Developmental toxicity of perfluorooctane sulfonate (PFOS) is not dependent on expression of peroxisome proliferator activated receptor-alpha (PPAR alpha) in the mouse

Perfluorooctane sulfonate (PFOS) and perfluorooctanoic acid (PFOA) are members of a family of perfluorinated compounds. Both are environmentally persistent and found in the serum of wildlife and humans. PFOS and PFOA are developmentally toxic in laboratory rodents. Exposure to these chemicals in utero delays development and reduces postnatal survival and growth. Exposure to PFOS on the last 4 days of gestation in the rat is sufficient to reduce neonatal survival. PFOS and PFOA are weak agonists of peroxisome proliferator activated receptor-alpha (PPAR alpha). The reduced postnatal survival of neonatal mice exposed to PFOA was recently shown to depend on expression of PPAR alpha. This study used PPAR alpha knockout (KO) and 129S1/SvlmJ wild type (WT) mice to determine if PPAR alpha expression is required for the developmental toxicity of PFOS. After mating overnight, the next day was designated gestation day (GD) 0. WT females were weighed and dosed orally from GD15 to 18 with 0.5% Tween-20, 4.5, 6.5, 8.5, or 10.5mg PFOS/kg/day. KO females were dosed with 0.5% Tween-20, 8.5 or 10.5mg PFOS/kg/day. Dams and pups were observed daily and pups were weighed on postnatal day (PND) 1 and PND15. Eye opening was recorded from PND12 to 15. Dams and pups were killed on PND15, body and liver weights recorded, and serum collected. PFOS did not affect maternal weight gain or body or liver weights of the dams on PND15. Neonatal survival (PND1-15) was significantly reduced by PFOS in both WT and KO litters at all doses. WT and KO pup birth weight and weight gain from PND1 to 15 were not significantly affected by PFOS exposure. Relative liver weight of WT and KO pups was significantly increased by the 10.5mg/kg dose. Eye opening of PFOS-exposed pups was slightly delayed in WT and KO on PND13 or 14, respectively. Because results in WT and KO were comparable, it is concluded that PFOS-induced neonatal lethality and delayed eye opening are not dependent on activation of PPAR alpha.     

Developmental toxicity of perfluorooctane sulfonate (PFOS) is not dependent on expression of peroxisome proliferator activated receptor-alpha (PPARα) in the mouse

Perfluorooctane sulfonate (PFOS) and perfluorooctanoic acid (PFOA) are members of a family of perfluorinated compounds. Both are environmentally persistent and found in the serum of wildlife and humans. PFOS and PFOA are developmentally toxic in laboratory rodents. Exposure to these chemicals in utero delays development and reduces postnatal survival and growth. Exposure to PFOS on the last 4 days of gestation in the rat is sufficient to reduce neonatal survival. PFOS and PFOA are weak agonists of peroxisome proliferator activated receptor-alpha (PPARα). The reduced postnatal survival of neonatal mice exposed to PFOA was recently shown to depend on expression of PPARα. This study used PPARα knockout (KO) and 129S1/SvlmJ wild type (WT) mice to determine if PPARα expression is required for the developmental toxicity of PFOS. After mating overnight, the next day was designated gestation day (GD) 0. WT females were weighed and dosed orally from GD15 to 18 with 0.5% Tween-20, 4.5, 6.5, 8.5, or 10.5 mg PFOS/kg/day. KO females were dosed with 0.5% Tween-20, 8.5 or 10.5 mg PFOS/kg/day. Dams and pups were observed daily and pups were weighed on postnatal day (PND) 1 and PND15. Eye opening was recorded from PND12 to 15. Dams and pups were killed on PND15, body and liver weights recorded, and serum collected. PFOS did not affect maternal weight gain or body or liver weights of the dams on PND15. Neonatal survival (PND1–15) was significantly reduced by PFOS in both WT and KO litters at all doses. WT and KO pup birth weight and weight gain from PND1 to 15 were not significantly affected by PFOS exposure. Relative liver weight of WT and KO pups was significantly increased by the 10.5 mg/kg dose. Eye opening of PFOS-exposed pups was slightly delayed in WT and KO on PND13 or 14, respectively. Because results in WT and KO were comparable, it is concluded that PFOS-induced neonatal lethality and delayed eye opening are not dependent on activation of PPARα.     

