Transplacental induction of mixed-function oxygenases in extra-hepatic tissues by 2,3,7,8-tetrachlorodibenzo-p-dioxin

Treatment of pregnant rats with 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) resulted in a dose-dependent induction of a mixed-function oxidase system in fetal and maternal extra-hepatic tissues. At doses of 6 μmg/kg, aryl hydrocarbon hydroxylase (AHH) activity was increased 24-, 22- and 4-fold in fetal lung, kidney and skin, respectively, while maternal lung, kidney and adrenal AHH activity was increased 4-, 2- and 2-fold respectively. High-pressure liquid chromatographic (H.P.L.C.) analysis of benzo(a)pyrene (BP) metabolism after TCDD induction indicated that fetal lung, kidney and skin produced significant quantities of benzo(a)pyrene-7,8-dihydrodiol (BP-7,8-diol), benzo(a)pyrene-4,5-dihydrodiol (BP-4,5-diol) and 9- and 3-phenols of BP. The fetal liver produced benzo(a)pyrene-9,10-dihydrodiol (BP-9,10-diol), BP-4,5-diol, BP-7,8-diol and 9- and 3-phenols of BP. Maternal lung also produced BP-9,10-diol, while maternal adrenal gland yielded primarily the 9-phenol of BP. Epoxide hydratase activity was increased 2- to 3-fold in maternal lung, fetal lung and skin after TCDD pretreatment, but was not affected significantly in liver, kidney or placenta. Treatment of pregnant rats with TCDD increased the covalent binding of BP to DNA in preparations containing maternal liver, lung and placenta as well as fetal liver, lung and skin. Pretreatment with TCDD resulted in increased epoxide hydratase and AHH activities in extra-hepatic tissues but only AHH was increased in hepatic tissues, indicating that the inducing capabilities of TCDD differ from, but share some similarities with, both phenobarbital (PB) and 3-methylcholanthrene (MC). Thus, TCDD appears to provide an exceptionally potent and broad-spectrum transplacental induction of carcinogen-transforming enzymes in extra-hepatic tissues.     

Toxic interaction of specific polychlorinated biphenyls and 2,3,7,8-tetrachlorodibenzo-p-dioxin: Increased incidence of cleft palate in mice

The induction of cleft palate in $\text{C57BL}\text{6N}$ mice is an extremely reproducible and sensitive indicator of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) toxicity. This endpoint was used to look for potential interactions between two polychlorinated biphenyl (PCB) congeners and TCDD. Both 2,3,4,5,3′,4′-hexachlorobiphenyl (HCB) and 2,4,5,2′,4′,5′-HCB are of relatively low toxic potency, but their biological properties differ. Pregnant mice were treated with TCDD and either HCB on gestation Days 10 through 13, and the fetuses examined for the presence of cleft palate and renal abnormalities on gestation Day 18. At a dose of TCDD which caused a low level of cleft palate, moderate hydronephrosis was observed. No renal or palatal anomalies were detected after 2,4,5,2′,4′,5′-HCB treatment, and the combination of this isomer with TCDD had no effect on the incidence of TCDD-induced cleft palate. 2,3,4,5,3′,4′-HCB caused mild renal toxicity, but no cleft palate. However, treatment of pregnant mice with a combination of TCDD and 2,3,4,5,3′,4′-HCB resulted in a 10-fold increase in the incidence of cleft palate. Thus, the toxicity of compounds such as TCDD may be enhanced by compounds of relatively low acute toxicity such as selected PCBs. The widespread environmental occurrence of such combinations suggests a need for further evaluation of the mechanism of this interaction.     

Induction of aryl hydrocarbon hydroxylase and epoxide hydrolase in rat liver,kidney, testis,prostate glands,and stomach by a potent nematocide, 1,2-dibromo-3-chloropropane

