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.     

Protection of the Cyp1a2(-/-) null mouse against uroporphyria and hepatic injury following exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin

The effect of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) on the liver of C57BL/6J mice is a model for clinical sporadic porphyria cutanea tarda (PCT). There is massive uroporphyria, inhibition of uroporphyrinogen decarboxylase (UROD) activity, and hepatocellular damage. A variety of evidence implicates the CYP1A2 enzyme as necessary for mouse uroporphyria. Here we report that, 5 weeks after a single oral dose of TCDD (75 microg/kg), Cyp1a2(+/+) wild-type mice showed severe uroporphyria and greater than 90% decreases in UROD activity; in contrast, despite exposure to this potent agent Cyp1a2(-/-) knockout mice displayed absolutely no increases in hepatic porphyrin levels, even after prior iron overload, and no detectable inhibition of UROD activity. Plasma levels of alanine-aminotransferase (ALT) and aspartate aminotransferase (AST)-although elevated in both genotypes after TCDD exposure-were significantly less in Cyp1a2(-/-) than in Cyp1a2(+/+) mice, suggesting that the absence of CYP1A2 also affords partial protection against TCDD-induced liver toxicity. Histological examination confirmed a decrease in hepatocellular damage in TCDD-treated Cyp1a2(-/-) mice; in particular, there was no bile duct damage or proliferation that in the Cyp1a2(+/+) mice might be caused by uroporphyrin. We conclude that CYP1A2 is both necessary and essential for the potent uroporphyrinogenic effects of TCDD in mice, and that CYP1A2 also plays a role in contributing to TCDD-induced hepatocellular injury. This study has implications for both the toxicity assessment of TCDD and the hepatic injury seen in PCT patients.     

Hepatic toxicity and uroporphyrinogen decarboxylase activity following a single dose of 2, 3, 7, 8-tetrachlorodibenzo-p-dioxin to mice

A single, low-lethality, oral dose of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) (75 μg/kg) induces an hepatic porphyria in C57BL/10 mice of either sex. The hepatic porphyrin levels are maximal 4–6 weeks after dosing and are still elevated 12 weeks after the dose. The activity of hepatic uroporphyrinogen decarboxylase is depressed within one week of the dose and this appears to precede the onset of porphyria. Mice of the DBA/2 strain show no changes of the same magnitude at doses of 1200 μg/kg, for porphyrin levels, or 75 μg/kg, for decarboxylase activity. In both strains there is an increase in hepatic iron content 3 weeks after the 75 μg/kg dose. Female C57BL/10 mice are more resistant than males to the lethal effects of TCDD.     

Genetic variation of basal iron status, ferritin and iron regulatory protein in mice: potential for modulation of oxidative stress

Toxic and carcinogenic free radical processes induced by drugs and other chemicals are probably modulated by the participation of available iron. To see whether endogenous iron was genetically variable in normal mice, the common strains C57BL/10ScSn, C57BL/6J, BALB/c, DBA/2, and SWR were examined for major differences in their hepatic non-heme iron contents. Levels in SWR mice were 3- to 5-fold higher than in the two C57BL strains, with intermediate levels in DBA/2 and BALB/c mice. Concentrations in kidney, lung, and especially spleen of SWR mice were also greater than those in C57BL mice. Non-denaturing PAGE of hepatic ferritin from all strains showed a major holoferritin band at approximately 600 kDa, with SWR mice having > 3-fold higher levels than C57BL strains. SDS PAGE showed a band of 22 kDa, mainly representing L-ferritin subunits. A trace of a subunit at 18 kDa was also detected in ferritin from SWR mice. The 18 kDa subunit and a 500 kDa holoferritin from which it originates were observed in all strains after parenteral iron overload, and there was no major variation in ferritin patterns. Although iron uptake studies showed no evidence for differential duodenal absorption between strains to explain the variation in basal iron levels, acquisition of absorbed iron by the liver was significantly higher in SWR mice than C57BL/6J. As with iron and ferritin contents, total iron regulatory protein (IRP-1) binding capacity for mRNA iron responsive element (IRE) and actual IRE/IRP binding in the liver were significantly greater in SWR than C57BL/6J mice. Cytosolic aconitase activity, representing unbound IRP-1, tended to be lower in the former strain. SWR mice were more susceptible than C57BL/10ScSn mice to the toxic action of diquat, which is thought to involve iron catalysis. If extrapolated to humans, the findings could suggest that some people might have the propensity for greater basal hepatic iron stores than others, which might make them more susceptible to iron-catalysed toxicity caused by oxidants.     