Developmental triclosan exposure decreases maternal, fetal, and early neonatal thyroxine: a dynamic and kinetic evaluation of a putative mode-of-action

This work tests the mode-of-action (MOA) hypothesis that maternal and developmental triclosan (TCS) exposure decreases circulating thyroxine (T4) concentrations via up-regulation of hepatic catabolism and elimination of T4. Time-pregnant Long-Evans rats received TCS po (0-300mg/kg/day) from gestational day (GD) 6 through postnatal day (PND) 21. Serum and liver were collected from dams (GD20, PND22) and offspring (GD20, PND4, PND14, PND21). Serum T4, triiodothyronine (T3), and thyroid-stimulating hormone (TSH) concentrations were measured by radioimmunoassay. Ethoxy-O-deethylase (EROD), pentoxyresorufin-O-depentylase (PROD) and uridine diphosphate glucuronyltransferase (UGT) enzyme activities were measured in liver microsomes. Custom Taqman(?) qPCR arrays were employed to measure hepatic mRNA expression of select cytochrome P450s, UGTs, sulfotransferases, transporters, and thyroid hormone-responsive genes. TCS was quantified by LC/MS/MS in serum and liver. Serum T4 decreased approximately 30% in GD20 dams and fetuses, PND4 pups and PND22 dams (300mg/kg/day). Hepatic PROD activity increased 2-3 fold in PND4 pups and PND22 dams, and UGT activity was 1.5 fold higher in PND22 dams only (300mg/kg/day). Minor up-regulation of Cyp2b and Cyp3a expression in dams was consistent with hypothesized activation of the constitutive androstane and/or pregnane X receptor. T4 reductions of 30% for dams and GD20 and PND4 offspring with concomitant increases in PROD (PND4 neonates and PND22 dams) and UGT activity (PND22 dams) suggest that up-regulated hepatic catabolism may contribute to TCS-induced hypothyroxinemia during development. Serum and liver TCS concentrations demonstrated greater fetal than postnatal internal exposure, consistent with the lack of T4 changes in PND14 and PND21 offspring. These data support the MOA hypothesis that TCS exposure leads to hypothyroxinemia via increased hepatic catabolism; however, the minor effects on thyroid hormone metabolism may reflect the low efficacy of TCS as thyroid hormone disruptor or highlight the possibility that other MOAs may also contribute to the observed maternal and early neonatal hypothyroxinemia.     

Exposure to perfluorooctane sulfonate during pregnancy in rat and mouse. I: maternal and prenatal evaluations.

The maternal and developmental toxicities of perfluorooctane sulfonate (PFOS, C8F17SO3-) were evaluated in the rat and mouse. PFOS is an environmentally persistent compound used as a surfactant and occurs as a degradation product of both perfluorooctane sulfonyl fluoride and substituted perfluorooctane sulfonamido components found in many commercial and consumer applications. Pregnant Sprague-Dawley rats were given 1, 2, 3, 5, or 10 mg/kg PFOS daily by gavage from gestational day (GD) 2 to GD 20; CD-1 mice were similarly treated with 1, 5, 10, 15, and 20 mg/kg PFOS from GD 1 to GD 17. Controls received 0.5% Tween-20 vehicle (1 ml/kg for rats and 10 ml/kg for mice). Maternal weight gain, food and water consumption, and serum chemistry were monitored. Rats were euthanized on GD 21 and mice on GD 18. PFOS levels in maternal serum and in maternal and fetal livers were determined. Maternal weight gains in both species were suppressed by PFOS in a dose-dependent manner, likely attributed to reduced food and water intake. Serum PFOS levels increased with dosage, and liver levels were approximately fourfold higher than serum. Serum thyroxine (T4) and triiodothyronine (T3) in the PFOS-treated rat dams were significantly reduced as early as one week after chemical exposure, although no feedback response of thyroid-stimulating hormone (TSH) was observed. A similar pattern of reduction in T4 was also seen in the pregnant mice. Maternal serum triglycerides were significantly reduced, particularly in the high-dose groups, although cholesterol levels were not affected. In the mouse dams, PFOS produced a marked enlargement of the liver at 10 mg/kg and higher dosages. In the rat fetuses, PFOS was detected in the liver but at levels nearly half of those in the maternal counterparts, regardless of administered doses. In both rodent species, PFOS did not alter the numbers of implantations or live fetuses at term, although small deficits in fetal weight were noted in the rat. A host of birth defects, including cleft palate, anasarca, ventricular septal defect, and enlargement of the right atrium, were seen in both rats and mice, primarily in the 10 and 20 mg/kg dosage groups, respectively. Our results demonstrate both maternal and developmental toxicity of PFOS in the rat and mouse.     

