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. Author manuscript; available in PMC: 2019 Mar 1.
Published in final edited form as: Neurotoxicology. 2018 Mar;65:125–134. doi: 10.1016/j.neuro.2018.01.008

Ahr and Cyp1a2 genotypes both affect susceptibility to motor deficits following gestational and lactational exposure to polychlorinated biphenyls

Breann T Colter 1,*, Helen Frances Garber 1,2,*, Sheila M Fleming 3,4, Jocelyn Phillips Fowler 1, Gregory D Harding 1, Molly Kromme Hooven 1, Amy Ashworth Howes 1, Smitha Krishnan Infante 1, Anna L Lang 1,5, Melinda Curran MacDougall 6, Melinda Stegman 1, Kelsey Rae Taylor 1, Christine Perdan Curran 1
PMCID: PMC5857246  NIHMSID: NIHMS943873  PMID: 29409959

Abstract

Polychlorinated biphenyls (PCBs) are persistent organic pollutants known to cause adverse health effects and linked to neurological deficits in both human and animal studies. Children born to exposed mothers are at highest risk of learning and memory and motor deficits. We developed a mouse model that mimics human variation in the aryl hydrocarbon receptor and cytochrome P450 1A2 (CYP1A2) to determine if genetic variation increases susceptibility to developmental PCB exposure. In our previous studies, we found that high-affinity AhrbCyp1a2(−/−) and poor-affinity AhrdCyp1a2(−/−) knockout mice were most susceptible to learning and memory deficits following developmental PCB exposure compared with AhrbCyp1a2(+/+) wild type mice (C57BL/6J strain). Our follow-up studies focused on motor deficits, because human studies have identified PCBs as a potential risk factor for Parkinson’s disease. Dams were treated with an environmentally relevant PCB mixture at gestational day 10 and postnatal day 5. We used a motor battery that included tests of nigrostriatal function as well as cerebellar function, because PCBs deplete thyroid hormone, which is essential to normal cerebellar development. There was a significant effect of PCB treatment in the rotarod test with impaired performance in all three genotypes, but decreased motor learning as well in the two Cyp1a2(−/−) knockout lines. Interestingly, we found a main effect of genotype with corn oil-treated control Cyp1a2(−/−) mice performing significantly worse than Cyp1a2(+/+) wild type mice. In contrast, we found that PCB-treated high-affinity Ahrb mice were most susceptible to disruption of nigrostriatal function with the greatest deficits in AhrbCyp1a2(−/−) mice. We conclude that differences in AHR affinity combined with the absence of CYP1A2 protein affect susceptibility to motor deficits following developmental PCB exposure.

Keywords: Polychlorinated biphenyls, aryl hydrocarbon receptor, CYP1A2, nigrostriatal pathways, motor function, cerebellum

1. Introduction

Polychlorinated biphenyls (PCBs) are widespread persistent organic pollutants linked to numerous human health problems, with the most serious effects seen in children of exposed mothers (Ross 2004; Schantz et al., 2003; Jacobson et al., 2003). They are number 5 on the U.S. government’s list of priority pollutants (ATSDR 2015). Worldwide, an estimated 200 billion kg remain in the environment (WHO 2003). The primary route of exposure is contaminated food, especially fatty fish, meat and dairy products (Malisch and Kotz, 2014; Langer et al., 2007; Gomara et al., 2005). New sources of PCB exposure have been reported with the inadvertent production of highly toxic PCB congeners (e.g. PCB 77 and PCB 153) during the synthesis of paint pigments (Anezaki et al., 2015; Hu & Hornbuckle, 2010) and the discovery of airborne PCBs near rural and urban schools (Marek et al., 2017). PCBs will remain a problem for generations because highly exposed cohorts are now reaching reproductive age (Bányiová et al., 2017; Quinn et al,. 2011).

Multiple human studies found deficits in motor function in children exposed to high levels of PCBs (Boucher et al., 2016; Wilhelm et al., 2008; Vreugdenhil et al., 2002; Stewart et al., 2000). The hydroxylated metabolite 4-OH-CB 107 can also cause motor deficits in highly exposed children (Berghuis et al., 2013). In adults, an increased risk of Parkinson’s disease (PD) was reported in women with high workplace exposures (Steenland et al., 2006) and in adults who consumed contaminated whale meat and blubber (Petersen et al., 2008). Rodent studies found adverse PCB effects in both the striatum (Caudle et al., 2006; Chishti et al., 1996) and cerebellum (Nguon et al., 2005). PCB effects on dopamine, the major neurotransmitter associated with motor function, are well known (Seegal et al., 1986, 1994, 1997, 2005).

PCBs occur as mixtures of coplanar congeners which can bind and activate the aryl hydrocarbon receptor and non-coplanar congeners which do not. Human studies clearly show differential responses to PCBs and related AHR agonists (Marek et al., 2014; Novotna et al., 2007; van Duursen et al., 2005; Tsuchiya et al., 2003). The AHR regulates three members of the cytochrome P450 family: CYP1A1, CYP1A2 and CYP1B1.The level of CYP1A2 found in human livers varies about 60-fold (Nebert & Dalton 2006), and maternal CYP1A2 can sequester planar pollutants to prevent transfer to offspring (Curran et al., 2011a; Dragin et al., 2006,). In humans, there is a greater than 12-fold difference in the inducibility of CYP1A1, although the polymorphism responsible has not been identified (Nebert 2017; Nebert et al. 2013).

We developed a mouse model to mimic human variation in the AHR and CYP1A2 to better understand genetic susceptibility to PCBs and similar pollutants. High-affinity Ahrb mice will respond to low levels of xenobiotics such as dioxin and coplanar PCBs whereas poor-affinity Ahrd mice are considered non-responders. To model the wide variation in CYP1A2, we used wild type Cyp1a2(+/+) mice and Cyp1a2(−/−) knockout mice (Denison & Faber 2017; Nebert 2017; Nebert & Dalton, 2006). We previously showed that both high-affinity AhrbCyp1a2(−/−) knockout mice and poor-affinity AhrdCyp1a2(−/−) mice were more susceptible to learning and memory deficits when exposed to an environmentally relevant mixture of PCBs during gestation and lactation compared with AhrbCyp1a2(+/+) wild type mice (Curran et al. 2011a-b, 2012). The studies described here extend those findings by testing the hypothesis that there is similar genetic susceptibility to PCB-induced motor deficits. Our motor battery was also designed to help clarify if PCBs affect motor function by disruption of nigrostriatal pathways or primarily target the cerebellum.

2. Materials and Methods 2.1 Animals

Three genotypes of mice were included. High-affinity AhrbCyp1a2(+/+) wild type mice were purchased from The Jackson Laboratory (Bar Harbor, ME) as C57BL/6J mice, which was the background strain for the two knockout lines used: AhrbCyp1a2(−/−) and poor-affinity AhrdCyp1a2(−/−). All animals were housed in standard shoebox polysulfone cages with corncob bedding and one 5.1 cm2 nestlet per week as enrichment. Water and Lab Diet 5015 chow were provided ad libitum.

Animals were kept on a 12h/12h light-dark cycle with all experiments conducted during the light cycle. Genotype was confirmed at the end of the behavior experiments. All experiments were approved by the Northern Kentucky University Institutional Animal Care and Use Committee. All husbandry and handling was in accordance with the Eighth Guide for the Care and Use of Laboratory Animals and the ARRIVE guidelines.

2.2 Breeding

Nulliparous females between 2.5 and 4 months of age were mated on a four-day breeding cycle with males of the same genotype. Females were separated from males the morning when a vaginal plug was found. Litters were culled or cross-fostered to balance litter size at 6 pups per dam, matching pups with dams of the same genotype and treatment. Pups were weaned at postnatal day 25, group housed by genotype, sex and treatment, and behavioral testing began at P60.

