NIEHS-funded research pursues thyroid findings.

Clinical studies and animal experiments have established the essential role of thyroid hormone for normal brain development. A lack of thyroid hormone in pregnancy can result in congenital hypothyroidism, which causes moderate to severe mental retardation when untreated. Subtle thyroid hormone deficits in pregnant rats disrupt the migration of neurons in the fetal cortex and hippocampus. Several epidemiological studies have shown that mothers with subtle thyroid hormone deficiencies give birth to children with lower IQs than mothers with normal thyroid hormone levels. Exposure to certain environmental contaminants, including polychlorinated biphenyls (PCBs), organochlorine pesticides, and phthalates, has been reported to interfere with the production, transportation, and metabolism of thyroid hormone by a variety of mechanisms. Yet, despite the many puzzle pieces already in place, questions remain regarding whether--and to what extent--such effects by environmental contaminants include brain development problems.

The thyroid secretes thyroxine ([T.sub.4]), which is converted in tissues to triiodothyronine ([T.sub.3]); together, these two products generally are known collectively as "thyroid hormone." The activity of the thyroid gland is predominantly regulated by the pituitary glycoprotein hormone known as thyroid-stimulating hormone (TSH). Thyroid hormone exerts a negative feedback effect on pituitary secretion of TSH, so that when [T.sub.4] concentrations drop, TSH concentrations increase, thus keeping [T.sub.3] levels stable. Thyroid hormones exert their action at a nuclear level in the brain by regulating the transcription of thyroid hormone-responsive genes. This process is initiated when [T.sub.3] binds to thyroid hormone receptors. Perturbations in this system and the possible consequences are the topic of several NIEHS-funded studies.

Intramural and extramural research approaches range from animal studies of brain effects to studying U.S. populations that are exposed to relatively high levels of PCBs and other contaminants in comparison to the general population. In addition, a research group within NIEHS is exploring a novel mechanism by which thyroid hormone may affect brain development.

PCBs: A Target of Research

A number of NIEHS-funded researchers are focusing on the effects of PCBs. PCBs are a well-studied class of industrial compounds that consist of paired phenyl rings with various degrees of chlorination. They are now ubiquitous and persistent environmental contaminants routinely found in samples of human and animal tissues. Although adverse effects have been linked to PCB exposure, it has not proved possible to tease out the mechanism of action for most of these effects.

Several studies have shown that exposure to this group of contaminants has been linked in humans with neurological effects similar to those associated with thyroid hormone deficiencies. In addition, there is strong evidence that exposure to PCBs can reduce circulating levels of thyroid hormone in animals (as reported by Ellen S. Goldey and Kevin M. Crofton in the September 1998 issue of Toxicological Sciences) and mimic thyroid hormone's effect on gene expression in the brain (as reported by R. Thomas Zoeller and colleagues in the January 2000 issue of Endocrinology). However, this evidence comes from studies using much larger PCB doses than those found in nature, says David Armstrong, a senior investigator in the NIEHS Membrane Signaling Group.

An association between PCB exposure and thyroid hormone levels in humans has been cited in multiple papers, including reports by Lars Hagmar and colleagues in the 10 April 2001 issue of International Archives of Occupational and Environmental Health, M. Sala and colleagues in the March 2001 issue of Occupational & Environmental Medicine, and Victoria Persky and colleagues in the December 2001 issue of EHP. PCBs can enhance liver metabolism of thyroid hormone, which increases biliary excretion, or interfere with serum proteins that bind and transport thyroid hormone. Both effects may also reduce circulating levels of thyroid hormone.

Laboratory Studies

Zoeller, an endocrinologist at the University of Massachusetts Amherst, has, with his colleagues, identified a number of genes whose expression in the rat brain can be used as biomarkers of thyroid hormone action. Zoeller is particularly interested in the effect of thyroid hormone on early brain development, when the fetal brain gets thyroid hormone from the mother. By focusing on genes that are expressed before the onset of fetal thyroid function, Zoeller's group can study the effects of maternal exposure to thyroid-disruptive agents.