Developmental exposure to brominated diphenyl ethers results in thyroid hormone disruption.

The objective of the current study was to characterize the effects of DE-71 (a commercial polybrominated diphenyl ether mixture containing mostly tetra- and penta-bromodiphenyl ethers) on thyroid hormones and hepatic enzyme activity in offspring, following perinatal maternal exposure. Primiparous Long-Evans rats were orally administered DE-71 (0, 1, 10, and 30 mg/kg/day) in corn oil from gestation day (GD) 6 to postnatal day (PND) 21. Serum and liver samples obtained from dams (GD 20 and PND 22), fetuses (GD 20), and offspring (PNDs 4, 14, 36, and 90) were analyzed for circulating total serum thyroxine (T(4)) and triiodothyronine (T(3)), or hepatic microsomal ethoxy- and pentoxy-resorufin-O-deethylase (EROD and PROD), and uridine diphosphoglucuronosyl transferase (UDPGT) activity. There were no significant effects of treatment on maternal body weight gain, litter size, or sex ratio, nor were there any effects on any measures of offspring viability or growth. Serum T(4) was reduced in a dose-dependent manner in fetuses on GD 20 (at least 15%) and offspring on PND 4 and PND 14 (50 and 64% maximal in the 10 and 30 mg/kg/day groups, respectively), but recovered to control levels by PND 36. Reduction in serum T(4) was also noted in GD 20 dams (48% at highest dose), as well as PND 22 dams (44% at highest dose). There was no significant effect of DE 71 on T(3) concentrations at any time in the dams or the offspring. Increased liver to body weight ratios in offspring were consistent with induction of EROD (maximal 95-fold), PROD (maximal 26-fold) or UDPGT (maximal 4.7-fold). Induction of PROD was similar in both dams and offspring; however, EROD and UDPGT induction were much greater in offspring compared to dams (EROD = 3.8-fold; UDPGT = 0.5-fold). These data support the conclusion that DE-71 is an endocrine disrupter in rats during development.     

Lack of differential sensitivity to cholinesterase inhibition in fetuses and neonates compared to dams treated perinatally with chlorpyrifos.

Pregnant Sprague-Dawley rats were exposed to chlorpyrifos (CPF; O,O-diethyl-O-[3,5,6-trichloro-2-pyridinyl] phosphorothioate) by gavage (in corn oil) from gestation day (GD) 6 to postnatal day (PND) 10. Dosages to the dams were 0 (control), 0.3 (low), 1.0 (middle) or 5.0 mg/kg/day (high). On GD 20 (4 h post gavage), the blood CPF concentration in fetuses was about one half the level found in their dams (high-dose fetuses 46 ng/g; high-dose dams 109 ng/g). CPF-oxon was detected only once; high-dose fetuses had a blood level of about 1 ng/g. Although no blood CPF could be detected (limit of quantitation 0.7 ng/g) in dams given 0.3 mg/kg/ day, these dams had significant inhibition of plasma and red blood cell (RBC) ChE. In contrast, fetuses of dams given 1 mg/kg/day had a blood CPF level of about 1.1 ng/g, but had no inhibition of ChE of any tissue. Thus, based on blood CPF levels, fetuses had less cholinesterase (ChE) inhibition than dams. Inhibition of ChE occurred at all dosage levels in dams, but only at the high-dose level in pups. At the high dosage, ChE inhibition was greater in dams than in pups, and the relative degree of inhibition was RBC approximately plasma > or = heart > brain (least inhibited). Milk CPF concentrations were up to 200 times those in blood, and pup exposure via milk from dams given 5 mg/kg/day was estimated to be 0.12 mg/kg/day. Therefore, the dosage to nursing pups was much reduced compared to the dams exposure. In spite of exposure via milk, the ChE levels of all tissues of high-dosage pups rapidly returned to near control levels by PND 5.     