2.3 Treatments

We used the same dosing regimen (Table 1) described in our prior studies (Curran et al. 2011a,b), which was based on the toxic equivalency factors for coplanar PCBs 77, 126 and 169. Noncoplanar (PCB 105, 118, 138, 153, 180) congeners were selected based on reports of their high toxicity and prevalence in the human food supply (Van den berg et al., 2006; Costabeber et al., 2006; Langer et al., 2007). Controls were treated with the corn oil vehicle (Kroger, unstripped). Dams were randomly assigned to treatment groups and treated by gavage at a volume of 10μl/g body weight. at gestational day 10 (GD 10) and postnatal day 5 (PND 5). The PCB doses used in the current study were higher than typical human exposures, but consistent with risk assessment principles requiring higher doses of a toxicant to observe toxicity in the short timeframe of a typical animal study (Doull 2003) and at the low end of cumulative exposures used in other rodent studies of PCB neurotoxicity (Lein et al. 2007; Lee et al. 2011).

Table 1.

PCB congeners and concentrations used in dosing mixture.

PCB congener Planarity IUPAC # Dose
2,3,3',4,4'-Pentachlorobiphenyl non-coplanar 105 10 mg/kg
2,3',4,4',5-Pentachlorobiphenyl non-coplanar 118 10 mg/kg
2,2',3,4,4',5'-Hexachlorobiphenyl non-coplanar 138 10 mg/kg
2,2',4,4',5,5'-Hexachlorobiphenyl non-coplanar 153 10 mg/kg
2,2',3,4,4',5,5'-Heptachlorobiphenyl non-coplanar 180 10 mg/kg
3,3'4,4'-Tetrachlorobiphenyl coplanar 77 5 mg/kg
3,3',4,4',5-Pentachlorobiphenyl coplanar 126 25 μg/kg
3,3',4,4',5,5'-Hexachlorobiphenyl coplanar 169 250 μg/kg
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2.4 Chemicals

Polychlorinated biphenyl congeners were ordered from UltraScientific (N. Kingstown, RI). Unless otherwise noted, all other reagents were purchased from Sigma-Aldrich (St. Louis, MO).

2.5 Western blot

CYP1A1 induction was confirmed in high-affinity Ahrb mice using livers collected from P30 littermates of animals used in behavior. The liver was removed, rinsed in ice-cold phosphate buffered saline, blotted and stored at −80°C until processing. Approximately 500 mg of tissue per animal was homogenized using a polytron homogenizer and a buffer of 0.25 M sucrose, 10 mM HEPES, 1 mM Na2EDTA, and 1 mM EGTA with 0.1% bovine serum albumin (BSA). The buffer was adjusted to pH 7.2 using KOH. Microsomes were prepared using multiple centrifugations to remove cellular debris and other organelles before ultracentrifugation at 40,000 g for 40 min. Microsomes were resuspended in 1 ml of the homogenization buffer. Protein concentrations were determined by the Bradford assay (Sigma-Aldrich, St. Louis MO), following the manufacturer’s protocol. Microsomal proteins (10 μg/lane) were separated on 12% mixed alcohol-detergent-polyacrylamide gel electrophoresis (MAD–PAGE) under denaturing conditions (Brown, 1988). Separated proteins were transferred to PVDF membranes. Western blot analysis was performed using rabbit polyclonal anti-CYP1A1 antibody (Millipore AB1247) and a horseradish peroxidase-conjugated secondary antibody (Daiichi). The SuperSignal Pico enhanced chemiluminescence system (Pierce) was to detect primary antibody binding, with exposure times ranging from 10 to 60 sec. N = 4–6 per group.

2.6 EROD assay

Microsomes from liver, cortex and cerebellum were prepared as described in the previous section to measure CYP1A1 enzymatic activity in PCB-exposed and control animals using the well-known ethoxyresorufin-O-deethylase (EROD) assay. Unknowns were quantified using a standard curve, and purified human CYP1A1 was used as a positive control following the methods of Thompson et al. (2010). N = 5–9 per group.

2.7 Glutathione assay

Reduced glutathione (GSH) and oxidized GSSG were measured in liver, cortex and cerebellum using a standard kit (Cayman Chemicals, Ann Arbor MI) and following the manufacturer’s protocols N = 3–5 per group.

2.8 Motor function tests

One male and one female from each litter were randomly assigned to behavioral testing. A comprehensive battery of tests was used to assess function in both the cerebellum and nigrostriatal pathways. Each animal went through the same experimental protocol, and each animal was limited to one test per day. Experiments were conducted within a 4 h time block to avoid confounding by circadian rhythms. Experiments are described in the order in which they were performed. Animal handlers and those analyzing the data were naïve to the genotype and treatment. N ≥ 16 litters per group (Table 2) Video demonstrating the techniques and equipment can be viewed at: https://www.youtube.com/watch?v=TximAxZcomk

Table 2.

Group sizes for behavioral experiments

Genotype Treatment N per group
AhrbCyp1a2(+/+) Corn oil 16
AhrbCyp1a2(+/+) PCB 19
AhrbCyp1a2(−/−) Corn oil 16
AhrbCyp1a2(−/−) PCB 17
AhrdCyp1a2(−/−) Corn oil 21
AhrdCyp1a2(−/−) PCB 16
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2.8.1 Rotarod

Mice were acclimated to the rotarod apparatus for one day at 0 rpm and one day at a constant 2 rpm speed. Testing was conducted with the rotarod set to accelerate from 1–20 rpm over 180 s, for a maximum trial of 300 s. Latency to fall was recorded. Mice received 3 trials per day for 5 days with an inter-trial interval of 5 min. Rotarod is one of the most widely used tests of cerebellar function (Nadler et al. 2006).

2.8.2 Gait analysis

The hind paws of mice were coated with nontoxic paint, then the mice walked down a 5 cm x 28 cm alley into a black-lined escape cage. Two days were used for training followed by a test day with three trials per day. Trials where mice ran or stopped were not included in the analysis. Stride length and stride width were measured, and the differential between the longest and shortest strides was calculated during analysis. Deficits in hind limb control result in uneven gait patterns in rodents (Fleming et al., 2004). Striatal lesions would result in more steps and shorter steps. Cerebellar lesions would result in unequal stride lengths. No mouse received more than 6 trials over 2 days before completing the task successfully.

2.8.3 Sticker removal

The sticker removal test was used to assess sensorimotor function (Schallert, 1988). Mice must detect the presence of a round 6.35 mm sticker on their snout and use their front paws to remove it. The latency to remove the sticker was recorded. Each mouse received a single test unless the sticker fell off or was not correctly placed on the snout. Mice not removing the sticker after 60 s were assigned a time of 60 s. No mouse received more than 3 trials.

2.8.4 Pole test

The pole test measures gross motor coordination and can detect impairments in nigrostriatal pathways (, Fernagut et al., 2003; Sedelis et al. 2001), which can be reversed by treatment with L-DOPA (Matsuura et al., 1997; Ogawa et al. 1985, 1987). A 50cm vertical pole of 8 mm diameter was placed inside the animal’s home cage. Each mouse was placed at the top of the pole, with its head facing upward. Mice were trained to turn and climb down to the home cage for 2 days and tested on the Day 3. The time to turn and time to descend were recorded. Each mouse received 5 trials per day with a 60 s inter-trial interval.

