Neurotoxicological outcomes of perinatal heptachlor exposure in the rat.

The developing nervous system has been identified as a potential target of pesticide exposure. Heptachlor is a cyclodiene pesticide that was widely used for many years, and for which inadvertent exposure to children and fetuses took place in the early 1980s; yet little is known regarding the developmental neurotoxicity of it and other cyclodienes. The aim of this study was to determine whether perinatal heptachlor exposure results in persistent alterations in nervous system function. Pregnant Sprague-Dawley dams were dosed from gestational day (GD) 12 to postnatal day (PND) 7, whereupon the rat pups were dosed directly until PND 21 (group A) or PND 42 (group B). Dose levels were 0, 0.03, 0.3, or 3 mg/kg/day, po. There were no dose-related effects on maternal weight, litter size, or pup growth. GABA(A) receptor binding (using [(35)S] tert-butylbicyclophosphorothionate; TBPS) and GABA-stimulated Cl- flux were evaluated in control and high-dose brain tissues taken on PND 7, 21, and 43. The B(max) values for [(35)S]-TBPS binding in brainstem, but not cortex, were decreased in female rats across all ages tested. There were no such changes in male rats, nor were K(D) values altered in either tissue or gender. GABA-stimulated Cl- flux was decreased in female cortex synaptoneurosomes only on PND 21. The ontogeny of the righting response (PND 2-5) was delayed in the high-dose females. All subsequent testing took place a week to months after dosing ceased. The functional observational battery (FOB) showed treatment-related, but not necessarily dose-related, changes in different aspects of the rat's reactivity and activity levels. Group-A rats also showed altered within-session habituation of motor activity. There were no heptachlor-related differences in motor activity following challenge with a range of chlordiazepoxide doses. Cognitive assessments were conducted in both groups of rats. There were no statistically significant differences among treatment groups in a one-trial passive avoidance test, although there was a trend toward less learning. In group B, rats (both sexes), heptachlor altered spatial learning in the Morris water maze during two weeks of daily training (2 trials/day). On probe trials, heptachlor-treated rats did not show significant preference for the correct quadrant (all dose groups in males, high dose in females). These rats did not show alterations on subsequent working-memory training (where the platform position was relearned each day). Thus, perinatal exposure to heptachlor produced neurochemical and persistent neurobehavioral changes, including alterations in spatial learning and memory.     

Dopamine transporter binding in the rat striatum is increased by gestational, perinatal, and adolescent exposure to heptachlor.

Heptachlor is a persistent cyclodiene pesticide that affects GABAergic function. Recent reports indicate that heptachlor exposure also alters dopamine transporter (DAT) expression and function in adult mice. The aim of this study was to determine whether gestational, perinatal, and/or adolescent heptachlor exposure in rats altered dopamine-receptor and DAT binding. Adolescent exposure to dieldrin was included to evaluate the generality of the findings. Sprague-Dawley rats received doses (po) ranging from 0 to 8.4 mg/kg/day of heptachlor, or dieldrin, 3 mg/kg/day, during different developmental periods. There were dose-related decreases in maternal weight gain and pup survival, as well as delayed righting reflex, at heptachlor doses > or =3 mg/kg/day. There were no changes in striatal dopamine receptor-D1 ([(3)H]SCH-23390) and -D2 ([(3)H]spiperone) binding in preweanling pups exposed perinatally to heptachlor, and no differences in the response of adult rats to the motor activity-increasing effects of d-amphetamine. However, there were significant (27-64%) increases in striatal DAT binding of [(3)H]mazindol in preweanling rats exposed only gestationally. In rats exposed perinatally and/or during adolescence, there were also increases (34-65%) in striatal DAT binding at postnatal days (PND) 22, 43, and 128. Adolescent exposure to dieldrin also increased DAT binding. In other rats exposed perinatally and throughout adolescence, even the lowest dose of heptachlor 0.3 mg/kg/d increased DAT binding on PND 130. The DAT affinity for mazindol was unchanged in heptachlor-exposed striata. In vitro binding studies indicated that heptachlor (> or =10 microM) displaced mazindol binding. Thus, gestational, perinatal, and/or adolescent exposure to heptachlor produced an increase in DAT binding as early as PND 10, and this change persisted into adulthood.     

