William F. Bousquet

Correlation of hepatic metallothionein concentrations with acute cadmium toxicity in the mouse

Abstract Dose-dependent induction of metallothionein by cadmium was measured by gel filtration of liver supernatants prepared from mice 48 hr following single doses of 0.2–2.0 mg of Cd/kg (ip). Hepatic metallothionein concentrations increased in proportion to the cadmium pretreatment dose. A dose-dependent tolerance to acute cadmium toxicity was simultaneously evidenced by elevated LD50 values in cadmium-pretreated mice. A positive correlation between dose-related increases in hepatic metallothionein and cadmium LD50 suggests a protective function for metallothionein. The data suggest that a threshold dose of cadmium must be exceeded in order to induce concentrations of metallothionein adequate to ameliorate acute cadmium toxicity.

Cadmium inhibition of hepatic drug metabolism in the rat

Pretreatment of rats and mice of either sex with a single 2.0 mg/kg dose of cadmium acetate (840 μg Cd ion) potentiates the action of subsequently administered hexobarbital. Three days after cadmium administration to the male rat the duration of response to hexobarbital, but not that to barbital was significantly prolonged. Plasma concentrations of hexobarbital at awakening were not different from controls, indicating that cadmium treatment had not altered central nervous system sensitivity to the barbiturate. Such treatment of male rats 3 days prior to sacrifice leads to a significant inhibition of the metabolism of aminopyrine, hexobarbital, p-nitroanisole and zoxazolamine in microsomal subfractions obtained from these animals; microsomal cytochrome P-450 is reduced to 50% of control values by the cadmium treatment. Inhibition of the metabolism of these substrates by cadmium was also achieved when the metal was added to isolated microsomes in concentrations ranging from 5 × 10−4 to 5 × 10−7 m.

Effects of cadmium on nucleic acid and protein synthesis in rat liver

In male rats, cadmium (0.5 or 2.5 mg Cd 2+ /kg, ip) distributes among the major hepatic organelles, with the cytoplasmic fraction showing the greatest accumulation. Cadmium (0.34, 1.70, or 3.41 mg Cd 2+ /kg, ip) administration showed a biphasic effect with time on RNA synthesis in vivo . Upon its addition to isolated hepatic nuclei cadmium inhibited RNA synthesis. Administration of cadmium (0.34 or 1.70 mg Cd 2+ /kg, ip) to partially hepatectomized rats, decreased DNA synthesis. It was suggested from (DNA) melting profile measurements that cadmium may interact directly with DNA. The incorporation of l -[ 14 C]phenylalanine by a microsomal preparation from rats pretreated with cadmium (0.34, 1.70, or 3.41 mg Cd 2+ /kg, ip) was increased relative to controls. When hepatic microsomes were preincubated to destroy mRNA, the polyuridylic acid-directed incorporation of l -[ 14 C]phenylalanine was significantly inhibited by the cadmium pretreatment. This activity was likewise inhibited when cadmium was added to preincubated microsomes from control animals. The mRNA directed incorporation of amino acid by hepatic microsomes from control animals showed a significant biphasic effect dependent upon the concentration of added cadmium. At the doses of cadmium used in this study histologic examination of liver indicated focal necrosis, hemorrhage, inflammation, and congestion in some treatment groups.

Metabolism and tolerance studies with chlordiazepoxide-2-14C in the rat

Abstract Male rats made tolerant to chlordiazepoxide exhibit increased rates of tissue disappearance and excretion of 14 C-labeled chlordiazepoxide. A cross-tolerance is evident to hexobarbital, pentobarbital, and meprobamate, but not to phenobarbital. Stimulation by chlordiazepoxide of ascorbic acid excretion, and the ability of dl -ethionine to block the drug-induced stimulation of liver weight, liver protein content, and hepatic N -demethylase activity, suggest that the mechanism of tolerance development involves stimulation of hepatic microsomal enzymes responsible for drug metabolism.

Kinetics of cadmium-induced hepatic and renal metallothionein synthesis in the mouse.

Abstract Rates of hepatic and renal metallothionein synthesis were estimated by measuring the incorporation of [ 3 H]cysteine into metallothionein prepared from mice at various times following a single intraperitioneal injection of cadmium acetate (2 mg of Cd/kg). Tissue metallothionein concentrations were measured indirectly as a function of the total cadmium-binding capacity of the isolated metallothionein. Maximal incorporation of [ 3 H]cysteine into hepatic metallothionein occurred 6–12 hr following cadmium exposure, while renal metallothionein synthesis was maximal after 3 hr. Incorporation of [ 3 H]cysteine into metallothionein as well as metallothionein concentrations was greater in the liver than in the kidney. It is concluded that the liver is the primary site of cadmium-induced metallothionein synthesis.