Neonatal death of mice treated with perfluorooctane sulfonate

Pregnant mice exposure to perfluorooctane sulfonate (PFOS) causes neonatal death. Ten pregnant ICR mice per group were given 1, 10 or 20 mg/kg PFOS daily by gavage from gestational day (GD) 0 to the end of the study. Five dams per group were sacrificed on GD 18 for prenatal evaluation, the others were left to give birth. Additional studies were conducted for histopathological examination of lungs and heads of fetuses and neonates at birth. PFOS treatment (20 mg/kg) reduced the maternal weight gain and feed intake but increased the water intake. The liver weight increased in a dose-dependent manner accompanied by hepatic hypertrophy at 20 mg/kg. PFOS reduced the fetal body weight in a dose-dependent manner and caused a bilateral enlargement in the neck region in all fetuses at 20 mg/kg and mild enlargement in some fetuses at 10 mg/kg, in addition to skeletal malformations. Almost all fetuses at 20 mg/kg were alive on GD18 and showed normal lung structure; but at parturition, all neonates were inactive and weak, showed severe lung atelectasis and severe dilatation of intracranial blood vessel, and died within a few hours. At 10 mg/kg, all neonates were born alive, 27% showed slight lung atelectasis, all of them had mild to severe dilatation of the intracranial blood vessel, and 45% of neonates died within 24 hr. The cause of neonatal death in mice exposed to PFOS may be attributed either to the intracranial blood vessel dilatation or to respiratory dysfunction. The former might be a cause of the latter.     

Endocrine-disrupting activities in vivo of the fungicides tebuconazole and epoxiconazole.

The triazole fungicides tebuconazole and epoxiconazole were investigated for reproductive toxic effects after exposure during gestation and lactation. Rats were dosed with epoxiconazole (15 or 50 mg/kg bw/day) or tebuconazole (50 or 100 mg/kg bw/day) during pregnancy from gestational day (GD) 7 and continued during lactation until postnatal day (PND) 16. Some dams were randomly chosen for cesarean section at GD 21 to evaluate effects on sexual differentiation in the fetuses. Other dams delivered normally, and the pups were examined (e.g., anogenital distance [AGD] and hormone levels) at birth, at PND 13 or PND 16, and semen quality was assessed in adults. Both tebuconazole and epoxiconazole affected reproductive development in the offspring after exposure in utero. Both compounds virilized the female offspring as shown by an increased AGD PND 0. Furthermore, tebuconazole had a feminizing effect on male offspring as shown by increased nipple retention. This effect was likely caused by the reduced testosterone levels seen in male fetuses. Tebuconazole increased the testicular concentrations of progesterone and 17alpha-hydroxyprogesterone in male fetuses, indicating a direct impact on the steroid synthesis pathway in the Leydig cells. The high dose of epoxiconazole had marked fetotoxic effects, while the lower dose caused increased birth weights. The increased birth weights may be explained by a marked increase in testosterone levels in dams during gestation. Common features for azole fungicides are that they increase gestational length, virilize female pups, and affect steroid hormone levels in fetuses and/or dams. These effects strongly indicate that one major underlying mechanism for the endocrine-disrupting effects of azole fungicides is disturbance of key enzymes like CYP17 involved in the synthesis of steroid hormones.     

Developmental toxicology of trimethyltin in the rat

Pregnant rats were treated on either gestational day (GD) 7, 12, or 17 with single doses of trimethyltin chloride (TMT) ip at either 0, 5, 7, or 9 mg/kg. A significant effect of dose was manifest as decreased maternal weight at term, which persisted during lactation until postnatal day (PND) 15 in some groups. For all treatments combined, term weights of dams exposed on GDs 7 and 12 were greater than those treated on GD 17. Litter sizes were decreased for groups treated on GD 17 with 9 mg/kg TMT. Pups treated in utero and exhibiting treatment-induced decreases in weight at or near birth remained smaller than untreated animals into adulthood (PND 280). By PND 20, weights of pups treated on GD 7 greater than GD 12 greater than GD 17. Neuropathology of pups sacrificed on PND 1 was minimal in all animals with lesions only identified in animals treated on GDs 12 or 17 which consisted of subtle degenerative changes in the CA3 and CA4 regions of Ammon's horn of the hippocampus. Muscarinic cholinergic receptor binding in whole brains from pups on PND 1 did not show any significant changes compared to controls for any dose or day of exposure. These data indicate that prenatal TMT exposure results in postnatal toxicity in treated pups but only in the presence of maternal toxicity.     

