedly on the biochemical components of the hepatic microsomal monooxygenase enzyme system was examined in male rats. 72 h following acute administration of selenium (2.4 mg Se/kg, i.p.), there was a significant decrease in ethylmorphine-N-demethylase activity and cytochrome P-450 levels but no change in aniline hydroxylase or NADPH cytochrome c reductase activity. Following repeated administration of selenite in the drinking water (1, 2, or 4 ppm Se) for 30 days, there was no alteration in any of the parameters measured. Following the in vitro additions of selenite to microsomes obtained from untreated rats, ethylmorphine-N-demethylase and aniline hydroxylase activities were inhibited at selenium concentrations of 10(-4) M or greater, but the inhibition achieved was less than 50%. No alterations in cytochrome P-450 levels were observed. These results indicate that selenium is a rather weak, indirect, and substrate-specific inhibitor of the hepatic monooxygenase enzyme system.

Effect of glutathione depletion on selenium lethality and hepatic drug metabolism in male rats

Pretreatment of male rats with diethylmaleate (DEM) (3.1 mmol/kg) markedly decreased the levels of reduced glutathione in both liver (81%) and kidney (44%). Acute lethality produced by sodium selenite (0.4 to 2.4 mg Se/kg) was greatly enhanced in DEM-treated rats; however, no alterations in hepatic drug metabolism by selenium were observed.

Cadmium-induced alteration of the microsomal mono-oxygenase system in male rat liver: Effects on hemoprotein turnover and phospholipid content

Abstract After cadmium treatment (2.0 mg Cd/kg, i.p.) a marked inhibition is observed in microsomal drug metabolizing enzyme activity which is correlated with a reduction in the hemoprotein, cytochrome P-450, content. Incorporation of the heme precursor, 3H-δ-aminolevulinic acid (3H-δ-ALA), into microsomal CO-binding particles was decreased in cadmium-treated rats. Cadmium treatment produced a differential effect in the biphasic disappearance of radioactivity from the CO-binding particles in that the metal decreased the half-life of the fast-phase component and increased the half-life of the slow-phase component. Thus, the reduction in microsomal levels of cytochrome P-450 induced by cadmium apparently results from a stimulation of the degradation of the fast-phase hemoprotein fraction and a reduction in the synthesis of the slow-phase and fast-phase hemoprotein fractions. Cadmium treatment did not quantitatively alter microsomal phospholipid content.