2.8.5 Challenging balance beam

Mice were trained for two days (5 trials per day) on a smooth beam decreasing in width from 35 to 5cm. Each beam segment was 25 cm in length for a total length of 1 m. Mice were tested on the third day with the beam covered by a wire mesh. Latency to cross, steps and slips were recorded, and the slip:step ratio was calculated during analysis. This protocol has been successfully used to assess nigrostriatal deficits in alpha-synuclein over-expressing mice and other PD mouse models. (Schultheis 2013, Fleming et al. 2004).

2.9 Data analysis

Behavior and EROD data were analyzed using SAS Proc Mixed Models Analysis of Variance with litter as the unit of analysis. We examined the main effects of treatment, genotype, and sex and their interactions. For rotarod, day was included as a repeated measure. When differences were found, we examined simple effects using the SAS “Slice” command with a correction for multiple post-hoc analyses. Measurements of reduced glutathione and oxidized GSSG were compared against control mice using the Holm-Sidak method. Data are presented as least square means ± the standard error of the mean (SEM). Western blot data is simply reported as CYP1A1 present or not, indicating whether or not AHR activation occurred.

3. Results

3.1 Assessment of AHR activation by CYP1A1 upregulation

Our Western blot and EROD results confirm that the AHR is only activated in PCB-treated high-affinity Ahrb mice and not in poor-affinity Ahrd mice or the corn oil-treated controls. At P30, levels of CYP1A1 protein were highest in livers of PCB-treated AhrbCyp1a2(−/−) knockout mice, but CYP1A1 protein was also present in PCB-treated AhrbCyp1a2(+/+) wild type mice (Fig 1). The EROD assays showed significantly higher CYP1A1 activity in the livers of P30 Ahrb mice (P < 0.001) with nearly undetectable activity in PCB-treated AhrdCyp1a2(−/−) knockout mice and the corn oil-treated controls (Fig 2A). There was a trend for higher EROD activity in the cerebellum of AhrbCyp1a2(−/−) knockout mice (P = 0.08) compared with all other groups (Fig 2B). Similar trends (P = 0.06) were seen in cortex (data not shown).

Fig. 1.

Fig. 1

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Western blot of CYP1A1 in liver. CYP1A1 is an inducible enzyme typically not detectable in tissues unless the aryl hydrocarbon (AHR) is activated. Our Western blot analysis confirmed the AHR was only activated in high-affinity Ahrb mice compared with corn oil-treated controls and poor-affinity Ahrd mice.

Fig. 2.

Fig. 2

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Fig. 2A EROD activity in liver. There was a significant gene x treatment interaction with higher EROD activity in liver of Ahrb mice compared with poor-affinity Ahrd mice. N ≥ 9 per group. *** P < 0.001

Fig 2B EROD activity in cerebellum. There was a trend for higher EROD activity in the cerebellum of AhrbCyp1a2(−/−) mice. P = 0.08. N ≥ 5 per group.

3.2 Assessment of oxidative stress

Glutathione levels were significantly higher (F2,11 = 8.48; P < 0.05) in the liver of PCB-treated AhrbCyp1a2(−/−) knockout mice as well as levels of oxidized glutathione (GSSG) (F2,11 = 16.75; P < 0.01), indicating that oxidative stress response had been induced in these animals. There were no differences by treatment or genotype for GSH levels in the cortex or cerebellum; however PCB-treated AhrbCyp1a2(−/−) knockout mice had significantly higher levels of GSSG in the cortex (F2,11 = 9.97; P < 0.05) (Table 3)

Table 3.

Measurements of reduced and oxidized glutathione in live and cortex. Bold indicates significant differences from control mice

Genotype Treatment GSH-Liver GSSG-Liver GSSG-Cortex
AhrbCyp1a2(+/+) corn oil 51 ± 5.8 12.5 ± 0.5 6.0 ± 0.8
AhrbCyp1a2(+/+) PCB 43.5 ± 4.5 11.3 ± 0.4 7.0 ± 0.6
AhrbCyp1a2(−/−) PCB 64.8 ± 5.8 14.3 ± 0.5 9.7 ± 0.8
AhrdCyp1a2(−/−) PCB 50.8 ± 6.9 12.8 ± 0.7 9.1 ± 1.0
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3.3 Motor function test results

Data from behavior experiments are presented in the order in which they were performed.

3.3.1 Rotarod results

There was a significant main effect of genotype with both groups of Cyp1a2(−/−) mice having shorter latencies to fall compared with wild type mice (F2,205 = 14.74; P < 0.0001) and a significant main effect of treatment with PCB-treated mice having shorter latencies to fall (F2,205 = 16.81; P < 0.0001). All groups of mice did show motor learning over the 5 days of testing with a significant main effect of day (F4,587 = 75.83; P < 0.0001); however, there was a significant gene x day interaction with AhrbCyp1a2(+/+) wild type mice showing significantly greater improvements over the 5 days of testing (F8,656 = 4.26; P< 0.0001. Female knockouts also showed the greatest impairments compared with controls (Figs. 3A–D).

Fig. 3.

Fig. 3

Fig. 3

Fig. 3

Fig. 3

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Fig. 3A Rotarod performance by genotype. There was a main effect of genotype with Cyp1a2(−/−) having significantly shorter latencies to fall off the rotarod. *** P < 0.001.

Fig. 3B Rotarod performance by treatment. There was a main effect of treatment on rotarod performance with PCB-treated mice having significantly shorter latencies to fall off the rotarod. *** P < 0.001.

Fig. 3C Rotarod performance over time (motor learning). All groups of mice showed improvement over 5 days of testing, but high-affinity AhrbCyp1a2(+/+) showed greater motor learning compared with Cyp1a2(−/−) knockout mice with a significant gene x day interaction. P < 0.001.

Fig. 3D. Sex differences in rotarod performance. Female mice with the Cyp1a2(−/−) genotype showed the greatest impairments on the rotarod test compared with Cyp1a2(+/+) wild type mice. *P < 0.05, ** P < 0.01, *** P <0.001.

3.3.2 Gait analysis

We found a significant main effect of genotype with both groups of Cyp1a2(−/−) mice having longer strides compared with wild type mice (F2,248 = 16.95; P < 0.0001) and a significant gene x treatment interaction (F2,248 = 5.67; P < 0.01). Both lines of Cyp1a2(−/−) knockout mice had longer strides than wild type mice, and PCB-treated AhrbCyp1a2(+/+) wild type mice had shorter stride lengths than all other groups (Fig. 4A–B). There was also a significant main effect of genotype for stride width (Fig. 4C) with AhrbCyp1a2(−/−) knockout mice having significantly narrower strides (F2,248 = 10.48; P < 0.001). There were no significant differences by genotype or treatment for stride differential (P > 0.05).

Fig. 4.

Fig. 4

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A. Gait stride length. AhrbCyp1a2(+/ + ) wild type mice had significantly shorter strides than all other groups. ** P < .001. B. Gait stride length. PCB-treated AhrbCyp1a2(+/ + ) wild type mice had significantly shorter strides than all other groups. ** P < .01.C. Gait stride width. AhrbCyp1a2(−/−) mice had significantly narrower strides compared with the other two genotypes, but there was no effect of PCB treatment. *** P < .001.

3.3.3 Sticker removal

PCB-treated AhrbCyp1a2(−/−) knockout mice had the longest latencies to remove the adhesive sticker; however, the differences were not statistically significant (F2,281 = 1.37; P = 0.26). There was a trend for a genotype effect (F2,281 = 2.80; P = 0.062) with AhrbCyp1a2(−/−) knockout mice having the longest latencies compared with wild type mice and AhrdCyp1a2(−/−) knockout mice (Fig. 5).

Fig. 5. Latency to remove adhesive sticker.

Fig. 5

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There was a trend for a gene x treatment interaction with PCB-treated AhrbCyp1a2(−/−) mice having the longest latencies of all groups. P = 0.06.