Gestational and lactational exposure to heptachlor does not alter reproductive system development in rats

Like other organochlorine insecticides, heptachlor is lipophilic and accumulates in fatty tissues. Earlier studies suggested that in utero exposure to heptachlor decreased fertility in the offspring; neonatal exposure to the closely related insecticide chlordane reportedly delayed puberty and disrupted estrous cycling in females. We hypothesized that in utero and lactational exposure to heptachlor would disrupt the development of the reproductive system in males and females, resulting in altered timing of puberty and abnormal gonadal histology and reproductive hormone levels. Timed pregnant Sprague-Dawley rats were treated by oral gavage with heptachlor in corn oil at doses of 0.5 and 5.0 mg/kg or an equivalent volume of corn oil alone daily from gestational day 8 through post-natal day (PND) 21, the day of weaning (n = 7-8/group). Litters were standardized to 4 males and 4 females on the day of birth. Two of the dams in the 5.0 mg/kg/d group died. Pups in the highest dose group weighed significantly less than those in the other 2 groups on PND 0. All but 1 litter of the 5.0 mg/kg/d group died within the first 4 post-natal days. Age at eye opening was delayed with increasing heptachlor dose. There was no effect of treatment on pup weight gain in the surviving litters, on anogenital distance, age at puberty, nipple retention past the infantile period in males, estrous cycling, serum sex steroid concentrations, reproductive organ weights, or testicular or ovarian histology. These results suggest that heptachlor exposure during gestation and lactation does not disrupt development of the reproductive system in rats.     

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.     

Fertility and developmental neurotoxicity effects of inhaled hydrogen sulfide in Sprague-Dawley rats

In this study, we examined whether perinatal exposure by inhalation to hydrogen sulfide (H2S) had an adverse impact on pregnancy outcomes, offspring prenatal and postnatal development, or offspring behavior. Virgin male and female Sprague-Dawley rats (12 rats/sex/concentration) were exposed (0, 10, 30, or 80 ppm H2S; 6 h/day, 7 days/week) for 2 weeks prior to breeding. Exposures continued during a 2-week mating period (evidence of copulation = gestation day 0 = GD 0) and then from GD 0 through GD 19. Exposure of dams and their pups (eight rats/litter after culling) resumed between postnatal day (PND) 5 and 18. Adult male rats were exposed for 70 consecutive days. Offspring were evaluated using motor activity (PND 13, 17, 21, and 60+/-2), passive avoidance (PND 22+/-1 and 62+/-3), functional observation battery (PND 60+/-2), acoustic startle response (PND 21 and 62+/-3), and neuropathology (PND 23+/-2 and 61+/-2). There were no deaths and no adverse physical signs observed in F0 male or female rats during the study. A statistically significant decrease in feed consumption was observed in F0 male rats from the 80-ppm H2S exposure group during the first week of exposure. There were no statistically significant effects on the reproductive performance of the F0 rats as assessed by the number of females with live pups, litter size, average length of gestation, and the average number of implants per pregnant female. Exposure to H2S did not affect pup growth, development, or performance on any of the behavioral tests. The results of our study suggest that H2S is neither a reproductive toxicant nor a behavioral developmental neurotoxicant in the rat at occupationally relevant exposure concentrations (< or =10 ppm).     

Decreased endurance to cold water swimming and delayed sexual maturity in the rat following neonatal lead exposure

The effects of neonatal lead (Pb) exposure on ability to endure stress and on the onset of sexual maturity were investigated using rats. Sprague-Dawley dams (n = 17/treatment) were treated with or without lead acetate (0.3%) in drinking water from parturition until postnatal day (PND) 21, at which time the pups were weaned. A set of sexbalanced pairs of pups (24 male and 24 females/treatment) from randomly selected control and Pb-treated dams was tested for cold water (4 °C) swimming-endurance on PND 15, 21, 25 and 30. Lead treated-female pups showed significantly (P < 0.05) lower endurance on PND 21 and 30, while Pb-treated males exhibited lower (P < 0.05) endurance on PND 21 compared to their respective controls. The results of this study indicate that neonatal exposure to Pb decreased cold water swimming-endurance. Neonatal exposure to either Pb or swimming stress delayed (P < 0.002) the onset of sexual maturity in both sexes. However, exposure to both treatments masked the effect of swimming stress on the onset of maturity in females but not in males.     