Prenatal PFOS exposure induces oxidative stress and apoptosis in the lung of rat off-spring

Perfluorooctane sulfonate (PFOS) could induce neonatal pulmonary injuries in rodents. The aim of this study was to investigate the underlying mode of action. Pregnant rats were dosed orally with PFOS (0, 0.1 and 2.0mg/kgd) from gestation days (GD) 1 to 21. Lung samples from postnatal day (PND) 0 and 21 pups were analyzed for the toxic effects of PFOS. The results showed that maternal exposure to 2.0mg/kgd PFOS caused severe histopathological changes along with marked oxidative injuries and cell apoptosis in offspring lungs; at the same time, the ratio of Bax to Bcl-2, release of cytochrome c (Cyt c) from mitochondria to cytoplasm, expressions of Fas and Fas-L, and activities of caspase-3, -8 and -9 were up-regulated correspondingly. The results indicate that oxidative stress and both intrinsic and extrinsic cell death pathways were involved in prenatal PFOS exposure-induced injuries in postnatal lungs.     

Differences in metabolite burden of di(2-ethylhexyl)phthalate in pregnant and postpartum dams and their offspring in relation to drug-metabolizing enzymes in mice

Di(2-ethylhexyl)phthalate (DEHP) induced adverse effects on mice offspring, and the metabolite mono(2-ethylhexyl)phthalate (MEHP) may be essential to determine the toxicity. In this experiment, we measured liver MEHP levels and the factors determining the metabolism, two enzyme activities [lipase and uridine 5′-diphosphate-glucuronosyltransferase (UGT)] or expression of cytochrome P450 4A14 (CYP4A14) in dams (on gestational day 18 and postnatal day 2) and their offspring. MEHP concentrations in the liver from pregnant dams were 1.5 times higher than those of postpartum dams at exposure to 0.05% DEHP. Accordingly, MEHP concentrations were 1.7 times higher in fetuses than in pups at the dose. Interestingly, lipase activity was 1.8-fold higher in pregnant dams than postpartum ones, but no such difference was noted in the activity between fetuses and pups. UGT activity was also 1.5-fold higher in pregnant dams than postpartum ones, whereas the activity in the fetuses was 1/2 that of pups. No difference was noted in CYP4A14 levels between pregnant and postpartum mice, whereas the levels in the fetuses were <1/10 those of pups. DEHP exposure did not influence lipase activity, whereas it slightly enhanced UGT activity and exclusively increased CYP4A14 levels in pregnant and/or postpartum dams. Taken together, the higher MEHP levels in pregnant dams than postpartum ones may be primarily due to higher lipase activities in pregnant dams, which may closely reflect those in fetuses and pups.     

Perfluorooctanoic acid and perfluorononanoic acid in fetal and neonatal mice following in utero exposure to 8-2 fluorotelomer alcohol.

8-2 Fluorotelomer alcohol (FTOH) and its metabolites, perfluorooctanoic acid (PFOA) and perfluorononanoic acid (PFNA), are developmental toxicants but metabolism and distribution during pregnancy are not known. To examine this, timed-pregnant mice received a single gavage dose (30 mg 8-2 FTOH/kg body weight) on gestational day (GD) 8. Maternal and neonatal serum and liver as well as fetal and neonatal homogenate extracts were analyzed using gas chromatography coupled with mass spectrometry. During gestation (GD9 to GD18), maternal serum and liver concentrations of PFOA decreased from 789 +/- 41 to 668 +/- 23 ng/ml and from 673 +/- 23 to 587 +/- 55 ng/g, respectively. PFOA was transferred to the developing fetuses as early as 24-h posttreatment with concentrations increasing from 45 +/- 9 ng/g (GD10) to 140 +/- 32 ng/g (GD18), while PFNA was quantifiable only at GD18 (31 +/- 4 ng/g). Post-partum, maternal serum PFOA concentrations decreased from 451 +/- 21 ng/ml postnatal day (PND) 1 to 52 +/- 19 ng/ml (PND15) and PFNA concentrations, although fivefold less, exhibited a similar trend. Immediately after birth, pups were cross-fostered with dams that had been treated during gestation with 8-2 FTOH (T) or vehicle (C) resulting in four treatment groups in which the first letter represents in utero (fetal) exposure and the second represents lactational (neonatal) exposure: C/C, T/C, C/T, T/T. On PND1, neonatal whole-body homogenate concentrations of PFOA from T/T and T/C groups averaged 200 +/- 26 ng/g, decreased to 149 +/- 19 ng/g at PND3 and this decreasing trend was seen in both neonatal liver and serum from PND3 to PND15. Based on detectible amounts of PFOA in neonatal serum in the C/T group on PND3 (57 +/- 11 ng/ml) and on PND15 (58 +/- 3 ng/ml), we suggest that the neonates were exposed through lactation. In conclusion, exposure of neonates to PFOA and PFNA occurs both pre- and postnatally following maternal 8-2 FTOH exposure on GD8.     