Because of environmental contamination with 1,2-dibromo-3-chloropropane (DBCP), a potent nematocide, it has become necessary to assess its adverse biological effects. The present data describe the ability of DBCP to induce aryl hydrocarbon hydroxylase (AHH) and epoxide hydrolase (EH) activities in the liver, kidney, testis, prostate glands, and stomach of the male rat. DBCP induced AHH in all organs studied except the liver and prostate glands. Maximum induction of AHH appeared 8, 8, and 16 hr following treatment for stomach, kidney, and testis, respectively. At the maximum induction, AHH activity in testis, kidney, and stomach was 2.1, 3.8, and 4.7 times that of controls, respectively. EH activity was also induced at varying times following DBCP treatment in the liver, kidney, testis, and prostate glands. The maximum induction occurred at 48 hr in kidney and testis, and at 74 hr in liver and prostate glands. At the maximally induced state, EH activity in prostate, testis, kidney, and liver was 2.3, 1.4, 1.9, and 2.6 times that of controls, respectively. Thus, EH induction was slightly less than that of AHH in respective organs. Furthermore, DBCP induction of AHH and EH activity was inhibited by either actinomycin D or cycloheximide, suggesting that this enzyme induction is due to de novo synthesis of new hemoproteins.     

Increased epidermal transglutaminase activity following 2,3,7,8-tetrachlorodibenzo-p-dioxin: In vivo and in vitro studies with mouse skin

In previous studies it has been shown that topical treatment of hairless mice with 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD, dioxin) induces hyperproliferation and hyperkeratinization in the epidermis of hairless mice. The present investigation demonstrated that such TCDD-induced morphological changes in skin in vivo are accompanied by increased levels in activity of epidermal transglutaminase (ETG), the enzyme associated with terminal epidermal differentiation. Exposure of mouse epidermal cells in tissue culture to 10^?9m TCDD also resulted in a significant increase in ETG activity, despite the fact that morphologically these cultures (grown at 0.07 mm ionic calcium concentrations) exhibited no signs of terminal differentiation. Thus one mechanism of action of TCDD in inducing cutaneous changes appears to relate to the stimulation of increased ETG levels.     

Aryl hydrocarbon hydroxylase induction in rat lung, liver, and male reproductive organs following inhalation exposure to diesel emission

Due to increasing use of diesel-powered vehicles, it has become necessary to assess adverse biological effects of diesel emissions in our environment. The present data describe the effects of inhalation exposure of diesel emission on the specific activities of aryl hydrocarbon hydroxylase (AHH) and epoxide hydrase (EH) in rat lung, liver, testis, and prostate gland. Adult CD strain, Sprague-Dawley male rats (8–10 weeks old) were exposed to diesel emission (1:13 dilution) containing 14.2 ppm ($\text{v}\text{v}$) total hydrocarbon, 20 hr/day for 42 days. Although liver exhibited the greatest overall AHH activity among the organs tested, the percentage increase over control values was highest in the prostate gland. On Day 15 of exposure, AHH activity had increased to 4.5-fold that of the control and remained elevated throughout the entire exposure period. In contrast, AHH induction in liver and lung occurred on Day 33 of exposure, and the maximum AHH induction was observed on Day 42 (1.4- and 1.3-fold increase in lung and liver, respectively). In all the organs tested, EH activity was not changed significantly during the experimental period.     

Incomplete correlation of 2,3,7,8-tetrachlorodibenzo-p-dioxin hepatotoxicity with Ah phenotype in mice

Pretreatment of male mice of the inbred strains A2G, BALB/c, C57BL/10, and AKR with iron dextran synergized the action of a single dose of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD, 75 micrograms/kg) in causing hepatic porphyria and necrosis 35 days later. There was no effect in DBA/2 mice. Increased porphyrin levels were associated with decreased hepatic activity of uroporphyrinogen decarboxylase. Iron alone had no effect on porphyrin levels or decarboxylase activity. In male BALB/c mice given TCDD alone there was a delay in the onset of porphyria. Female BALB/c, AKR, and AKR X DBA/2 F1 mice were more resistant to the porphyrinogenic effect of TCDD than males. Development of porphyria did not correlate with Ah phenotype of the mice. The inheritance of sensitivity to TCDD in crosses of the AKR and DBA/2 strains, both Ah nonresponsive, was studied by a biometrical genetic analysis. The inheritances of increased porphyrin levels and of increased plasma activity of enzymes indicative of hepatic necrosis were both complex. Segregation of alleles at more than one locus was required to explain the data. A lack of correlation of porphyrins with plasma enzyme levels in the F2 generation suggested that the expression of these traits was determined independently. Genes other than Ah influence the development of TCDD-induced hepatotoxicity in mice.     

