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Developmental Thyroid Hormone Insufficiency Induces Cortical Brain Malformation and Learning Impairments: A Cross-Fostering Study
OShaughnessy, K., Pat Kosian, J. Ford, W. Oshiro, S. Degitz, AND M. Gilbert. Developmental Thyroid Hormone Insufficiency Induces Cortical Brain Malformation and Learning Impairments: A Cross-Fostering Study. TOXICOLOGICAL SCIENCES. Society of Toxicology, RESTON, VA, 163(1):101-115, (2018).
The risk of thyroid disrupting chemicals is a high priority of concern when considering children’s health, as normal brain development is dependent on thyroid hormones (THs). What is less understood is how the timing of a maternal xenobiotic exposure can differentially affect the thyroid axis in the fetus and/or child, and how TH insufficiency across discrete points in development may lead to differential adverse outcomes of the central nervous system. To begin to elucidate these issues we performed a cross-fostering experiment in rats to parse the effects of maternal TH disruption during pregnancy, during lactation, or across neurodevelopment. First, pregnant dams were dosed with the goitrogen, propylthiouracial (PTU, 3 ppm or 0 ppm control), from gestational day (GD) 6 until postnatal day (PN) 21. On PN2, half of the pups from each litter were fostered to a dam of the opposite dose group. This resulted in four treatment conditions: pups that received PTU only during gestation, pups that received PTU primarily during lactation, those that received PTU across neurodevelopment, and pups that were control. Next, PTU and TH concentrations were measured in the serum of both dams and pups across developmental time. These measurements, coupled with traditional apical endpoints like body weight changes, provided an integration of chemical dosimetry and resulting TH effects in each exposure condition. This resulted in a comprehensive estimate of exposure kinetics in both mother and offspring. To begin to address the neurodevelopmental consequences of each exposure, the neonatal brain was screened for the presence of a heterotopia. The heterotopia represents a morphological defect that we have previously identified in the brains of developmentally hypothyroid animals. However, how the timing of TH insufficiency relates to heterotopia penetrance and severity is unknown. Our results demonstrate that PTU treatment during gestation is both sufficient and necessary for heterotopia formation in the neonatal brain. Pups that were exposed to PTU only during lactation did not form this birth defect. Furthermore, results identified that heterotopia formation is not sex dependent, and is a permanent defect persisting to PN300 in the rat. Given this morphological data, we next investigated whether learning impairments, a behavioral phenotype previously implicated in hypothyroid animals, was also associated with a specific exposure scenario. Interestingly, results show that only male animals that were dosed with PTU across neurodevelopment exhibited any alterations in behavior. This is reminiscent of the male-bias in neurodevelopmental disorders in humans, and suggests that this neurobehavioral outcome may be sex dependent. These results also suggest a dissociation between our structural and behavioral indices of TH disruption, and illustrate how differing windows of susceptibility may confer distinct phenotypes. This work will be used to further our quantitative adverse outcome pathway development for thyroid and mammalian brain function and forms the basis for further investigations to refine the window for mechanistic studies of heterotopia formation.
Thyroid hormones (TH) are essential for brain development, but animal models of well-defined and sensitive downstream apical neurotoxic outcomes associated with developmental TH disruption are lacking. A structural anomaly, a cortical heterotopia, in the brains of hypothyroid rats treated in utero with the TH synthesis inhibitor propylthiouracil (PTU) has been previously described. This study aims to clarify the exposure and internal dosimetry associated with its formation, define the window of exposure required for its induction, and more fully characterize its physical parameters. Pregnant LE rats (n=32) were exposed to PTU (0 or 3ppm) through the drinking water from gestational day (GD) 6 until postnatal day (PN) 14. On PN2 a subset of pups was cross-fostered to a dam of the opposite treatment, in order to create four conditions: pups that were exposed to PTU primarily prenatally, primarily postnatally, during both periods, or not at all (control). Both PTU and TH concentrations were characterized in both the mother and offspring over time, in order to capture the dynamics of a developmental xenobiotic exposure. Additionally, the brains of offspring were examined for heterotopia presence and severity, and adult littermates were assessed for memory impairments using a trace fear conditioning paradigm. Heterotopia were reliably observed under conditions of prenatal exposure, but heterotopia volume increased in animals that experienced a more sustained exposure from GD6-PN14. This defect is permanent persisting in animals assessed ~PN300. Heterotopia volume did not change significantly between PN14 and adulthood, nor was it sex-dependent. In contrast, behavioral impairments were limited to male offspring from the treatment group exposed to PTU during both the gestational and postnatal periods. This suggests a distinct TH-dependent etiology for both morphological and functional phenotypes, and illustrates how timing of hypothyroxinemia can induce abnormal brain structure and function. These findings inform the design of studies aimed at identifying biomarkers of TH-action associated with heterotopia formation, improving our ability to assess neurotoxicity of endocrine disrupting chemicals. These findings further underscore the need to examine hormonal status in the pre and postnatal period and to target investigations to appropriate apical endpoints.