Low levels of p53 are associated with resistance to tetrachlorodibenzo-p-dioxin toxicity in DBA/2 mice.

We show here that DBA/2 strain mice have a complex mutation/polymorphism in the promoter region of the Trp53 locus (the mouse p53 locus). This region has previously been shown to be essential for p53 expression. We further show that the DBA/2 mutation is associated with approximately fourfold lower p53 levels in thymocytes treated with the DNA-damaging agent etoposide in-vitro, and with relative resistance of these thymocytes to apoptosis induced by the DNA-damaging agent etoposide compared with C57BL/6 mice. When part of the promoter containing this mutation was inserted into a plasmid containing a luciferase reporter gene but lacking eukaryote promoter sequences and transfected into MCF-7 human breast cell line cells, the mean luciferase activity was slightly less with the DBA/2 than with the C57BL/6 promoter-reporter construct (p < 0.01). We found that DBA/2xC57BL/6 F2 hybrid mice with the DBA/2 genotype at the Trp53 locus were relatively resistant to tetrachlorodibenzo-p-dioxin toxicity, and this resistance was additive with resistance associated with the Ahr locus. DBA/2 mice are long-lived and do not have particularly high levels of cancer, suggesting either that they carry other compensatory tumour resistance alleles (such as Ahr(d)), or that, while there may be a p53 protein dosage effect for acute toxicity, lower than normal levels of p53 may still be sufficient to protect against cancer. In evolutionary terms, it may be better to maintain low levels of p53 in order to avoid death from acute toxicity, even at the expense of a higher incidence of cancer in later life.     

Uroporphyria produced in mice by iron and 5-aminolevulinic acid

Porphyria cutanea tarda and the analogous hepatic uroporphyria produced in rodents by aromatic hydrocarbons result from inactivation of hepatic uroporphyrinogen decarboxylase (UROD). Inactivation appears to be iron-dependent and may require induction of cytochromes of the P450IA subfamily. To investigate the hypothesis that the mechanism of inactivation involves an intermediate of haem biosynthesis, we administered iron and the haem precursor, 5-aminolevulinate (ALA), to mice. Iron-overloaded male mice of the Ah-responsive C57BL/6 strain, given ALA solution as their only drink, developed severe uroporphyria after 49 days. ALA did not produce uroporphyria in iron-overloaded male mice of the Ah-nonresponsive DBA/2 strain. Iron or ALA alone did not produce porphyria in either strain. Hepatic iron concentrations and rates of ethoxyresorufin deethylation (an indicator of cytochrome P450IA-mediated activity) were similar in both strains. These experiments show that a haem precursor is involved in iron-dependent inactivation of UROD. They emphasize the importance of inherited factors in determining susceptibility to this type of porphyria, even in the absence of administration of compounds that act through the Ah locus to induce cytochromes of the P450IA subfamily.     