3.3.4 Pole test

There were no differences in the time to turn or the time to descend the pole based on treatment; however, females had significantly shorter turn (F1,295 = 5.56; P <0.05) and descent times compared with males (F1,295 = 4.74; P <0.05), and AhrdCyp1a2(−/−) knockout mice had significantly shorter turn (F2,295 = 4.27; P <0.05) and descent times (F2,295 = 7.23; P <0.001) compared with Ahrb knockout and wild type mice (Figs. 6A–B).

Fig. 6.

Fig. 6

Fig. 6

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Fig. 6A Pole turn time.

There was a main effect of genotype, but no effect of PCB treatment with poor-affinity AhrdCyp1a2(−/−) mice having the shortest latencies to turn downward on the pole. * P < 0.05.

Fig. 6B Pole descent time. There was a main effect of genotype, but no effect of PCB treatment with poor-affinity AhrdCyp1a2(−/−) having the shortest latencies to descend the pole back to the home cage. ** P < 0.01.

3.3.5 Challenging balance beam

There was a significant main effect of genotype for latency to cross the balance beam with AhrdCyp1a2(−/−) knockout mice having significantly shorter latencies (F2,121 = 3.36; P <0.05) compared with Ahrb knockout and wild type mice (Fig. 7A). There was a significant gene x treatment interaction (F2,117 = 6.54; P <0.01) for the number of slips with PCB-treated Ahrb mice having more slips while PCB-treated AhrdCyp1a2(−/−) knockout mice had significantly fewer slips (Fig. 7B). There was also a significant gene x treatment interaction (F2,117 = 6.07; P <0.01) for the ratio of slips/steps with PCB-treated Ahrb mice having more slips per step while PCB-treated AhrdCyp1a2(−/−) knockout mice had significantly fewer slips per step (Fig. 7C).

Fig. 7.

Fig. 7

Fig. 7

Fig. 7

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Fig. 7A. Challenging beam latency. AhrdCyp1a2(−/−) mice had the shortest latency to cross the balance beam, regardless of treatment. * P < 0.05

Fig. 7B Mean slips on challenging beam. There was a significant gene x treatment interaction with PCB-treated Ahrb mice having more slips than corn oil-treated controls of the same genotype and poor-affinity Ahrd mice having significantly fewer than corn oil-treated controls of the same genotype. ţ P < 0.1, ** P < 0.01.

Fig. 7C Slip to Step ratio on challenging beam. There was a significant gene x treatment interaction with PCB-treated Ahr bCyp1a2(−/−) mice having more slips per step than their corn oil-treated controls mice and poor-affinity Ahrd Cyp1a2(−/−) mice having significantly fewer slips per step than their corn oil-treated controls. ** P < 0.01.

4. Discussion and conclusions

Our data confirm that an environmentally relevant PCB mixture only activates the AHR in high-affinity Ahrb mice and that Cyp1a2(+/+) wild type mice show fewer motor impairments following developmental PCB exposure. This supports our previous findings (Curran et al. 2011b, 2012) and extends them to motor deficits. We did not find evidence of significant oxidative stress following PCB exposure, although it appears that the antioxidant response system had been activated, since both oxidized and reduced glutathione levels were significantly increased in the most susceptible AhrbCyp1a2(−/−) mice (Table 3).

Our motor battery was designed to assess both nigrostriatal pathways and cerebellar function, and the data are consistent with impairments in those brain regions. PCB-treated mice from all three genotypes showed impaired performance on the rotarod compared to corn oil-treated controls, suggesting a cerebellar deficit. Motor learning was reduced in the two knockout lines compared with wild type mice. Interestingly, there was also a motor deficit in both corn oil-treated lines of Cyp1a2(−/−) mice. There is evidence to suggest CYP1A2 has a normal function in the brain. CYP1A2 is differentially regulated in the cortex and cerebellum (Iba et al., 2003). The enzyme can also be induced in the brain in a region-specific manner with highest levels seen in the pons, medulla, cerebellum, frontal cortex (Yadav et al., 2006) and hypothalamus (Korkalainen et al., 2005). CYP1A2 is normally down-regulated during cerebellar granule cell migration (Mulero-Navarro et al. 2003). Together with our findings, these data warrant further study of CYP1A2’s role in cerebellar development and function.

Results from the gait analysis did not support our hypothesis of genetic susceptibility in Cyp1a2(−/−) mice. In contrast, PCB-treated AhrbCyp1a2(+/+) mice had shorter strides than corn oil-treated control mice and both groups of PCB-treated knockout mice. Nigrostriatal lesions result in shorter strides (Fleming et al. 2004), so this suggests some impairment in the PCB-treated wild type mice. We previously reported striatal dopamine levels were significantly lower in PCB-treated AhrbCyp1a2(−/−) mice compared with PCB-treated AhrbCyp1a2(+/+) mice (p < 0.01) (Curran et al. 2011a), so it’s unlikely this impairment was caused by an absolute loss of dopamine.

Our findings are consistent with Caudle et al. (2006) who reported a significant decrease in expression of the dopamine transporter (DAT) in PCB-treated C57BL/6J mice, but no difference in striatal dopamine levels. Bemis and Seegal (2004) reported that PCB mixtures inhibit the vesicular monoamine transporter (VMAT) in synaptosomes from adult Long-Evans rats. Interestingly, Akahoshi et al. (2009, 2012) demonstrated that the liganded AHR upregulates tyrosine hydroxylase, the rate-limiting enzyme in dopamine production. Together, this suggests future work is needed to examine the inter-related effects of PCB mixtures on dopamine production, metabolism and transport.

The major finding in the pole test was a shorter latency to turn and descend the pole by AhrdCyp1a2(−/−) knockout mice. This difference could result from increased anxiety and greater motivation to return to the home cage or hyperactivity which has also been reported in PCB neurotoxicity studies. Our prior studies with poor affinity AhrdCyp1a2(+/+) and AhrdCyp1a2(−/−) mice found no evidence of hyperactivity in open field locomotor testing, fewer zone crossings in the elevated zero maze (Curran et al. 2011b) and a heighted startle response in AhrdCyp1a2(−/−) mice (Curran et al. 2012). This argues in favor of an anxiety-like phenotype in Ahrd mice and against hyperactivity. Since latencies were shorter than control mice, the results cannot be interpreted as an impairment in motor function.

The challenging balance beam results support the hypothesis that AhrdCyp1a2(−/−) knockout mice have higher motivation to reach the goal box or home cage. Both PCB-treated and control mice from this line had significantly shorter latencies to traverse the beam compared with Ahrb mice. Both PCB-treated Ahrb groups showed a trend toward more slips, which can be an indication of an impairment in nigrostriatal function.

Efforts to identify a mode of action responsible for the distinct patterns of PCB-induced motor impairments observed in these experiments will need to consider both the genetic differences and the well-established model of noncoplanar PCB neurotoxicity mediated by the ryanodine receptor (RyR) and calcium dysregulation (Dingemans et al. 2016, Pessah et al. 2010, Roegge et al. 2006, Gafni et al. 2004). We note that our PCB mixture does contain noncoplanar PCBs, but the congeners included do not have the same high potency at the ryanodine receptor as its prototypical agonist PCB 95. Therefore, it is likely that an alternate mechanism or mechanisms are needed to explain all of the observed effects. In support of that concept, Roegge et al. (2006) found increased levels of RyR1 in PCB-treated Long-Evans rats, but no evidence of increased receptor binding when looking at changes in the cerebellum. Meanwhile, Nguon et al (2005) reported increased GFAP expression and reduced cerebellar mass in Sprague-Dawley rats exposed to Aroclor during development. Males had more severe motor impairments on rotarod and greater increases in GFAP compared with females. In contrast, we found greater impairments in female mice. Llansola et al. (2009) identified the glutamate–NO–cGMP pathway as a cerebellar target of both coplanar PCB 126 and noncoplanar PCB 153 in Wistar rats. A follow-up study confirmed motor deficits for both PCB126 and PCB 153 with younger males at higher risk (Cauli et al. 2013).