2,3,7,8-Tetrachlorodibenzofuran tissue distribution and excretion in guinea pigs

The tissue distribution and excretion of [¹⁴C]2,3,7,8-tetrachlorodibenzofuran (TCDF) in adult male guinea pigs was studied up to 9 days following an iv or po administration (6.0 μg/kg, 0.02 μmol/kg). Greater than 90% of the dose was absorbed after po administration. Tissue concentrations and excreta were examined for [¹⁴C]TCDF-derived material for periods ranging from 1 hr to 9 days after administration. Liver, muscle, skin, and adipose tissue were found to be the most important tissues for the distribution of this compound. The highest levels of radioactivity were initially located in the liver and muscle, and then redistributed to the skin and adipose tissue. At 1 day the radioactivity began to be redistributed back to the liver; this second redistribution appeared to be due to the mobilization of fat stores associated with the overt toxicity of TCDF. At all time points, the specific activity of TCDF-derived radioactivity (percentage dose/g tissue) was highest in liver, fat, and adrenals, respectively. The initial phase for the elimination of radioactivity from the liver and muscle had a half-life of 3.9 and 15.7 hr, respectively. Excretion in the urine and feces each accounted for only 6.6 and 6.5% of the dose, respectively, during the 9 days following administration. Greater than 90% of the extracted radioactivity in feces and tissues examined cochromatographed with parent TCDF. However, the radioactivity in the urine did not chromatograph with TCDF and appeared to be a metabolite(s) more polar than TCDF. Studies of the accumulation and distribution of TCDF-derived material after six or more weekly doses of 1 μg/kg TCDF each demonstrated that TCDF is accumulated in liver, adipose tissue, and skin, and that when a body burden of approximately 5.6 to 6.6 μg/kg was attained, the toxicity of this compound was irreversible and resulted in progressive weight loss and death.     

Acute myelotoxic responses in mice exposed to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)

Blood cells are derived from a multitiered system of progenitor stem cells that lose their capacity for proliferation and self-renewal as they continue along pathways of differentiation. Since these hematopoietic events can be readily monitored in vivo and in vitro in the mouse, we have utilized this system to examine altered cellular differentiation associated with 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) toxicity. Progenitor cells were suppressed following acute exposure of mice to TCDD at doses as low as 1.0 micrograms/kg body wt. In vitro studies demonstrated that myelotoxicity occurs by a direct inhibition of proliferating stem cells. Genetic studies indicated that the myelotoxic responses to TCDD, both in vivo and in vitro, segregate with the Ah locus. In addition, the in vitro myelotoxicity of various polyhalogenated aromatic hydrocarbon congeners correlated with their previously reported ability to induce hepatic microsomal enzyme activity and to bind to an intracellular receptor for TCDD. TCDD was also found to bind specifically to bone marrow cells from Ah-responsive, but not nonresponsive mice, indicating that bone marrow cells possess a specific receptor for TCDD. These data indicate that the myelotoxic response to TCDD is regulated by the Ah receptor present in the target tissue and demonstrates the utility of this system for examining the cellular and molecular events associated with the toxicity of polyhalogenated aromatic hydrocarbons, the prototype for which is TCDD.     

Acute toxic effects in mice of an extract from the marine algae Gonyaulax monilata

Unialgal cultures of Gonyaulax monilata were cultured and harvested. A modified Westphall procedure was used to prepare an extract which did not contain saxitoxin, the gonyautoxins and structurally related toxins. The extract was administered i.p. to young adult, male CD-1 mice and produced: sedation, abdominal constriction, fecal clumping in the perianal area, ataxia, tremors, cyanosis, loss of reflexes, convulsions and death (LD50 = 2.28 mg/kg). Gross and microscopic pathology in the treated mice included: acute active hyperemia of the viscera, multifocal areas of necrosis of the musculature of the intestinal wall and diaphragm and the presence of cytoplasmic vacuoles in the peripheral margins of the acinar portion of the pancreas. Clinical pathology of the mice which survived 24 hr included significant elevation in the levels of serum lactic dehydrogenase, glutamic pyruvic and glutamic oxaloacetic transaminases. Some of these mice also had significantly decreased white blood cell counts. The extract administered orally produced similar signs without the abdominal constriction and convulsions (median lethal oral dose = 6.73 mg/kg). Gross pathology findings included extensive and severe congestion of the abdominal visceral organs. Vehicle control mice were normal. In conclusion, G. monilata, previously reported as nontoxic in homeotherms, yields an extract which contains a water soluble glycosidic substance(s) which is lethal to mice.     