Cadmium: inability to induce hypertension in the rat.

Abstract Cadmium has been suggested as a potential etiologic agent in human hypertension and has been reported to induce hypertension in experimenta animals. Cadmium acetate was administered ip to Sprague-Dawley derived and hooded Long-Evans rats at several dose levels and under a variety of treatment schedules and conditions. Under none of these circumstances was hypertension produced. These findings indicate that cadmium does not act as a proximate hypertension-inducing agent under all experimental conditions.

Age-related responsiveness of the rat to drugs affecting the central nervous system.

With few exceptions, the preclinical evaluation of drugs is undertaken in healthy, young adult animals. The available literature on the interaction between the age of animals and drug effects are largely concerned with the age span from neonate to young adult, i.e., developmental studies. Kato and Takanaka (1) have demonstrated depressed in vitro drug metabolism of several drugs including hexobarbital and me-probamate in 250-600 day old rats as compared to 100 day old rats. These investigators (2) also found an increased duration of action and a decreased rate of decline of plasma pentobarbital levels with increasing age. Ziem et al. (3) have reported increased central nervous system sensitivity to amphetamine in 12-14 mo old rats when compared with 3 mo old animals. The work presented here was performed to demonstrate age-related alterations in the responsiveness of mature rats to centrally acting drugs and to emphasize the importance of regarding the age of the animal as a significant factor in assessing drug response. Responses to sodium hexobarbital, chlorpromazine HCl, morphine SO4 and d-amphetamine SO4 were evaluated in 2.5-3 mo old and 9-10 mo old male rats. Methods. Male, albino Cox/Sprague-Dawley rats were obtained from Laboratory Supply Co., Indianapolis and housed in community cages for 1-2 weeks prior to use. Wayne Lab Bloks and tap water were supplied ad libitum. Room lights were cycled from 6 am to 8 pm daily and temperature was kept constant at 25 ± 1.0°. The electroencephalogram was recorded from three cortical electrodes (SS fillister head machine screws 0-80 × 1/8 in.) implanted 2 mm to either side of the saggital suture and 2 mm posterior to the bregma. A third screw was placed 1 cm anterior to the bregma and 1 mm to the right of the saggital suture.

Phenobarbital and methylcholanthrene stimulation of rat liver chromatin template activity

Abstract Both phenobarbital and 3-methylcholanthrene increase the activity of liver microsomal drug metabolizing enzymes, as discussed by Conney (1967) . This increased enzyme activity is due to de novo protein synthesis, as indicated by the studies of Von der Decken and Hultin (1960) , Gelboin and Sokoloff (1961) , and Conney and Gilman (1963) . Madix and Bresnick (1967) have shown that chromatin isolated at 7 and 16 hours after administration of the carcinogen 3-methylcholanthrene is a more effective template for in vitro RNA synthesis than chromatin from control animals. The results presented in the present report indicate that chromatin isolated from rat liver 12 hours after administration of phenobarbital or 3-methylcholanthrene is a more effective template for RNA synthesis when compared to controls, and that differences in template activity are abolished when basic proteins are removed from the chromatin preparations.

Stress stimulation of drug metabolism in the rat

Abstract Stressed rats have been shown to exhibit significantly decreased levels of hexobarbital, pentbarbital and meprobamate, but phenobarbital, in blood after administration of these drugs. The ability of stress to decrease drug levels in blood was shown to be dependent upon an intact pituitary-adrenal axis, since hypophysectomy and adrenalectomy blocked such responses. The response could be simulated in adrenalectomized animals by corticosterone, but not by ACTH, and in hypophysectomized animals by both corticosterone and ACTH. Actinomycin-D pretreatment blocked the ability of stress to decrease pentobarbital blood levels. Liver perfusion experiments show that pentobarbital metabolism is stimulated in stressed rats and that this stimulation can likewise be blocked by actinomycin-D.

Cadmium and vascular reactivity in the rat

Abstract Cadmium has been reported to induce hypertension accompanied by decreased vascular responsiveness to autonomic drugs in the rat and rabbit. Cadmium acetate administration to female Sprague-Dawley rats failed to elevate blood pressure, although cadmium-treated animals exhibited decreased blood pressure,responses to intravenously administered norepinephrine, acetylcholine, isoproterenol, and atropine. Aortic strips from cadmium acetate-treated animals demonstrated diminished reactivity to angiotensin, epinephrine, barium, and tyramine. It is concluded that cadmium treatment of the rat produces desensitization of the vasculature to vasoconstrictor and vasodilator substances independently of its ability to produce hypertension and, thus, that these facets of its biological activity are unrelated.