A limitation of our studies was the use of gavage dosing. Although this allows precise dosing, it would be important to repeat these experiments using a chronic, low-level exposure in food. This is commonly done in rat studies (Miller et al. 2017, Roegge et al. 2006, Widholm et al. 2004), but can be a challenge when using mice which consume much smaller quantities of food each day. In addition, longer-term studies would allow an assessment of motor function in aged mice, which would better match the typical course of human Parkinson’s disease patients. Since behavioral testing began at P60, it is possible that only minor impairments would be seen in nigrostriatal pathways at that age with more pronounced effects in animals over 1 y of age.

Further work is needed to assess gene expression in the substantia nigra, striatum and cerebellum at multiple time points to verify that there were no adverse effects on dopaminergic neurons involved in motor function and to identify potential modes of action for the observed motor deficits. A histological examination of those tissues could also reveal morphological changes related to abnormal development. It would also be important to look at changes related to thyroid hormone depletion, because we previously found that circulating thyroxine (T4) levels were reduced 80% in AhrbCyp1a2(−/−) knockout mice compared with control levels at P6 (Curran et al. 2011a). Hydroxylated PCBs can cross the placenta (Meerts et al. 2002) and mimic the action of thyroid hormone (Giera et al. 2011, Darras et al. 2008). Recent work has implicated CYP1A1 in metabolism of non-coplanar PCB congeners (Wadzinski et al. 2014, Gauger et al. 2007) to thyromimetic forms. Both 4-OH-PCB 107 (Berghuis et al. 2013) and OH-PCB 106 (Haijima et al. 2017) cause motor deficits, so it will be important to look into differential metabolism of parent congeners in the susceptible and resistant lines of mice.

4.1 Conclusions

Our data provides evidence that developmental exposure to PCBs lead to motor deficits consistent with adverse effects on cerebellar development and function as well as disruption of nigrostriatal pathways. The data presented here and our previous studies (Curran et al. 2011a and 2011b) provide strong evidence that allelic differences at the Ahr and Cyp1a2 loci affect developmental neurotoxicity to polychlorinated biphenyls. We have extended our previous studies on learning and memory deficits to demonstrate that high-affinity AhrbCyp1a2(−/−) knockout mice are uniquely susceptible to motor dysfunction following PCB exposure during gestation and lactation. By including corn oil-treated control mice of three genotypes in our studies, we also uncovered a previously unreported motor deficit in Cyp1a2(−/−) mice, regardless of Ahr genotype. Given known human variation in the aryl hydrocarbon receptor and CYP1A2, these studies support the use of our mouse model to explore developmental neurotoxicity of similar persistent organic pollutants and the role of CYP1A2 in normal brain development and function.

Highlights.

  • Genetic variation in the aryl hydrocarbon receptor and cytochrome P450 1A2 (CYP1A2) alter susceptibility to developmental PCB exposure.

  • PCB-treated high-affinity Ahrbmice had greater deficits in motor function tests associated with nigrostriatal pathways compared with poor-affinity Ahrdmice.

  • PCB-treated Cyp1a2(−/−) knockout mice were more susceptible to motor deficits on tests associated with cerebellar function.

  • Unexpectedly, corn oil-treated control Cyp1a2(−/−) knockout mice also had deficits on the rotarod test compared with Cyp1a2(−/−) wild type mice.

  • Our findings cannot rule out PCBs as a risk factor for Parkinson’s disease, but our data indicate more than one brain region required for normal motor function are affected by developmental PCB exposure.

Acknowledgments

This work was supported by the National Institutes of Health [R15ES020053, P20 GM103436], the National Science Foundation [RSF-034-07, DUE-STEP-096928], and the following grants from Northern Kentucky University: Faculty Development Project Grants, College of Arts & Sciences Collaborative Faculty-Student Award, Center for Integrative Natural Science and Mathematics Research Grants, and Dorothy Westerman Herrmann funds. We thank Dr. David Thompson of Northern Kentucky University’s Department of Biological Sciences for assistance with the EROD assays, Collin Johnson for help with the Western blots, Mary Moran of Cincinnati Children’s Research Foundation for assistance with data analysis, and we acknowledge the generous donation of Cyp1a2(−/−) knockout mice from Dr. Daniel W. Nebert, University of Cincinnati Department of Environmental Health.

Footnotes

5. Conflicts of interest.

The authors report no conflicts of interest.