The effects of methapyrilene hydrochloride on hepatocarcinogenicity and pentobarbital-induced sleeping time in rats and mice

In a 6- to 8-month dose-response toxicity study, methapyrilene hydrochloride (HCl) was administered in the feed to $\text{Fischer-}\text{344}\text{N}$ rats and B6C3F1 mice at concentrations of 125, 250, 500, 1000, or 2000 ppm. After 26 weeks, rats fed the higher dose levels of methapyrilene HCl developed cholangiocellular carcinomas in addition to severe hepatotoxic lesions. After 31 weeks, mice exhibited only mild hepatotoxicity from ingesting methapyrilene HCl, even at the higher dose levels. The duration of sodium pentobarbital-induced sleeping time (PEN-induced ST) is determined by the rate of hepatic metabolism of PEN and can therefore be used as an in vivo measurement of the level of hepatic detoxifying enzymes. A subsequent study was undertaken with PEN-induced ST to investigate differences in the effect of methapyrilene HCl on hepatic detoxifying enzymes between the two species; in this study, prior treatment with methapyrilene HCl significantly increased PEN-induced ST of rats but not of mice, indicating that methapyrilene HCl had a suppressive effect on detoxifying enzymes in rats but not in mice. In other groups in this study, phenobarbital, a known inducer of hepatic detoxification enzymes, was administered simultaneously with methapyrilene HCl to both rats and mice. Methapyrilene HCl plus phenobarbital exerted a synergistic effect of enzyme induction in mice. In rats, the phenobarbital-induced hepatic enzymes increased the rate of hepatic detoxification of methapyrilene HCl, as measured by a reduction of PEN-induced ST as compared with that in control rats or rats receiving methapyrilene HCl alone. The difference in the effects of methapyrilene HCl on hepatic detoxifying mechanisms of rats and mice, as indicated by the metabolism of PEN, may explain the difference between the two species in their susceptibility to the hepatocarcinogenic effects of methapyrilene HCl.     

Species differences in response to trichloroethylene. II. Biotransformation in rats and mice

Detailed analysis of urine from two strains of rats and mice dosed po with trichloroethylene at four doses from 10 to 2000 mg/kg failed to detect any major species or strain differences in the metabolism of trichloroethylene. Although a greater proportion of the dose was metabolized in mice than in rats, the relative proportions of the major metabolites were very similar in both strains and were unaffected by the dose amount. Analysis of the same urine samples for minor metabolites failed to establish a major species difference. Small amounts of dichloroacetic acid (less than 1% of the dose) were present in both rat and mouse urine and were not considered significant. Monochloroacetic acid accounted for less than 0.1% of the dose. Daily dosing of trichloroethylene (1000 mg/kg po) for 180 days did not induce the overall metabolism of trichloroethylene but did double the urinary excretion of trichloroacetic acid. This finding was accompanied by an equivalent percentage decrease in the concentration of trichloroethanol. CO2 has been shown to be a major metabolite of trichloroacetic acid, suggesting that this is the source of trichloroethylene-derived CO2. Trichloroacetic acid was also excreted in bile in both rats and mice suggesting possible conjugation of this metabolite in the liver. Very little evidence was found for the formation of chemically reactive species from trichloroethylene in either rats or mice and none that could be the basis of a major species difference. The increased rate of metabolism in the mouse, the resulting high blood concentrations of trichloroacetic acid, and stimulation of hepatic peroxisome proliferation in this species appears to be the major species difference possibly related to tumor formation in the liver. The conjugation of trichloroacetic acid and its metabolism to CO2 may be related to peroxisome proliferation.     

Dissociation between ornithine decarboxylase activity and aryl hydrocarbon hydroxylase induction in cell cultures treated with benz[a]anthracene and inhibitors of ornithine decarboxylase

The induction of aryl hydrocarbon hydroxylase and ornithine decarboxylase by benz[a]-anthracene in the presence or absence of ornithine decarboxylase inhibitors was studied in three different cell culture systems. An almost complete abolishment of ornithine decarboxylase activity by 1,3-diamino-2-propanol or α-difluoromethyl ornithine before the addition of the inducer did not affect appreciably the induction of aryl hydrocarbon hydroxylase by benz[a]anthracene in human embryo, HeLa and Reuber H-II-4-E cells in culture. These results suggest that the induction of aryl hydrocarbon hydroxylase does not require ornithine decarboxylase activity per se and can be expressed in the absence of continuous polyamine synthesis.     