Organotin speciation and tissue distribution in rat dams, fetuses, and neonates following oral administration of tributyltin chloride

Tributyltin (TBT) is a biocide that contaminates human foodstuffs, especially shellfish. TBT is an endocrine disrupter, producing imposex in several marine gastropods. Previous studies showed that oral dosing of rat dams with TBT chloride leads to abnormal fetal and postnatal development. In this study, the tissue distribution and speciation of organotins in tissues were examined in dams, fetuses, and neonates following dosing of rat dams commencing on gestational day (GD) 8 by oral gavage with TBT in olive oil at 0, 0.25, 2.5, or 10 mg/kg body weight (BW)/d. Dams' body weights were significantly reduced by the 10-mg/kg BW/d TBT treatment. At GD20, there were no significant effects of any TBT treatment on pup weights, litter size, sex ratio, or tissue weights. However, at postnatal day (PND) 6 and 12, neonatal pup weights were reduced by the 10-mg/kg BW/d TBT treatment but tissue weights were unaffected, except for the liver weight of female pups, which was reduced by the 10-mg/kg BW/d TBT treatment. Tissues harvested on GD20 and PND6 and PND12 were extracted for determination of organotins by gas chromatography-atomic emission detection (GC-AED). In most tissues, TBT and its metabolite dibutyltin (DBT) were evident but monobutyltin (MBT) was rarely measured above the detection limit. The livers and brains of fetuses contained TBT and DBT at levels that were approximately 50% of the equivalent tissues in the dams. Furthermore, these tissues appeared to preferentially absorb/retain organotins, since the concentrations were greater than were found for the total loading in whole pups. The placenta also contained relatively large quantities of TBT and DBT. Postnatally, the TBT levels in pups decreased markedly, a probable consequence of the extremely low levels of organotins in rat milk. However, DBT levels in pups livers and brains were maintained, probably due to metabolism of TBT to DBT. Similarly, while dams' spleens contained significant quantities of organotins, the pups' spleens contained smaller quantities, and these decreased rapidly between PND6 and PND12. These results show that organotins cross the placenta and accumulate in fetal tissues but that during lactation, the pups would receive minimal organotins through the milk and during this period, the levels of TBT in pups' tissues decreases rapidly. Consequently, fetuses would be at greater risk of the adverse effects of TBT, but due to the lack of transfer through milk, the risk would be reduced during the lactational period.     

Exposure to perfluorooctane sulfonate during pregnancy in rat and mouse. II: postnatal evaluation.