2,3,7,8-Tetrachlorodibenzo-p-dioxin: its effect on genes for mandible traits in mice

2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) is a powerful toxicant that exerts its effects through the aryl hydrocarbon receptor (AHR) governed by the Ahr locus that in mice is located on chromosome 12. We used single marker analyses of the offspring of female mice treated/not treated with TCDD to search for a gene (quantitative trait locus or QTL) on chromosome 12 near the site of the Ahr locus to test whether this locus appeared to affect mandible size, shape, and/or asymmetry especially in the treated mice. These mice were sampled from the F(2) generation of an original intercross of two strains (C57BL/6J and AKR/J) known to be divergent in their response to TCDD. A QTL affecting mandible shape was found on chromosome 12, but its effect on mice in the treated and control groups did not differ and it was concluded that this QTL probably was not the Ahr locus itself. We also probed a second chromosome (11) and found a QTL whose effects on asymmetry of mandible shape differed in the two environments. These results suggested that the entire genome in these mice should be scanned to search for additional QTLs that might be affected by TCDD to learn more about the potential effects of this powerful toxicant on these genes.     

Quantitative trait loci for acute behavioral sensitivity to paraoxon

Genetic mechanisms responsible for organophosphate (OP)-induced behavioral changes remain obscure. In the present study, provisional quantitative trait loci (QTL) associated with acute sensitivity or insensitivity to hypolocomotion produced by the OP paraoxon were identified. Naive adult male and female mice of the BXD/Ty series (22 different BXD strains plus C57BL/6J and DBA/2J progenitor strains) received 0 or 0.25 mg/kg paraoxon (IP), immediately before placement in an activity chamber for a 30-min test. As expected, based on dose-response and time course studies with Swiss-Webster, C57BL/6, and DBA/2 mice, paraoxon treatment reduced locomotor activity in most, but not all BXD strains. Heritability (proportion of phenotypic variability attributed to genetic differences) was 0. 58 for the paraoxon treatment effect. Difference scores (strain mean for vehicle activity minus strain mean for paraoxon activity), and percent change in activity of paraoxon-treated mice compared to vehicle-treated mice were calculated for each BXD strain. QTL analyses using activity difference scores and percentage change in activity were conducted using a database with over 1300 unique genetic markers. Several provisional QTL found on different chromosomes were associated with the activity phenotype. Of these, several markers attained p<0.01 or greater. These were as follows: Chr 1: Ly9, p<0.006; Chr 6: D6Ncvs44, p<0.0005; Chr 9: D9Mit15, p<0. 003; Chr 11: D11Ncvs76, p<0.002; Chr 15: Tstap198, p<0.008. In addition, several markers on chromosome 3 approached p<0.01. Identified genes found near these regions include two plasma carboxylesterase alleles on chromosomes 6 and 9, a glutamate receptor subtype on chromosome 11 and a glycine receptor subunit on chromosome 11, raising the possibility that these genes could be the basis for these provisional QTLs.     

A verification of previously identified QTLs for cocaine-induced activation using a panel of B6.A chromosome substitution strains (CSS) and A/J x C57Bl/6J F2 mice

Background

The objective of this study was to confirm provisional quantitative trait loci (QTL) for cocaine-induced locomotor activation, on chromosomes 1, 5, 6, 9, 12, 15, 16, 17, and 18, previously identified in the AXB/BXA recombinant inbred (RI) and AcB/BcA recombinant congenic (RC) strains of mice derived from A/J (A) and C57BL/6J (B6) progenitors. This was accomplished through a genetic analysis of cocaine-induced activity in an AxB6 F2 cross and a phenotypic survey across a panel of B6.A chromosome substitution strains (CSS) mice. Mice were tested for cocaine-induced activity, following administration of saline and cocaine (20 mg/kg), utilizing an open-field procedure.

Results

Among AxB6 F2 mice, differences in cocaine-induced activity were associated with loci on chromosome 1 (D1Mit305), 5 (D5Mit409), 16 (D16Mit131), and 18 (D18Mit189). A survey of the CSS panel confirmed cocaine QTLs on chromosomes 5 and 15, previously identified in RI or RC strains. Overall, the regions on chromosomes 5 and 18 represent verification of QTL previously identified in both the RC and RI strains. Additionally, the B6 allele for these QTL was consistently associated with greater relative cocaine activation.