Species differences in response to trichloroethylene. I. Pharmacokinetics in rats and mice

The elimination of radioactivity in two strains of rats and mice following a single po dose of trichloro[14C]ethylene at dose levels from 10 to 2000 mg/kg has shown a marked dose dependence in rats but not in mice. The metabolism of trichloroethylene in the mouse was linear over the range of doses used, whereas in the rat it became constant and independent of dose at 1000 mg/kg and above. At the 10-mg/kg dosage, both species metabolized trichloroethylene almost completely, 60% of the dose being excreted in urine with only 1 to 4% being eliminated unchanged in expired air in the first 24 hr. At 2000 mg/kg, 78% of the dose was eliminated unchanged in the rat, but only 14% in the mouse. Consequently at high dosages, the mouse was exposed to significantly higher concentrations of trichloroethylene metabolites than the rat. Blood level kinetics of trichloroethylene and its metabolites confirmed a faster rate of metabolism in the mouse than in the rat. Peak concentrations of the metabolites were reached within 2 hr of dosing in the mouse compared to 10 to 12 hr in the rat. The concentrations of both trichloroethanol (4X) and trichloroacetic acid (7X) were significantly higher in the mouse than in the rat. Whereas trichloroethanol was rapidly eliminated from blood, the higher concentrations of trichloroacetic acid were maintained for over 30 hr. The high blood quantities of trichloroethylene-derived trichloroacetic acid are known to induce hepatic peroxisome proliferation in mice but are insufficient to induce this response in rats. These data suggest that trichloroacetic acid blood amounts, peroxisome proliferation, and the link between peroxisomes and liver cancer are the basis of species difference in response to trichloroethylene.     

The effects of methyl mercury binding to microtubules

The effects of methyl mercury hydroxide (MeHg) on the in vitro polymerization and depolymerization of microtubules were studied. Polymerization was totally inhibited at 3.0 X 10(-5) M MeHg and depolymerization occurred at concentrations above 1.0 X 10(-5) M MeHg, reaching a maximal rate of -0.33%/min at 5.0 X 10(-5) M MeHg. At or above 1.0 X 10(-4) M MeHg, a mercury-protein aggregate formed in both the polymerization and depolymerization systems. Fifteen free sulfhydryl groups per tubulin dimer were determined, and MeHg bound to all 15. When MeHg bound to only 2 free sulfhydryl groups per dimer, it inhibited polymerization. MeHg bound to free sulfhydryl groups exposed uniquely on the surface of microtubules, as well as those free sulfhydryl groups exposed on the ends. These results show MeHg in vitro to be a potent microtubule assembly inhibitor at ratios stoichiometric with the tubulin dimer. The effects of MeHg on microtubules are presumably mediated through MeHg binding to free sulfhydryl groups both on the ends and on the surface of microtubules. The presence of binding sites (free sulfhydryl groups) on the microtubule surface suggests multiple classes of binding sites for MeHg.     

Resistance to cadmium-induced necrosis in testes of inbred mice: possible role of a metallothionein-like cadmium-binding protein

Altered accumulation and subcellular disposition of testicular Cd were examined as possible explanations for the resistance to Cd-induced testicular damage observed in certain inbred strains of mice. Mouse strains susceptible (129/J) or resistant (A/J) to Cd-induced testicular necrosis were injected iv with 10 or 45 mumol CdCl2/kg, respectively. This dosing regimen compensated for decreased Cd accumulation by A/J testes and established similar concentrations of Cd in testes of both strains (approximately 4 nmol Cd/g tissue). Twenty-four hours later, 129/J testes showed marked interstitial hemorrhage and seminiferous tubule necrosis, while A/J testes showed no microscopic evidence of damage. Two hours postinjection, no histopathologic changes were detected in testes of either strain; however, A/J testes had 15% more Cd associated with the cystosol than 129/J testes, and three times more Cd bound to a 14,500 MW cytosolic protein which had gel filtration and ion-exchange chromatography properties in common with metallothionein (MT). Therefore, resistance of A/J testes to Cd does not appear to be determined solely by decreased Cd accumulation, but is associated with increased binding of testicular Cd to a MT-like protein. However, this increase is not accompanied by a proportional increase in the total Cd-binding capacity of the MT-like protein in A/J testes compared to 129/J testes.     