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References

  1. [ATSDR] Agency for Toxic Substances and Disease Registry. 2015 CERCLA National Priorities List of Hazardous Substances. U.S. Department of Health and Human Services, Public Health Service; Atlanta, GA: 2015. https://www.atsdr.cdc.gov/spl/ [Google Scholar]
  2. Akahoshi E, Yoshimura S, Uruno S, Ishihara-Sugano M. Effect of dioxins on regulation of tyrosine hydroxylase gene expression by aryl hydrocarbon receptor: a neurotoxicology study. Environ Health Glob Access Sci Source. 2009;8:24. doi: 10.1186/1476-069X-8-24. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Akahoshi E, Yoshimura S, Uruno S, Itoh S, Ishihara-Sugano M. Tyrosine hydroxylase assay: a bioassay for aryl hydrocarbon receptor-active compounds based on tyrosine hydroxylase promoter activation. Toxicol Mech Methods. 2012;22:458–460. doi: 10.3109/15376516.2012.668574. [DOI] [PubMed] [Google Scholar]
  4. Anezaki K, Kannan N, Nakano T. Polychlorinated biphenyl contamination of paints containing polycyclic- and Naphthol AS-type pigments. Environ Sci Pollut Res Int. 2015;22(19):14478–88. doi: 10.1007/s11356-014-2985-6. [DOI] [PubMed] [Google Scholar]
  5. Bányiová K, Černá M, Mikeš O, Komprdová K, Sharma A, Gyalpo T, Čupr P, Scheringer M. Long-term time trends in human intake of POPs in the Czech Republic indicate a need for continuous monitoring. Environ Int. 2017;108:1–10. doi: 10.1016/j.envint.2017.07.008. [DOI] [PubMed] [Google Scholar]
  6. Bemis JC, Seegal RF. PCB-induced inhibition of the vesicular monoamine transporter predicts reductions in synaptosomal dopamine content. Toxicol Sci Off J Soc Toxicol. 2004;80:288–295. doi: 10.1093/toxsci/kfh153. [DOI] [PubMed] [Google Scholar]
  7. Berghuis SA, Soechitram SD, Hitzert MM, Sauer PJJ, Bos AF. Prenatal exposure to polychlorinated biphenyls and their hydroxylated metabolites is associated with motor development of three-month-old infants. NeuroToxicology. 2013;38:124–130. doi: 10.1016/j.neuro.2013.07.003. [DOI] [PubMed] [Google Scholar]
  8. Boucher O, Muckle G, Ayotte P, Dewailly E, Jacobson SW, Jacobson JL. Altered fine motor function at school age in Inuit children exposed to PCBs, methylmercury, and lead. Environ Int. 2016;95:144–151. doi: 10.1016/j.envint.2016.08.010. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Brown EG. Mixed anionic detergent/aliphatic alcohol-polyacrylamide gel electrophoresis alters the separation of proteins relative to conventional sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Anal Biochem. 1988;174:337–348. doi: 10.1016/0003-2697(88)90555-6. [DOI] [PubMed] [Google Scholar]
  10. Caudle WM, Richardson JR, Delea KC, Guillot TS, Wang M, Pennell KD, et al. Polychlorinated biphenyl-induced reduction of dopamine transporter expression as a precursor to Parkinson's disease-associated dopamine toxicity. Toxicol Sci. 2006;92:490–499. doi: 10.1093/toxsci/kfl018. [DOI] [PubMed] [Google Scholar]
  11. Cauli O, Piedrafita B, Llansola M, Felipo V. Gender differential effects of developmental exposure to methyl-mercury, polychlorinated biphenyls 126 or 153, or its combinations on motor activity and coordination. Toxicology. 2013;311(1–2):61–8. doi: 10.1016/j.tox.2012.11.016. [DOI] [PubMed] [Google Scholar]
  12. Chishti MA, Fisher JP, Seegal RF. Aroclors 1254 and 1260 reduce dopamine concentrations in rat striatal slices. Neurotoxicology. 1996;17:653–660. [PubMed] [Google Scholar]
  13. Costabeber I, Dos Santos JS, Xavier AA, Weber J, Leaes FL, Junior SB, et al. Levels of polychlorinated biphenyls (PCBs) in meat and meat products from the state of Rio Grande do Sul, Brazil. Food Chem Toxicol. 2006;44:1–7. doi: 10.1016/j.fct.2005.01.005. [DOI] [PubMed] [Google Scholar]
  14. Curran CP, Vorhees CV, Williams MT, Genter MB, Nebert DW. In utero and lactational exposure to a complex mixture of polychlorinated biphenyls: toxicity in pups dependent on the Cyp1a2 and Ahr genotypes. Toxicol Sci. 2011a;119:189–208. doi: 10.1093/toxsci/kfq314. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Curran CP, Nebert DW, Genter MB, Patel KV, Schafer TL, Skelton MR, Williams MT, Vorhees CV. In Utero and Lactational Exposure to PCBs in Mice: Adult offspring show altered learning and memory depending on Cyp1a2 and Ahr genotypes. Enironv Health Perspect. 2011b;119(9):1286–93. doi: 10.1289/ehp.1002965. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Curran CP, Altenhofen E, Ashworth AA, Brown A, Curran MA, Evans A, Floyd R, Fowler JP, Garber H, Hays B, Kamau-Cheggeh C, Kraemer S, Lang AL, Mynhier A, Samuels A, Strohamier C. AhrdCyp1a2(−/−) mice show increased susceptibility to PCB-induced developmental neurotoxicity. Neurotoxicology. 2012;33(6):1436–42. doi: 10.1016/j.neuro.2012.08.005. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Darras VM. Endocrine disrupting polyhalogenated organic pollutants interfere with thyroid hormone signalling in the developing brain. The Cerebellum. 2008:26–37. doi: 10.1007/s12311-008-0004-5. [DOI] [PubMed] [Google Scholar]
  18. Denison MS, Faber SC. And Now for Something Completely Different: Diversity in Ligand-Dependent Activation of Ah Receptor Responses. Current Opinion in Toxicology. 2017;2:124–131. doi: 10.1016/j.cotox.2017.01.006. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Dingemans MML, Kock M, van den Berg M. Mechanisms of action point towards combined PBDE/NDL-PCB risk assessment. Toxicol Sci. 2016;153:215–224. doi: 10.1093/toxsci/kfw129. [DOI] [PubMed] [Google Scholar]
  20. Doull J. The “Red Book” and other risk assessment milestones. Hum Ecol Risk Assess. 2003;9:1229–1238. [Google Scholar]
  21. Dragin N, Dalton TP, Miller ML, Shertzer HG, Nebert DW. For dioxin-induced birth defects, mouse or human CYP1A2 in maternal liver protects whereas mouse CYP1A1 and CYP1B1 are inconsequential. J Biol Chem. 2006;281:18591–18600. doi: 10.1074/jbc.M601159200. [DOI] [PubMed] [Google Scholar]
  22. Fernagut PO, Chalon S, Diguet E, Guilloteau D, Tison F, Jaber M. Motor behaviour deficits and their histopathological and functional correlates in the nigrostriatal system of dopamine transporter knockout mice. Neuroscience. 2003;116:1123–1130. doi: 10.1016/s0306-4522(02)00778-9. [DOI] [PubMed] [Google Scholar]
  23. Fleming SM, Salcedo J, Fernagut PO, Rockenstein E, Masliah E, Levine MS, Chesselet MF. Early and progressive sensorimotor anomalies in mice overexpressing wild-type human alpha-synuclein. J Neurosci Off J Soc Neurosci. 2004;24:9434–9440. doi: 10.1523/JNEUROSCI.3080-04.2004. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Gafni J, Wong PW, Pessah IN. Non-coplanar 2,2’,3,5’,6-pentachlorobiphenyl (PCB 95) amplifies ionotropic glutamate receptor signaling in embryonic cerebellar granule neurons by a mechanism involving ryanodine receptors. Toxicol Sci Off J Soc Toxicol. 2004;77:72–82. doi: 10.1093/toxsci/kfh004. [DOI] [PubMed] [Google Scholar]