Conclusions

Collectively, chromosome 5 and 18 QTL have now been replicated in multiple independent crosses derived from the A/J and C57BL/6J progenitors. The use of an in silico analysis highlighted potential candidate genes on chromosomes 5 and 18. The present results complement the targeted gene approach currently prevalent in the study of cocaine and provide a broader empirically based focus for subsequent candidate gene studies.     

Comparison of 2,3,7,8-tetrachlorodibenzo-p-dioxin-mediated hepatotoxicity in C57BL/6J and DBA/2J mice

The toxicity of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) was examined by clinical chemistry and liver histopathology in Ah-responsive C57BL/6J (C57) and Ah-nonresponsive DBA/2J (DBA) mice. Hepatotoxicity was assessed at 1, 3, and 7 d following a single ip injection of TCDD at doses that maximally induce hepatic aryl hydrocarbon hydroxylase (AHH) activity (3 micrograms/kg for C57 and 30 micrograms/kg for DBA mice) and at doses approaching the LD50 (150 micrograms/kg for C57 and 600 micrograms/kg for DBA mice). Histological examination of liver sections was found to be a more sensitive detection method for TCDD-induced hepatic changes than clinical chemistry analyses. Dramatic differences in the development and type of liver injury were observed between TCDD-treated C57 and DBA mice. C57 mice given 3 micrograms TCDD/kg developed mild to moderate hepatic lipid accumulation in the absence of both inflammation and necrosis. Severe fatty change and mild inflammation and necrosis occurred in C57 mice that received 150 micrograms TCDD/kg. In contrast, DBA mice exposed to 30 micrograms TCDD/kg developed hepatocellular necrosis and inflammation without any fatty change. Only slight hepatic lipid accumulation occurred with some necrosis and inflammation in DBA mice given 600 micrograms TCDD/kg. The Ah locus may play a role in determining the sensitivity of C57 mice to the steatotic effects of TCDD.     

Experimental hepatic uroporphyria induced by the diphenyl-ether herbicide fomesafen in male DBA/2 mice

Hepatic uroporphyria can be readily induced by a variety of treatments in mice of the C57BL strains, whereas DBA/2 mice are almost completely resistant. However, feeding of the protoporphyrinogen oxidase-inhibiting herbicide fomesafen (0.25% in the diet for 18 weeks) induced hepatic uroporphyria in male DBA/2N mice (liver porphyrin content up to 150 nmol/g, control animals 1 nmol/g), whereas fomesafen-treated male C57BL/6N mice displayed only a slight elevation of liver porphyrins (approximately 5 nmol/g). The profile of accumulated hepatic porphyrins in fomesafen-treated DBA/2N mice resembled the well-characterised uroporphyria induced by polyhalogenated aromatic hydrocarbons, while histological examination confirmed the presence of uroporphyria-specific cytoplasmic inclusions in the hepatocytes. Uroporphyrinogen decarboxylase activity decreased to about 30% of control values in fomesafen-treated DBA/2N mice; microsomal methoxyresorufin O-dealkylase activity was slightly reduced. The amount of CYP1A1 and CYP1A2 mRNA, as determined by real-time PCR, was not significantly changed; mRNA encoding the housekeeping 5-aminolevulinic acid synthase was elevated 10-fold. Total liver iron was slightly increased. A similar uroporphyria was induced by the herbicide formulation Blazer, containing a structurally related herbicide acifluorfen, when fed to DBA/2N mice at a dose corresponding to 0.25% of acifluorfen in the diet. Since DBA/2 mice are almost completely resistant to all well-characterised porphyrogenic chemicals, the results suggest the possible existence of a yet unknown mechanism of uroporphyria induction, to which the DBA/2 mouse strain is more sensitive than the C57BL strain.     