The postnatal effects of in utero exposure to perfluorooctane sulfonate (PFOS, C8F17SO3-) were evaluated in the rat and mouse. Pregnant Sprague-Dawley rats were given 1, 2, 3, 5, or 10 mg/kg PFOS daily by gavage from gestation day (GD) 2 to GD 21; pregnant CD-1 mice were treated with 1, 5, 10, 15, and 20 mg/kg PFOS from GD 1 to GD 18. Controls received 0.5% Tween-20 vehicle (1 ml/kg for rats and 10 ml/kg for mice). At parturition, newborns were observed for clinical signs and survival. All animals were born alive and initially appeared to be active. In the highest dosage groups (10 mg/kg for rat and 20 mg/kg for mouse), the neonates became pale, inactive, and moribund within 30-60 min, and all died soon afterward. In the 5 mg/kg (rat) and 15 mg/kg (mouse) dosage groups, the neonates also became moribund but survived for a longer period of time (8-12 h). Over 95% of these animals died within 24 h. Approximately 50% of offspring died at 3 mg/kg for rat and 10 mg/kg for mouse. Cross-fostering the PFOS-exposed rat neonates (5 mg/kg) to control nursing dams failed to improve survival. Serum concentrations of PFOS in newborn rats mirrored the maternal administered dosage and were similar to those in the maternal circulation at GD 21; PFOS levels in the surviving neonates declined in the ensuing days. Small but significant and persistent growth lags were detected in surviving rat and mouse pups exposed to PFOS prenatally, and slight delays in eye opening were noted. Significant increases in liver weight were observed in the PFOS-exposed mouse pups. Serum thyroxine levels were suppressed in the PFOS-treated rat pups, although triiodothyronine and thyroid-stimulating hormone [TSH] levels were not altered. Choline acetyltransferase activity (an enzyme that is sensitive to thyroid status) in the prefrontal cortex of rat pups exposed to PFOS prenatally was slightly reduced, but activity in the hippocampus was not affected. Development of learning, determined by T-maze delayed alternation in weanling rats, was not affected by PFOS exposure. These results indicate that in utero exposure to PFOS severely compromised postnatal survival of neonatal rats and mice, and caused delays in growth and development that were accompanied by hypothyroxinemia in the surviving rat pups.     

Adrenal-mediated rather than direct effects of nicotine as a basis of altered sex steroid synthesis in fetal and neonatal rat

Prenatal nicotine interferes with rat sexual brain differentiation and may influence human puberty. We studied effects of nicotine and cotinine on perinatal steroid synthesis in offspring of time-pregnant rats. In vitro, cotinine inhibited testosterone synthesis in neonatal rat testis. Both compounds inhibited the brain aromatase of gestational day (GD) 19 male fetuses. Effective concentrations were higher than levels in maternal plasma and fetal tissue at GD 19 after nicotine treatment from GD 12, even though nicotine accumulated in fetal brain. In view of a dual effect of nicotine in male GD 18 fetuses, decreasing plasma testosterone and increasing corticosterone [Dev Brain Res 1991;62:23-31], we administered metyrapone on GD 17 to nicotine-treated dams. 11beta-Hydroxylase inhibition completely reversed the nicotine-induced reduction of plasma testosterone at GD 18. POMC mRNA in anterior pituitary of nicotine-exposed GD 18 fetuses was reduced, probably as a result of corticosterone feedback. These data reveal a novel type of interaction of nicotine with the fetal gonadal axis involving the adrenal.     

Concentration and persistence of tin in rat brain and blood following dibutyltin exposure during development

Dibutyltin (DBT), a widely used plastic stabilizer, has been detected in the environment as well as human tissues. Although teratological and developmental effects are well documented, there are no published reports of DBT effects on the developing nervous system. As part of a developmental neurotoxicity study of DBT, tissue samples were periodically collected to determine the distribution of total tin (Sn) in brain and whole blood. Pregnant Sprague-Dawley rats were exposed to 0, 10, or 25 ppm DBT in drinking water from gestational day (GD) 6 to weaning at postnatal day (PND) 21. Beginning on PND 3, half of the litters were directly dosed every 2 to 3 d via oral gavage with 0, 1, or 2.5 mg/kg DBT such that the dose level matched the water concentration (for example, litters with 25 ppm DBT in the water received 2.5 mg/kg). For Sn analysis, brain and blood samples were collected from culled pups on PND2 (males and females pooled), from pups (males and females separately) as well as dams at weaning (PND21), and from adult offspring (males and females) at PND93. Total Sn was quantified using inductively coupled plasma-mass spectroscopy (ICP-MS). At all ages, brain Sn levels were higher than blood. At culling, in the directly dosed pups at weaning, and in dams at weaning, Sn levels in both tissues were linearly related to dose. Weanling pups without direct dosing showed lower levels than either culled pups or dams, indicating that lactational exposure was minimal or negligible even while maternal exposure is ongoing. In the adults, Sn levels persisted in brains of directly dosed rats, and the high-dose females had higher levels than did high-dose males. No Sn was detected in adult blood. Thus, during maternal exposure to DBT in drinking water, Sn is placentally transferred to the offspring, but lactational transfer is minimal, if any. Furthermore, Sn is concentrated in brain compared to blood, and its elimination is protracted, on the order of days to months after exposure ends.     