  25. Gauger KJ, Giera S, Sharlin DS, Bansal R, Iannacone E, Zoeller RT. Polychlorinated biphenyls 105 and 118 form thyroid hormone receptor agonists after cytochrome P4501A1 activation in rat pituitary GH3 cells. Environ Health Perspect. 2007;115:1623–1630. doi: 10.1289/ehp.10328. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Giera S, Bansal R, Ortiz-Toro TM, Taub DG, Zoeller RT. Individual polychlorinated biphenyl (PCB) congeners produce tissue- and gene-specific effects on thyroid hormone signaling during development. Endocrinology. 2011;152:2909–2919. doi: 10.1210/en.2010-1490. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Gomara B, Bordajandi LR, Fernandez MA, Herrero L, Abad E, Abalos M, et al. Levels and trends of polychlorinated dibenzo-p-dioxins/furans (PCDD/Fs) and dioxin-like polychlorinated biphenyls (PCBs) in Spanish commercial fish and shellfish products, 1995–2003. J Agric Food Chem. 2005;53:8406–8413. doi: 10.1021/jf050835z. [DOI] [PubMed] [Google Scholar]
  28. Haijima A, Lesmana R, Shimokawa N, Amano I, Takatsuru Y, Koibuchi N. Differential neurotoxic effects of in utero and lactational exposure to hydroxylated polychlorinated biphenyl (OH-PCB 106) on spontaneous locomotor activity and motor coordination in young adult male mice. J Toxicol Sci. 2017;42:407–416. doi: 10.2131/jts.42.407. [DOI] [PubMed] [Google Scholar]
  29. Hu D, Hornbuckle KC. Inadvertent polychlorinated biphenyls in commercial paint pigments. Environ Sci Technol. 2010;44(8):2822–7. doi: 10.1021/es902413k. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Iba MM, Storch A, Ghosal A, Bennett S, Reuhl KR, Lowndes HE. Constitutive and inducible levels of CYP1A1 and CYP1A2 in rat cerebral cortex and cerebellum. Arch Toxicol. 2003;77:547–554. doi: 10.1007/s00204-003-0488-1. [DOI] [PubMed] [Google Scholar]
  31. Jacobson JL, Jacobson SW. Prenatal exposure to polychlorinated biphenyls and attention at school age. J Pediatr. 2003;143:780–788. doi: 10.1067/S0022-3476(03)00577-8. [DOI] [PubMed] [Google Scholar]
  32. Korkalainen M, Linden J, Tuomisto J, Pohjanvirta R. Effect of TCDD on mRNA expression of genes encoding bHLH/PAS proteins in rat hypothalamus. Toxicology. 2005;208:1–11. doi: 10.1016/j.tox.2004.11.003. [DOI] [PubMed] [Google Scholar]
  33. Langer P, Kocan A, Tajtakova M, Petrik J, Chovancova J, Drobna B, et al. Fish from industrially polluted freshwater as the main source of organochlorinated pollutants and increased frequency of thyroid disorders and dysglycemia. Chemosphere. 2007;67:S379–S385. doi: 10.1016/j.chemosphere.2006.05.132. [DOI] [PubMed] [Google Scholar]
  34. Lee DW, Notter SA, Thiruchelvam M, Dever DP, Fitzpatrick R, Kostyniak PJ, Cory-Slechta DA, Opanashuk LA. Subchronic polychlorinated biphenyl (Aroclor 1254) exposure produces oxidative damage and neuronal death of ventral midbrain dopaminergic systems. Toxicol Sci. 2012 Feb;125(2):496–508. doi: 10.1093/toxsci/kfr313. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Lein PJ, Yang D, Bachstetter AD, Tilson HA, Harry GJ, Mervis RF, Kodavanti PR. Ontogenetic alterations in molecular and structural correlates of dendritic growth after developmental exposure to polychlorinated biphenyls. Environ Health Perspect. 2007;115(4):556–63. doi: 10.1289/ehp.9773. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Llansola M, Piedrafita B, Rodrigo R, Montoliu C, Felipo V. Polychlorinated biphenyls PCB 153 and PCB 126 impair the glutamate-nitric oxide-cGMP pathway in cerebellar neurons in culture by different mechanisms. Neurotox Res. 2009;16:97–105. doi: 10.1007/s12640-009-9055-8. https://doi.org/10.1007/s12640-009-9055-8. [DOI] [PubMed] [Google Scholar]
  37. Marek RF, Thorne PS, DeWall J, Hornbuckle KC. Variability in PCB and OH-PCB serum levels in children and their mothers in urban and rural U.S. communities. Environ Sci Technol. 2014;48(22):13459–67. doi: 10.1021/es502490w. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Malisch R, Kotz A. Dioxins and PCBs in feed and food--review from European perspective. Sci Total Environ. 2014;491–492:2–10. doi: 10.1016/j.scitotenv.2014.03.022. [DOI] [PubMed] [Google Scholar]
  39. Marek RF, Thorne PS, Herkert NJ, Awad AM, Hornbuckle KC. Airborne PCBs and OH-PCBs Inside and Outside Urban and Rural U.S. Schools. Environ Sci Technol. 2017;51(14):7853–7860. doi: 10.1021/acs.est.7b01910. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Matsuura K, Kabuto H, Makino H, Ogawa N. Pole test is a useful method for evaluating the mouse movement disorder caused by striatal dopamine depletion. J Neurosci Methods. 1997;73:45–48. doi: 10.1016/s0165-0270(96)02211-x. [DOI] [PubMed] [Google Scholar]
  41. Meerts IATM, Assink Y, Cenijn PH, Van Den Berg JHJ, Weijers BM, Bergman A, Koeman JH, Brouwer A. Placental transfer of a hydroxylated polychlorinated biphenyl and effects on fetal and maternal thyroid hormone homeostasis in the rat. Toxicol Sci Off J Soc Toxicol. 2002;68:361–371. doi: 10.1093/toxsci/68.2.361. [DOI] [PubMed] [Google Scholar]
  42. Miller MM, Sprowles JLN, Voeller JN, Meyer AE, Sable HJK. Cocaine sensitization in adult Long-Evans rats perinatally exposed to polychlorinated biphenyls. Neurotoxicol Teratol. 2017;62:34–41. doi: 10.1016/j.ntt.2017.04.005. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Mulero-Navarro S, Santiago-Josefat B, Pozo-Guisado E, Merino JM, Fernandez-Salguero PM. Down-regulation of CYP1A2 induction during the maturation of mouse cerebellar granule cells in culture: role of nitric oxide accumulation. Eur J Neurosci. 2003;18:2265–2272. doi: 10.1046/j.1460-9568.2003.02972.x. [DOI] [PubMed] [Google Scholar]
  44. Nadler JJ, Zou F, Huang H, Moy SS, Lauder J, Crawley JN, Threadgill DW, Wright FA, Magnuson TR. Large-scale gene expression differences across brain regions and inbred strains correlate with a behavioral phenotype. Genetics. 2006;174:1229–1236. doi: 10.1534/genetics.106.061481. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Nebert DW, Dalton TP. The role of cytochrome P450 enzymes in endogenous signalling pathways and environmental carcinogenesis. Nat Rev Cancer. 2006;6:947–960. doi: 10.1038/nrc2015. [DOI] [PubMed] [Google Scholar]
  46. Nebert DW, Wikvall K, Miller WL. Human cytochromes P450 in health and disease. Philos Trans R Soc Lond B Biol Sci. 2013;368(1612):20120431. doi: 10.1098/rstb.2012.0431. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Nebert DW. Aryl hydrocarbon receptor (AHR): “pioneer member” of the basic-helix/loop/helix per-Arnt-sim (bHLH/PAS) family of “sensors” of foreign and endogenous signals. Prog Lipid Res. 2017;67:38–57. doi: 10.1016/j.plipres.2017.06.001. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Nguon K, Baxter MG, Sajdel-Sulkowska EM. Perinatal exposure to polychlorinated biphenyls differentially affects cerebellar development and motor functions in male and female rat neonates. Cerebellum. 2005;4:112–122. doi: 10.1080/14734220510007860. [DOI] [PubMed] [Google Scholar]
  49. Novotna B, Topinka J, Solansky I, Chvatalova I, Lnenickova Z, Sram RJ. Impact of air pollution and genotype variability on DNA damage in Prague policemen. Toxicol Lett. 2007;172:37–47. doi: 10.1016/j.toxlet.2007.05.013. [DOI] [PubMed] [Google Scholar]
  50. Ogawa N, Hirose Y, Ohara S, Ono T, Watanabe Y. A simple quantitative bradykinesia test in MPTP-treated mice. Res Commun Chem Pathol Pharmacol. 1985;50:435–441. [PubMed] [Google Scholar]