Tissue manganese concentrations in lactating rats and their offspring following combined in utero and lactation exposure to inhaled manganese sulfate.

There is little information regarding the tissue distribution of manganese in neonates following inhalation. This study determined tissue manganese concentrations in lactating CD rats and their offspring following manganese sulfate (MnSO4) aerosol inhalation. Except for the period of parturition, dams and their offspring were exposed to air or MnSO4 (0.05, 0.5, or 1 mg Mn/m3) for 6 h/day, 7 days/week starting 28 days prior to breeding through postnatal day (PND) 18. Despite increased manganese concentrations in several maternal tissues, MnSO4 inhalation exposure did not affect body weight gain, terminal (PND 18) body weight, or organ weights in the dams. Exposure to MnSO4 at 1 mg Mn/m3 resulted in decreased pup body weights on PND 19 and decreased brain weights in some PND 14 to PND 45 pups. Exposure to MnSO4 at > or =0.05 mg Mn/m3 was associated with increased stomach content, blood, liver, and skull cap manganese concentrations in PND 1 pups, increased brain, lung, and femur manganese concentrations in PND 14 pups, and elevated olfactory bulb, cerebellum, and striatum manganese concentrations in PND 19 pups. When compared to controls, MnSO4 exposure to > or =0.5 mg Mn/m3 increased liver and blood manganese concentrations in PND 14 pups and increased liver, pancreas, and femur manganese concentrations in PND 19 pups. Manganese concentrations returned to control values in all offspring tissues by PND 45 +/- 1. Our data demonstrate that neonatal tissue manganese concentrations observed following MnSO4 inhalation are dependent on the MnSO4 exposure concentration and the age of the animal.     

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.     

Perinatal exposure to lead attenuates the conditioned reinforcing properties of cocaine in male rats

The purpose of this study was to examine the effects of developmental lead exposure on drug responsiveness later in the life cycle. Adult female rats were gavaged daily with 0, 8, or 16 mg lead for 30 days before breeding with non-exposed males. The respective exposure regimens were maintained throughout gestation and lactation (perinatal exposure). In Experiment 1, at postnatal day (PND) 30 or 90, pups were trained with 0, 1.25, 2.5, or 5 mg/kg cocaine HCl (IP) in a biased conditioned place preference (CPP) procedure. At both PND 30 and 90, an attenuation in CPP was present in animals exposed to 8 or 16 mg lead relative to control rats. Using an identical lead-exposure regimen, a conditioned place aversion (CPA) procedure with 0, 10, 20, or 40 mg/kg lithium chloride (IP) was employed for Experiment 2. No significant differences were present among pups from each lead-exposure group conditioned and tested at PND 30 or 90, thus suggesting that an impairment of associative mechanisms was not solely responsible for the pattern of attenuation present in Experiment 1. Subsequent analyses of blood-lead in all experiments demonstrated concentrations below 5 microg/dl for all animals at PND 30 and below detectable limits (<1 microg/dl) at PND 90. The findings suggested attenuation in cocaine reinforcement with perinatal lead exposure even though the metal apparently had gained clearance from soft tissue.     

The effect of route, vehicle, and divided doses on the pharmacokinetics of chlorpyrifos and its metabolite trichloropyridinol in neonatal Sprague-Dawley rats.