Two-generation reproduction and cross-foster studies of perfluorooctanesulfonate (PFOS) in rats

Perfluorooctanesulfonate (PFOS) is a persistent acid found widely distributed in wildlife and humans. To understand the potential reproductive and developmental effects of PFOS, a two-generation reproduction study was conducted in rats. Male and female rats were dosed via oral gavage at dose levels of 0, 0.1, 0.4, 1.6, and 3.2 mg/(kg day) for 6 weeks prior to mating, during mating, and, for females, through gestation and lactation, across two generations. Due to substantial F1 neonatal toxicity observed in the 1.6 and 3.2 mg/(kg day) groups, continuation into the second generation was limited to F1 pups from the 0, 0.1, and 0.4 mg/(kg day) groups. No adverse effects were observed in F0 females or their fetuses upon caesarean sectioning at gestation day 10. Statistically significant reductions in body-weight gain and feed consumption were observed in F0 generation males and females at dose levels of 0.4 mg/(kg day) and higher, but not in F1 adults. PFOS did not affect reproductive performance (mating, estrous cycling, and fertility); however, reproductive outcome, as demonstrated by decreased length of gestation, number of implantation sites, and increased numbers of dams with stillborn pups or with all pups dying on lactation days 1-4, was affected at 3.2 mg/(kg day) in F0 dams. These effects were not observed in F1 dams at the highest dose tested, 0.4 mg/(kg day). Neonatal toxicity in F1 pups, as demonstrated by reduced survival and body-weight gain through the end of lactation, occurred at a maternal dose of 1.6 mg/(kg day) and higher while not at dose levels of 0.1 or 0.4 mg/(kg day) or in F2 pups at the 0.1 or 0.4 mg/(kg day) dose levels tested. In addition to these adverse effects, slight yet statistically significant developmental delays occurred at 0.4 (eye opening) and 1.6 mg/(kg day) (eye opening, air righting, surface righting, and pinna unfolding) in F1 pups. Based on these data, the NOAELs were as follows: reproductive function: F0> or =3.2 and F1> or =0.4 mg/(kg day); reproductive outcome: F0=1.6 and F1> or =0.4 mg/(kg day); overall parental effects: F0=0.1 and F1> or =0.4 mg/(kg day); offspring effects: F0=0.4 and F1> or =0.4 mg/(kg day). To distinguish between maternal and pup influences contributing to the perinatal mortality observed in the two-generation study, a follow-up cross-foster study was performed. Results of this study indicated that in utero exposure to PFOS causally contributed to post-natal pup mortality, and that pre-natal and post-natal exposure to PFOS was additive with respect to the toxic effects observed in pups.     

Neurobehavioral toxicity study of dibutyl phthalate on rats following in utero and lactational exposure

To investigate the neurobehavioral effects of dibutyl phthalate (DBP), an important endocrine disruptor known for reproductive toxicity, on rodent offspring following in utero and lactational exposure, pregnant Wistar rats were treated with DBP (0, 0.037, 0.111, 0.333 and 1% in the diet) from gestation day (GD) 6 to postnatal day (PND) 28, and selected developmental and neurobehavioral parameters of the offspring were measured. There were no significant effects of DBP on body weight gain of the dams during GD 6–20 or on the pups' ages of pinna detachment, incisor eruption or eye opening. Exposure to 1% DBP prolonged gestation period, decreased body weight in both male and female pups, depressed surface righting (PND 7) in male pups, shortened forepaw grip time (PND 10), enhanced spatial learning and reference memory (PND 35) in male pups. Exposure to 0.037% DBP also shortened forepaw grip time (PND 10), but inhibited spatial learning and reference memory in male pups. Sex × treatment effects were found in forepaw grip time (PND 10), spatial learning and reference memory, and the male pups appeared to be more susceptible than the females. However, all levels of DBP exposure did not significantly alter surface righting (PND 4), air righting (PND 16), negative geotaxis (PND 4 or 7), cliff avoidance (PND 7) or open field behavior (PND 28) in either sex. Overall, the dose level of DBP in the present study produced a few adverse effects on the neurobehavioral parameters, and it may alter cognitive abilities of the male rodent. Copyright © 2009 John Wiley & Sons, Ltd.