  51. Ogawa N, Mizukawa K, Hirose Y, Kajita S, Ohara S, Watanabe Y. MPTP-induced parkinsonian model in mice: biochemistry, pharmacology and behavior. Eur Neurol. 1987;26(Suppl 1):16–23. doi: 10.1159/000116351. [DOI] [PubMed] [Google Scholar]
  52. Pessah IN, Cherednichenko G, Lein PJ. Minding the calcium store: Ryanodine receptor activation as a convergent mechanism of PCB toxicity. Pharmacol Ther. 2010;125:260–285. doi: 10.1016/j.pharmthera.2009.10.009. [DOI] [PMC free article] [PubMed] [Google Scholar]
  53. Petersen MS, Halling J, Bech S, Wermuth L, Weihe P, Nielsen F, et al. Impact of dietary exposure to food contaminants on the risk of Parkinson's disease. Neurotoxicology. 2008;29:584–590. doi: 10.1016/j.neuro.2008.03.001. [DOI] [PubMed] [Google Scholar]
  54. Quinn CL, Wania F, Czub G, Breivik K. Investigating intergenerational differences in human PCB exposure due to variable emissions and reproductive behaviors. Environ Health Perspect. 2011;119:641–646. doi: 10.1289/ehp.1002415. [DOI] [PMC free article] [PubMed] [Google Scholar]
  55. Roegge CS, Schantz SL. Motor function following developmental exposure to PCBS and/or MEHG. Neurotoxicol Teratol. 2006;28:260–277. doi: 10.1016/j.ntt.2005.12.009. [DOI] [PubMed] [Google Scholar]
  56. Ross G. The public health implications of polychlorinated biphenyls (PCBs) in the environment. Ecotoxicol Environ Saf. 2004;59:275–291. doi: 10.1016/j.ecoenv.2004.06.003. [DOI] [PubMed] [Google Scholar]
  57. Schallert T. Aging-dependent emergence of sensorimotor dysfunction in rats recovered from dopamine depletion sustained early in life. Ann NY Acad Sci. 1988;515:108–120. doi: 10.1111/j.1749-6632.1988.tb32972.x. [DOI] [PubMed] [Google Scholar]
  58. Schantz SL, Widholm JJ, Rice DC. Effects of PCB exposure on neuropsychological function in children. Environ Health Perspect. 2003;111:357–576. doi: 10.1289/ehp.5461. [DOI] [PMC free article] [PubMed] [Google Scholar]
  59. Schultheis PJ, Fleming SM, Clippinger AK, Lewis J, Tsunemi T, Giasson B, Dickson DW, Mazzulli JR, Bardgett ME, Haik KL, Ekhator O, Chava AK, Howard J, Gannon M, Hoffman E, Chen Y, Prasad V, Linn SC, Tamargo RJ, Westbroek W, Sidransky E, Krainc D, Shull GE. Atp13a2-deficient mice exhibit neuronal ceroid lipofuscinosis, limited α-synuclein accumulation and age-dependent sensorimotor deficits. Hum Mol Genet. 2013;22:2067–2082. doi: 10.1093/hmg/ddt057. [DOI] [PMC free article] [PubMed] [Google Scholar]
  60. Sedelis M, Schwarting RK, Huston JP. Behavioral phenotyping of the MPTP mouse model of Parkinson’s disease. Behav Brain Res. 2001;125:109–125. doi: 10.1016/s0166-4328(01)00309-6. [DOI] [PubMed] [Google Scholar]
  61. Seegal RF, Brosch KO, Bush B. Polychlorinated biphenyls produce regional alterations of dopamine metabolism in rat brain. Toxicol Lett. 1986;30:197–202. doi: 10.1016/0378-4274(86)90103-7. [DOI] [PubMed] [Google Scholar]
  62. Seegal RF, Brosch KO, Okoniewski RJ. Effects of in utero and lactational exposure of the laboratory rat to 2,4,2',4'- and 3,4,3',4'-tetrachlorobiphenyl on dopamine function. Toxicol Appl Pharmacol. 1997;146:95–103. doi: 10.1006/taap.1997.8226. [DOI] [PubMed] [Google Scholar]
  63. Seegal RF, Brosch KO, Okoniewski RJ. Coplanar PCB congeners increase uterine weight and frontal cortical dopamine in the developing rat: implications for developmental neurotoxicity. Toxicol Sci. 2005;86:125–131. doi: 10.1093/toxsci/kfi174. [DOI] [PubMed] [Google Scholar]
  64. Seegal RF, Bush B, Brosch KO. Decreases in dopamine concentrations in adult, non-human primate brain persist following removal from polychlorinated biphenyls. Toxicology. 1994;86:71–87. doi: 10.1016/0300-483x(94)90054-x. [DOI] [PubMed] [Google Scholar]
  65. Steenland K, Hein MJ, Cassinelli RT, Prince MM, Nilsen NB, Whelan EA, et al. Polychlorinated biphenyls and neurodegenerative disease mortality in an occupational cohort. Epidemiology. 2006;17:8–13. doi: 10.1097/01.ede.0000190707.51536.2b. [DOI] [PubMed] [Google Scholar]
  66. Stewart P, Reihman J, Lonky E, Darvill T, Pagano J. Prenatal PCB exposure and neonatal behavioral assessment scale (NBAS) performance. Neurotoxicol Teratol. 2000;22:21–29. doi: 10.1016/s0892-0362(99)00056-2. [DOI] [PubMed] [Google Scholar]
  67. Thompson ED, Burwinkel KE, Chava AK, Notch EG, Mayer GD. Activity of Phase I and Phase II enzymes of the benzo[a]pyrene transformation pathway in zebrafish (Danio rerio) following waterborne exposure to arsenite. Comp Biochem Physiol Toxicol Pharmacol CBP. 2010;152:371–378. doi: 10.1016/j.cbpc.2010.06.004. [DOI] [PubMed] [Google Scholar]
  68. Tsuchiya Y, Nakai S, Nakamura K, Hayashi K, Nakanishi J, Yamamoto M. Effects of dietary habits and CYP1A1 polymorphisms on blood dioxin concentrations in Japanese men. Chemosphere. 2003;52:213–219. doi: 10.1016/S0045-6535(03)00110-3. [DOI] [PubMed] [Google Scholar]
  69. Van den berg BM, Birnbaum LS, Denison M, De VM, Farland W, Feeley M, et al. The 2005 World Health Organization reevaluation of human and Mammalian toxic equivalency factors for dioxins and dioxin-like compounds. Toxicol Sci. 2006;93:223–241. doi: 10.1093/toxsci/kfl055. [DOI] [PMC free article] [PubMed] [Google Scholar]
  70. van Duursen MB, Sanderson JT, Van den BM. Cytochrome P450–1A1 and 1B1 in human blood lymphocytes are not suitable as biomarkers of exposure to dioxin-like compounds: polymorphisms and interindividual variation in expression and inducibility. Toxicol Sci. 2005;85:703–712. doi: 10.1093/toxsci/kfi089. [DOI] [PubMed] [Google Scholar]
  71. Vreugdenhil HJ, Lanting CI, Mulder PG, Boersma ER, Weisglas-Kuperus N. Effects of prenatal PCB and dioxin background exposure on cognitive and motor abilities in Dutch children at school age. J Pediatr. 2002;140:48–56. doi: 10.1067/mpd.2002.119625. [DOI] [PubMed] [Google Scholar]
  72. Wadzinski TL, Geromini K, McKinley Brewer J, Bansal R, Abdelouahab N, Langlois MF, Takser L, Zoeller RT. Endocrine disruption in human placenta: expression of the dioxin-inducible enzyme, CYP1A1, is correlated with that of thyroid hormone-regulated genes. J Clin Endocrinol Metab. 2014;99:E2735–2743. doi: 10.1210/jc.2014-2629. [DOI] [PMC free article] [PubMed] [Google Scholar]
  73. [WHO] World Health Organization. Polychlorinated Biphenyls: Human Health Aspects. World Health Organization; Geneva, Switzerland: 2003. [Google Scholar]
  74. Widholm JJ, Seo BW, Strupp BJ, Seegal RF, Schantz SL. Effects of perinatal exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin on spatial and visual reversal learning in rats. Neurotoxicol Teratol. 2003;25:459–471. doi: 10.1016/s0892-0362(03)00014-x. [DOI] [PubMed] [Google Scholar]
  75. Wilhelm M, Ranft U, Krämer U, Wittsiepe J, Lemm F, Fürst P, Eberwein G, Winneke G. Lack of neurodevelopmental adversity by prenatal exposure of infants to current lowered PCB levels: comparison of two German birth cohort studies. J Toxicol Environ Health A. 2008;71:700–702. doi: 10.1080/15287390801984904. [DOI] [PubMed] [Google Scholar]
  76. Yadav S, Johri A, Dhawan A, Seth PK, Parmar D. Regional specificity in deltamethrin induced cytochrome P450 expression in rat brain. Toxicol Appl Pharmacol. 2006;217:15–24. doi: 10.1016/j.taap.2006.07.008. [DOI] [PubMed] [Google Scholar]

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