There is a paucity of data on neonatal systemic exposure using different dosing paradigms. Male CD (Sprague-Dawley derived) rats at postnatal day (PND) 5 were dosed with chlorpyrifos (CPF, 1 mg/kg) using different routes of exposure, vehicles, and single versus divided doses. Blood concentrations of CPF and its primary metabolite, trichloropyridinol, were measured at multiple times through 24 h. Groups included were single gavage bolus versus divided gavage doses in corn oil (one vs. three times in 24h), single gavage bolus versus divided gavage doses in rat milk, and sc administration in dimethyl sulfoxide (DMSO). These data were compared with lactational exposure of PND 5 pups from dams exposed to CPF in the diet at 5 mg/kg/day for 4 weeks or published data from dams exposed to daily gavage with CPF at 5 mg/kg/day. Maternal blood CPF levels were an order of magnitude lower from dietary exposure than gavage (1.1 vs. 14.8 ng/g), and blood CPF levels in PND 5 pups that nursed dietary-exposed or gavage-exposed dams were below the limit of detection. Single gavage doses of 1 mg/kg CPF in corn oil vehicle in pups resulted in CPF blood levels of 49 ng/g and in milk vehicle about 9 ng/g. Divided doses led to lower peak CPF levels. A bolus dose of 1 mg/kg CPF in DMSO administered sc appeared to have substantially altered pharmacokinetics from orally administered CPF. To be meaningful for risk assessment, neonatal studies require attention to the exposure scenario, since route, vehicle, dose, and frequency of administration result in different systemic exposure to the test chemical and its metabolites.     

Development Neurotoxicity Evaluation of Orally Administered Isopropanol in Rats

Developmental Neurotoxicity Evaluation of Orally Administered Isopropanol in Rats. Bates, H. K., McKee, R. H., Bieler, G. S., Gardiner, T. H., Gill, M. W., Strother, D. E., and Masten, L. W. (1994). Fundam. Appl. Toxicol. 22, 152-158.Isopropanol was administered by gavage to timed-mated rats from Gestation Day (GD) 6 through Postnatal Day (PND) 21. Doses administered were 0, 200, 700, or 1200 mg/kg/day in a volume of 5 ml/kg. The dams were allowed to deliver and body weights and food consumption were recorded during gestation and lactation. Pups were counted, examined, sexed, and weighed on PND 0, 4, 7, 13, 17, 21, 36, 49, and 68. Litters were culled to eight pups (4:4 or 5:3 sex ratio) on PND 4 and litters without acceptable numbers of male and female pups were eliminated from the study. Pups were weaned on PND 22, and two pups from each litter and their dams were killed. Six of these pups from each dose group were perfused in situ for histopathological examination of the central and peripheral nervous system. Brains of the remaining pups were divided into four regions and weighed. Maternal liver and kidney weights were recorded. Weaned pups were assessed for day of testes descent or vaginal opening and for motor activity on PNDs 13, 17, 21, 47, and 58; auditory startle on PNDs 22 and 60; and active avoidance on PNDs 60-64. These pups were euthanized and examined on PND 68. One high-dose dam died on PND 15, but there were no other clinical observations or effects on maternal weight, food consumption, or gestation length. Pup survival, weight, sex ratio, and sexual maturation were unaffected. There were no biologically significant findings in the behavioral tests, no changes in organ weights, and no pathological findings that could be attributed to isopropanol exposure. In conclusion, there was no evidence of developmental neurotoxicity associated with isopropanol exposure as high as 1200 mg/kg/day.     

Perinatal Methanol Exposure in the Rat: I. Blood Methanol Concentration and Neural Cell Adhesion Molecules

Although the acute toxicity of methanol is well documented,few studies have addressed the consequences of perinatal exposuresto the low concentrations that are expected to arise from itsproposed use as a component of automobile fuel. This reportdescribes the general research design of a series of studies,the effects of methanol exposures on blood concentrations indams and neonates, and indices of brain development. Four cohortsof Long-Evans pregnant rats, each cohort consisting of an exposure(n=12) and a control (n=12) group, were exposed whole-body to4500 ppm methanol vapor or air for 6 hr daily beginning on GestationDay 6. Both dams and pups were then exposed through PostnatalDay 21 (PND 21). Blood methanol concentrations determined bygas chromatography from samples obtained immediately followinga 6-hr exposure reached approximately 500–800 µg/mlin the dams during gestation and lactation. Average concentrationsfor pups attained levels about twice those of the dams. Selectedoffspring from Cohort 4 were exposed for one additional 6-hrsession at ages that extended out to PND 52. Regression analysesshowed that the blood methanol concentrations of the pups declineduntil about PND 48, at which time their levels approximatedthose of their dams. Such pharmacokinetic differences mightincrease the risks posed to developing organisms. Light-microscopicanalysis showed no significant abnormalities in the brains ofthe methanol-treated animals. However, assays of neural celladhesion molecules (NCAMs) in brains of pups sacrificed on PND4 showed staining for both the 140 and the 180 kDa isoformsto be less intense in the cerebellum of exposed animals. NCAMdifferences were not apparent in animals sacrificed 15 monthsafter their final exposure.     