Time and dose responses of the reduction in retinoid concentrations in C57BL/Rij and DBA/2 mice induced by 3,4,3',4'-tetrachlorobiphenyl

A single ip dose of 15 mg 3,4,3',4'-tetrachlorobiphenyl (TCB)/kg induced a 30 to 40% reduction of retinol and retinyl palmitate concentrations in hepatic tissue of C57BL/Rij mice within 2 to 4 days. This level of reduction was maintained for about 14 days. The ED50 was 32 mg TCB/kg for hepatic retinol and 17 mg TCB/kg for hepatic retinyl palmitate. In DBA/2 mice, however, no reduction in hepatic retinoids was observed even at doses up to 729 mg TCB/kg. The duration of the reduction in hepatic retinoids did not correlate with the induced aryl hydrocarbon hydroxylase (AHH) activity in C57BL/Rij mice. These data suggest that AHH and related enzymes are not directly involved in the TCB-induced reduction in retinoids in these mouse strains. No significant differences in the accumulation of TCB in hepatic tissues of C57BL/Rij and DBA/2 mice were observed; however, the estimated elimination rate in C57BL/Rij was two times faster than that in DBA/2 mice. In serum of DBA/2 mice, a rapid 50% decline in the concentration of retinol was observed after only 2 to 6 hr, remaining at the reduced value for about 14 days. The ED50 for serum retinol was 1 to 2 mg TCB/kg. Only a transient reduction in serum retinol, disappearing within 48 hr, was observed in the C57BL/Rij mouse. TCB accumulated to an almost 15-fold higher extent in serum of DBA/2 mice as compared with C57BL/Rij mice. The effect of TCB on retinoids in a few other strains of mice and in the Sprague-Dawley rat showed a reduction in serum retinol rather than in hepatic retinoids.     

The effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin on molar and mandible traits in congenic mice: a test of the role of the Ahr locus

2,3,7,8-Tetracholorodibenzo-p-dioxin is a highly toxic substance that can cause a variety of adverse effects on organisms. While it has been shown that TCDD acts mainly through the aryl-hydrocarbon receptor (AHR), the mechanism of toxicity is not completely clear. To test the role of the AHR in mediating the effects of TCDD, we exposed two congenic strains of mice differing only at the Ahr locus (Ahr(b)/Ahr(b) and Ahr(d)/Ahr(d)) to TCDD (0, 0.01, 0, or 1 microg/kg body weight) in utero on gestation day 13 and examined the developmental effects on mandible and mandibular tooth row size and shape. Our hypothesis was that TCDD would significantly affect one or more of these endpoints in Ahr(b)/Ahr(b) mice, previously shown to be sensitive to the effects of TCDD, while causing little or no effect in mice carrying the less sensitive Ahr(d) allele. At the doses used in this study, TCDD did not alter the size of mandibles or molars in either Ahr(b)/Ahr(b) or Ahr(d)/Ahr(d) mice. However, we did find that the highest dose of TCDD altered mandible shape, but only in Ahr(b)/Ahr(b) (not Ahr(d)/Ahr(d)) male mice. Similarly, the highest dose of TCDD significantly altered molar shape in Ahr(b)/Ahr(b) but not Ahr(d)/Ahr(d) male mice, although females in both congenic strains were affected. These results suggest that the effects of TCDD on molar and mandible shape are influenced by the Ahr genotype but that males and females differ in sensitivity to both of these effects.     

Fate of hexachlorobenzene in C57BL/10 mice with iron overload

The distribution of radioactivity in male C57BL/10 mice dosed with [14C]hexachlorobenzene (HCB) was followed over 21 days and found to be high in adipose tissue and adrenals, moderate in thymus whereas liver was relatively poorly labelled. A predose of iron (500 mg/kg), which greatly promotes the porphyrogenic action of HCB in this strain, had only a small effect on the distribution of radioactivity in tissues and excreta. Iron induced excretion of urinary metabolites from HCB by C57BL/10 mice but not by the insensitive DBA/2 strain. However, there was no such difference in faecal metabolites, total metabolism was only slightly increased and there was no correlation between liver porphyrin levels and urinary excretion of metabolites by individual mice. At the end of 4 weeks exposure of iron-treated C57BL/10 mice to HCB urinary metabolites fell while porphyrin excretion continued to rise. Thus the considerable sensitisation of the C57BL/10 strain after iron overload to the induction of porphyria by HCB cannot be ascribed simply to enhancement of total metabolism but must be caused either by the formation of a specific undetected metabolite or induction of some other toxic process.     