Comparative toxicity of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in four (sub)strains of adult male rats

Four (sub)strains of adult male rats were given single oral doses of various concentrations of TCDD to establish and compare the oral 30-day LD50 values. The strains of rats were Fischer (F/334N) supplied by Harlan Industries, Frederick Cancer Research Center, and Charles River Breeding Laboratories; and CD supplied by Charles River Breeding Laboratories. The Charles River/Fischer rats were most sensitive to TCDD (LD50 = 164, 95% confidence limits 104-217 micrograms/kg), the Frederick/Fischer and Charles River/CD rats were moderately sensitive to TCDD (LD50 = 303, 250-360; and 297, 240-360 micrograms/kg, respectively), and the Harlan/Fischer rats were most resistant to TCDD (LD50 = 340, 281-409 micrograms/kg). The mean times of death were from 24.5 +/- 1.0 to 28.3 +/- 0.5 days and the percentage body weight loss at death was 37.4 +/- 1.2 to 42.7 +/- 1.3%. One week after exposure of the Charles River/Fischer animal to 45 micrograms TCDD/kg (1/4 the established 30-day LD50 dose), the same serum profile was induced as previously observed in the Harlan/Fischer rat, which includes hypoglycemia, hypertriglyceridemia, and hypercholesterolemia. These results emphasize the importance of indicating the precise dose, strain of rat, and time after dosing before termination in reporting the effects of TCDD on a particular biological response.     

The effects of the administration of fenoprofen or indomethacin to rat dams during late pregnancy,with special reference to the ductus arteriousus of the fetuses and neonates

Examination of a series of normal neonatal rats showed that the ductus arteriosus did not begin closing until approximately 16 to 20 min after birth and was usually completely closed by 85 to 98 min after birth. Pregnant rats were orally dosed twice daily for 1.5 to 5 days with 50 mg/kg of fenoprofen calcium, 1.6 mg/kg of indomethacin, or vehicle (1% methylcellulose), beginning 18 days after mating. The fenoprofen calcium treatment resulted in premature (in utero) closure of the ductus arteriosus in 4 to 10% of the offspring of dams treated for 3.5 to 5 days. The indomethacin treatment caused premature (in utero) ductal closure in 1 to 3% of the offspring after 4.5 to 5 days. Both treatments also caused a high incidence of maternal mortality after 4.5 to 5 days (38 to 50% for fenoprofen calcium and 13 to 25% for indomethacin) and increased the duration of gestation or blocked parturition. Treatment of rat dams with 4 mg of progesterone daily from 18 through 22 days after mating blocked parturition, but the ductus arteriosus was not closed in any of the fetuses that were retained in utero. This suggests that the in utero closure of the ductus found in fetuses of dams treated with fenoprofen calcium or indomethacin was not a consequence of retention of the fetuses in the uterus.     

Consideration of the mechanism of pulmonary adenogenesis in urethane-treated Swiss mice

A number of investigators have observed a quadratic relationship between acute urethane dose and cumulative pulmonary adenoma incidence in mice. The hypothesis was tested that this dose-effect relationship may be explained by consideration of the elimination kinetics of urethane. Single doses of 0.4 to 1.8 mg urethane per g body weight were given ip to 6-week-old Swiss-Cox mice. The measure of internal exposure to the intact urethane molecule for a given external urethane dose was taken to be the area under the curve (AUC alpha) of a blood urethane concentration versus time plot. AUC alpha was linearly related to tumor prevalence. When urethane elimination was induced by pretreatment of the mice with p,p'-DDT, the linear relationship of AUC alpha to tumor prevalence was shifted. Reduction of tumor prevalence by p,p'-DDT pretreatment was more marked than that predicted on the basis of exposure to urethane. Thus, the kinetic evidence is consistent with biochemical evidence from other investigators supporting the premise that activation of urethane to a reactive metabolite is required for adenoma formation in the mouse lung. While exposure to the adenogenic moiety is evidently closely proportional to internal exposure to urethane in both pretreated and non-pretreated mice, p,p'-DDT pretreatment causes a shift in this proportionality.