Developmental Neurotoxicity Evaluation of Orally Administered Isopropanol in Rats

Isopropanol was administered by gavage to timed-mated rats fromGestation Day (GD) 6 through Postnatal Day (PND) 21. Doses administeredwere 0, 200, 700, or 1200 mg/kg/day in a volume of 5 ml/kg.The dams were allowed to deliver and body weights and food consumptionwere recorded during gestation and lactation. Pups were counted,examined, sexed, and weighed on PND 0, 4, 7, 13, 17, 21, 36,49, and 68. Litters were culled to eight pups (4:4 or 5:3 sexratio) on PND 4 and litters without acceptable numbers of maleand female pups were eliminated from the study. Pups were weanedon PND 22, and two pups from each litter and their dams werekilled. Six of these pups from each dose group were perfusedin Situ for histopatho logical examination of the central andperipheral nervous sys tem. Brains of the remaining pups weredivided into four regions and weighed. Maternal liver and kidneyweights were re corded. Weaned pups were assessed for day oftestes descent or vaginal opening and for motor activity onPNDs 13, 17, 21, 47, and 58; auditory startle on PNDs 22 and60; and active avoidance on PNDs 60–64. These pups wereeuthanized and examined on PND 68. One high-dose dam died onPND 15, but there were no other clinical observations or effectson maternal weight, food consumption, or gestation length. Pupsurvival, weight, sex ratio, and sexual maturation were unaffected.There were no biologically significant findings in the behavioraltests, no changes in organ weights, and no pathological findingsthat could be attributed to isopropanol exposure. In conclusion,there was no evidence of developmental neurotoxicity associatedwith isopropanol exposure as high as 1200 mg/kg/day.     

Increased expression of glial fibrillary acidic protein in cerebellum and hippocampus: differential effects on neonatal brain regional acetylcholinesterase following maternal exposure to combined chlorpyrifos and nicotine

Cigarette smoking and environmental exposure to chlorpyrifos during pregnancy could lead to developmental toxicity in the offspring. In the present study, pregnant female Sprague-Dawley rats (300-350 g) were treated daily with nicotine (1 mg/kg, sc) or chlorpyrifos (0.1 mg/kg, dermal) or a combination of nicotine and chlorpyrifos from gestational days (GD) 4-20. Control animals were treated with saline and ethanol. Male offspring from the mothers treated with nicotine alone gained significantly less weight on postnatal day (PND) 30 as compared to control. On PND 7, there was a significant increase in brain acetylcholinesterase (AChE) activity in pups from nicotine- and chlorpyrifos-treated dams, whereas plasma butyrylcholinesterase (BChE) activity was significantly elevated in pups of mothers treated with either chlorpyrifos alone or pesticide combined with nicotine. On PND 30 there was a significant increase in AChE activity in brainstem and cerebellum in all treated male pups. In female pups on PND 30 there was a significant rise in AChE activity in brainstem of chlorpyrifos alone and in cerebellum of the combination nicotine and chlorpyrifos group. Histopathological evaluation demonstrated an increased neuronal cell death in the cerebellum granular cell layer of female offspring from nicotine or combined nicotine with chlorpyrifos group. A rise in glial fibrillary acidic protein (GFAP) immunostaining was observed in the CA1 subfield of hippocampus and cerebellum on PND 30 in female and male offspring of mothers treated with either nicotine or nicotine in combination with chlorpyrifos, but to a lesser extent in males. Data suggest that maternal exposure to nicotine and chlorpyrifos, alone or in combination, produces differential alterations in brain regional AChE activity and expression of GFAP in cerebellum and hippocampus in offspring on PND 30.     