Dioxin-induced retardation of development through a reduction in the expression of pituitary hormones and possible involvement of an aryl hydrocarbon receptor in this defect: A comparative study using two strains of mice with different sensitivities to dioxin

We have previously revealed that treating pregnant rats with 2,3,7,8-tetracholorodibenzo-p-dioxin (TCDD) reduces the expression of gonadotropins and growth hormone (GH) in the fetal and neonatal pituitary. A change in gonadotropin expression impairs the testicular expression of steroidogenic proteins in perinatal pups, and imprint defects in sexual behavior after reaching maturity. In this study, we examined whether TCDD also affects the expression of gonadotropin and GH in mice using C57BL/6J and DBA/2J strains which express the aryl hydrocarbon receptor (Ahr) exhibiting a different affinity for TCDD. When pregnant C57BL/6J mice at gestational day (GD) 12 were given oral TCDD (0.2–20 μg/kg), all doses significantly attenuated the pituitary expression of gonadotropin mRNAs in fetuses at GD18. On the other hand, in DBA/2J mice, a much higher dose of TCDD (20 μg/kg) was needed to produce a significant attenuation. Such reduction in the C57BL/6J strain continued until at least postnatal day (PND) 4. In agreement with this, TCDD reduced the testicular expression of steroidogenic proteins in C57BL/6J neonates at PND2 and 4, although the same did not occur in the fetal testis and ovary. Furthermore, TCDD reduced the perinatal expression of GH, litter size and the body weight of newborn pups only in the C57BL/6J strain. These results suggest that 1) also in mice, maternal exposure to TCDD attenuates gonadotropin-regulated steroidogenesis and GH expression leading to the impairment of pup development and sexual immaturity; and 2) Ahr activation during the late fetal and early postnatal stages is required for these defects.     

Strain-dependent behavioural sensitization to amphetamine: role of environmental influences

Repeated daily pairings of 1mg/kg of amphetamine and test environment induced a large, significant increase of locomotion in mice of the C57BL/6 strain, while a slight, non-significant increase was observed in mice of the DBA/2 strain. Neither C57BL/6 nor DBA/2 mice showed behavioral sensitization to amphetamine in the test cages when the drug was repeatedly administered in their home cage. Moreover, C57BL/6 but not DBA/2 mice showed conditioned hyperactivity. Subsequently six daily pairings of saline and test cage produced a slight, non-significant reduction of the hyperactive response shown by C57BL/6 mice, accompanied by a further increase in the behavioural effect of amphetamine. Finally, a similar, significant context-independent sensitization (unpaired vs control) was observed in mice of the two strains subjected to pairings of saline with the test cage; while context-dependent sensitization (paired vs unpaired) was observed only in C57BL/6 mice. Naive DBA/2 were less susceptible than C57BL/6 mice to the behavioural effect of high doses of amphetamine. However, effects of the low dose of amphetamine used in this experiment did not show strain differences in naive mice. These results show that C57BL/6 are more susceptible than DBA/2 mice to context-dependent behavioural sensitization to amphetamine. Moreover, they suggest that neither conditioned hyperactivity nor context-independent sensitization account for strain differences in environment-specific behavioural sensitization.     