Changes in rat brain cholinesterase activity and muscarinic receptor density during and after repeated oral exposure to chlorpyrifos in early postnatal development.

The effects of repeated oral exposures to the organophosphorus insecticide chlorpyrifos (CPS) on brain muscarinic receptor densities, together with cholinesterase (ChE) activity, were studied in early postnatal rats. Initially, the effects on esterases from lactational exposure to CPS were investigated in young rats by administering CPS (100, 150, or 200 mg/kg subcutaneously in corn oil) to dams 1 day postpartum, yielding a significant body burden of CPS in the dams for possible excretion in the milk. Brain ChE inhibition in pups was less severe than in dams, whereas liver carboxylesterase (CbxE) inhibition in pups was at the same level as in dams. Because of the limited brain ChE inhibition obtained following lactation, pups were exposed to CPS directly by gavage, using 3 dosing regimens to yield a dose response. The rats were gavaged with CPS in corn oil on alternate days from postnatal day (PND) 1 through PND 21. Rats in the low-dosage group received 11 treatments at 3 mg/kg, those in the medium-dosage group received 3 treatments at 3 mg/kg and 8 at 6 mg/kg, and those in the high dosage group received 3 treatments at 3 mg/kg, 4 at 6 mg/kg, and 4 at 12 mg/kg. ChE activity in brain homogenates were inhibited significantly by 29% and 63% in the low- and high-dosage groups, respectively, on PND 22 and by 17% in the high dosage group on PND 40. Muscarinic receptor densities in brain synaptosomes were reduced using 3H-N-methylscopolamine (NMS) and 3H-quinuclidinyl benzilate (QNB) as ligands, with the effects more prominent from 3H-NMS. Densities of both ligands recovered to the control level several days after terminating treatment. The results indicate that pups are apparently exposed to only limited amounts of chlorpyrifos and/or its oxon through the milk when dams are exposed to extremely high chlorpyrifos levels. In addition, repeated direct oral exposures of early postnatal rats to CPS will result in persistent brain ChE inhibition and will transiently reduce muscarinic receptor density.     

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.     

Localization of cytochrome P450 1A1 in a specific region of hydronephrotic kidney of rat neonates lactationally exposed to 2,3,7,8-tetrachlorodibenzo-p-dioxin

Hydronephrosis is typically observed in terata caused by in utero and lactational exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), via the arylhydrocarbon receptor, but the molecular mechanism underlying its pathogenesis is largely unknown. In the present study, pregnant Holtzman rats were treated once by gavage with TCDD (1.0 microg/kg bw) or corn oil on gestation day 15. All dams were allowed to litter, and standardized litters in terms of litter size were then reciprocally cross-fostered on postnatal day (PND) 1. On PND1, pups were divided into four experimental groups: pups exposed only in utero, pups exposed only lactationally, pups not exposed via either route (vehicle control), and pups exposed via both routes. Pups were euthanized on PND21 for further analyses. The TCDD dose used was not overtly toxic to the dams or neonates. The incidence and severity of hydronephrosis were markedly high in pups exposed to TCDD lactationally, but not those exposed in utero. On PND21, cytochrome P450 (CYP) 1A1 was detected predominantly in the outer zone of the medulla of the kidney from all the pups lactationally exposed to TCDD, regardless of the occurrence of hydronephrosis. Interestingly, TCDD concentrations in the cortex, the outer zone of the medulla and the inner zone of the medulla were similar. When adult Holtzman rats were administered TCDD, the induction of CYP1A1 was immunohistochemically detected in the liver but not in the kidney 7 days postadministration. The present findings suggest that TCDD-inducible genes via an AhR-dependent mechanism may be associated with the etiology of hydronephrosis in a particular region of the kidney.     

Effects on the reproductive parameters of two generations of Rattus norvegicus offspring from dams exposed to heptachlor during gestation and lactation

Heptachlor has been targeted for global elimination because of its toxicity and environmental persistence, in accordance with the Stockholm Convention on Persistent Organic Pollutants (POPs). However, there is no regulation of heptachlor use in Mexico, where relatively high levels have been found in maternal breast milk. The aim of this study was to determine the effects of heptachlor on the reproductive system of offspring of two consecutive generations of rats (F1 and F2) from dams orally administered heptachlor during midgestation and lactation. Female offspring were analyzed for different phenotypic, reproductive, and molecular parameters. In the F1 generation, heptachlor treatment induced decreased body weight at weaning, increased female anogenital distance, and delayed vaginal opening. In both generations, serum progesterone levels decreased and estradiol levels remained unchanged, while overexpression of the progesterone receptor was observed in uterine epithelial cells on estrus day. In the F2 generation, expression of the estrogen receptor α increased in the glandular epithelium. Finally, heptachlor treatment did not affect apoptosis in the uterine epithelial cells. Overall, the results indicate that heptachlor induced female reproductive alterations when administered to dams during the perinatal period. Accordingly, exposure to heptachlor may represent a risk for the reproductive health of humans. © 2016 Wiley Periodicals, Inc. Environ Toxicol 32: 856–868, 2017.     

Polybrominated diphenyl ether (PBDE)-induced alterations in vitamin A and thyroid hormone concentrations in the rat during lactation and early postnatal development

In experimental animals fed standard laboratory diets, penta-BDE mixtures can decrease circulating thyroid hormone and liver vitamin A concentrations. A substantial number of pregnant women and their children have marginal vitamin A status, potentially increasing their risk of adverse effects to penta-BDE exposure. The current study investigated the effects of maternal gestational and lactational penta-BDE exposure on thyroid hormone and vitamin A homeostasis in rats of sufficient vitamin A (VAS) or marginal vitamin A (VAM) status and their offspring. Dams were administered daily oral doses of 18 mg/kg DE-71 (a penta-BDE mixture) or a corn oil vehicle from gestation day 6 through lactation day (LD) 18. Thyroid hormone and vitamin A homeostasis were assessed in plasma and tissues of LD 19 dams and postnatal day (PND) 12, 18, and 31 pups. DE-71 exposure induced hepatomegaly in VAS and VAM pups at all timepoints and increased testes weights at PND 31. While liver vitamin A concentrations were low in DE-71 treated dams and pups, plasma retinol concentrations and plasma retinol binding protein levels were only low in VAM animals exposed to DE-71. DE-71 exposure lowered plasma thyroxine concentrations in VAS and VAM dams and pups. Plasma thyroid stimulating hormone concentrations were high in VAM dams exposed to DE-71, suggesting that marginal vitamin A status enhances the susceptibility to thyroid hormone axis disruption by DE-71. These results support the concept that marginal vitamin A status in pregnant women may increase the risk for PBDE-induced disruptions in vitamin A and thyroid hormone homeostasis.     

Gestational PFOA exposure of mice is associated with altered mammary gland development in dams and female offspring.

Perfluorooctanoic acid (PFOA), with diverse and widespread commercial and industrial applications, has been detected in human and wildlife sera. Previous mouse studies linked prenatal PFOA exposure to decreased neonatal body weights (BWs) and survival in a dose-dependent manner. To determine whether effects were linked to gestational time of exposure or to subsequent lactational changes, timed-pregnant CD-1 mice were orally dosed with 5 mg PFOA/kg on gestation days (GD) 1-17, 8-17, 12-17, or vehicle on GD 1-17. PFOA exposure had no effect on maternal weight gain or number of live pups born. Mean pup BWs on postnatal day (PND) 1 in all PFOA-exposed groups were significantly reduced and decrements persisted until weaning. Mammary glands from lactating dams and female pups on PND 10 and 20 were scored based on differentiation or developmental stages. A significant reduction in mammary differentiation among dams exposed GD 1-17 or 8-17 was evident on PND 10. On PND 20, delays in normal epithelial involution and alterations in milk protein gene expression were observed. All exposed female pups displayed stunted mammary epithelial branching and growth at PND 10 and 20. While control litters at PND 10 and 20 had average scores of 3.1 and 3.3, respectively, all treated litters had scores of 1.7 or less, with no progression of duct epithelial growth evident over time. BW was an insignificant covariate for these effects. These findings suggest that in addition to gestational exposure, abnormal lactational development of dams may play a role in early growth retardation of developmentally exposed offspring.