Localization of genes influencing ethanol-induced conditioned place preference and locomotor activity in BXD recombinant inbred mice

Genetic differences in ethanol's ability to induce conditioned place preference were studied in 20 BXD Recombinant Inbred (RI) mouse strains and in the C57BL/6J and DBA/2J progenitor strains. Male mice from each strain were exposed to a Pavlovian conditioning procedure in which a distinctive floor stimulus (CS+) was paired four times with ethanol (2 g/kg). A different floor stimulus (CS-) was paired with saline. Control mice were injected only with saline. Floor preference testing without ethanol revealed significant genetic differences in conditioned place preference, with some strains spending nearly 80% time on the ethanolpaired floor while others spent only 50% (i.e., no preference). Control mice showed genetic differences in unconditioned preference for the floor cues, but unconditioned preference was not genetically correlated with conditioned preference. There were also substantial genetic differences in ethanol-stimulated activity, but contrary to psychomotor stimulant theory, ethanol-induced activity on conditioning trials was not positively correlated with strength of conditioned place preference. However, there was a significant negative genetic correlation (r=–0.42) between test session activity and preference. Quantitative trait loci (QTL) analyses showed strong associations (P<0.01) between conditioned place preference and marker loci on chromosomes 4, 8, 9, 18 and 19. Weaker associations (0.01<P<0.05) were identified on several other chromosomes. Analysis also yielded several significant QTL for unconditioned preference, ethanol-stimulated activity, and sensitization. Overall, these data support the conclusion that genotype influences ethanol-induced conditioned place preference, presumably via genetic differences in sensitivity to ethanol's rewarding effects. Moreover, several chromosomal regions containing candidate genes of potential relevance to ethanol-induced conditioned place preference have been identified.     

2,3,7,8-Tetrachlorodibenzo-p-dioxin-induced porphyria in genetically inbred mice: partial antagonism and mechanistic studies

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) (233 nmol/kg) causes a significant increase of hepatic uroporphyrin, heptacarboxyporphyrin, and total porphyrins in female C57BL/6 mice, ovariectomized C57BL/6 mice, male C57BL/10 mice, and male C57BL/6 mice 3 weeks after treatment. In contrast, 6-methyl-1,3,8-trichlorodibenzofuran (MCDF) was inactive at a dose of 750 mumol/kg. Cotreatment of the mice with TCDD (233 mol/kg) plus MCDF (750 mumol/kg) resulted in partial antagonism of TCDD-induced hepatic porphyrin accumulation only in the female mice. Parallel studies in female C57BL/6 mice showed that the TCDD-induced porphyria was accompanied by the induction of hepatic microsomal aryl hydrocarbon hydroxylase (AHH) and ethoxyresorufin O-deethylase (EROD) activities and the depression of uroporphyrinogen decarboxylase (UROD). MCDF (750 mumol/kg) did not significantly affect these enzymes. In the cotreatment studies (MCDF plus TCDD), MCDF partially antagonized TCDD-induced hepatic porphyrin accumulation but did not affect the levels of hepatic AHH, EROD, or UROD. These results indicate that other factors, in addition to the induction of cytochrome P450-dependent monooxygenases and depressed UROD activity, are important in TCDD-induced porphyria in C57BL/6 female mice.     

The teratogenic sensitivity to 2,3,7,8-tetrachlorodibenzo-p-dioxin is modified by a locus on mouse chromosome 3

In an effort to understand how genetics can influence individual sensitivity to environmentally induced disease, we performed a linkage analysis to identify murine loci in addition to the Ahr locus that influence the incidence of cleft palate and hydronephrosis in developing mice exposed to the pollutant 2,3,7,8-tetrachlorodibenzo-p-dioxin (dioxin). Administration of 64 microg/kg dioxin to C57BL/6J (B6) dams at embryonic day 9 (E9) led to palatal clefting and hydronephrosis in nearly 100% of embryos by E17. In contrast, similar exposure of CBA/J (CBA) dams led to cleft palate in only 8% and hydronephrosis in 69% of embryos. To determine the genetic basis for this strain-dependent sensitivity, linkage analyses on the progeny of a B6CBAF1 intercross and a CBAxB6CBAF1 backcross were performed. The incidences of cleft palate and hydronephrosis were assessed and genomic DNA from embryos was analyzed at informative simple sequence length polymorphism (SSLP) markers. One locus segregating with dioxin-induced cleft palate was identified (p < 0.01) and designated as chemically mediated teratogenesis number 1 (Cmt1). The Cmt1 locus is located on chromosome 3.