PCBs seem to mimic thyroid hormone's effects on expression of these genes--interfering with thyroid hormone signaling without necessarily inhibiting the function of the thyroid gland. However, Zoeller's group has yet to identify any PCB congeners that bind with the thyroid hormone receptor. Such binding is the most obvious mechanism for PCBs' mimicry of thyroid hormone action in the brain. Other contaminants that can competitively bind to thyroid hormone receptors include bisphenol A (as reported by Kenji Moriyama and colleagues in the November 2002 issue of the Journal of Clinical Endocrinology & Metabolism) and halogenated bisphenol A derivatives (as reported by Shigeyuki Kitamura and colleagues in the 26 April 2002 issue of Biochemical and Biophysical Research Communications).

Lisa Opanashuk, an assistant professor of environmental medicine at the University of Rochester Medical Center in New York, has just begun working with in vitro cerebellar brain cell lines to look for indicators of contaminant action. While Zoeller started by studying the mechanisms of thyroid hormone action in the brain and then evaluating contaminant effects in this light, Opanashuk is first looking at contaminant action and then plans to eventually link these end points to thyroid hormone effects.

Armstrong and the NIEHS Membrane Signaling Group are taking yet another approach to studying thyroid hormone action in the brain by looking at basic mechanisms as opposed to toxicant action. Armstrong's team is pursuing tantalizing evidence that thyroid hormone receptors may also orchestrate nongenomic cellular functions in response to both thyroid hormone and environmental contaminants. In in vitro experiments using a rat pituitary cell line, Armstrong's team found that thyroid hormone stimulates potassium channel activity through a signal transduction cascade that involves PI3 kinase and Rac GTPase. Other investigators have already reported that both Rac and potassium channels are essential for normal neurite outgrowth in the developing brain. Hence, this discovery may provide a novel, nongenomic molecular explanation for how thyroid hormone system perturbations can affect brain development.

Epidemiological Studies

Studies that have attempted to relate PCB body burden to thyroid hormone levels have produced inconsistent results, as have studies aimed at linking PCB exposure to neurodevelopmental effects. As part of his investigations into the developmontal effects of organic contaminants, NIEHS epidemiologist Matthew Longnecker is exploring the reasons for these discrepancies.

Currently it's very difficult to compare studies because they use different measures of PCB exposure. To rectify this first barrier, Longnecker, together with other PCB investigators, has developed a uniform way to compare exposure levels so that data from all studies can be considered together. This method, reported in the January 2003 issue of EHP, focuses on the median level of PCB 153 in maternal serum. Longnecker and colleagues found that exposure in most studies overlapped substantially but that exposure levels determined in a study in the Faroe Islands, where people eat an especially high-PCB diet, were 3-4 times higher than the rest.

Epidemiologist Anne Sweeney and colleagues at Texas A&M University's School of Rural Public Health recently began a long-term prospective study of Asian-American children potentially exposed to high levels of PCBs and methylmercury through eating contaminated fish from Wisconsin's polluted Fox River. Researchers are currently recruiting reproductive-age couples with the goal of following 250 babies and parents from preconception through the children's growth.

Sweeney and colleagues will monitor changes in serum PCB and methylmercury concentrations and look for associations with perturbations in thyroid hormone status. The focus will be on specific aspects of cognitive, motor, and sensory function that are affected by PCBs and/or methylmercury in animal models and attributed to thyroid hormone disruption, but that have not been adequately assessed in human infants or children. These include auditory function, frontal lobe functions (such as working memory, planning, and response inhibition), and cerebellar functions (such as balance and coordination).

Lawrence Schell, an epidemiologist at the University at Albany School of Public Health, and colleagues are nearing completion of another population study to determine the effect of PCB exposure on thyroid hormone function in adolescents of the Akwesasne Mohawk Nation. Schell's team interviewed 270 mother-adolescent child pairs, collecting a diverse data set including information on maternal fish consumption; child diet; sociodemographic data; height, weight, and body composition measures; sexual maturation scale; and alcohol and cigarette use. Assessments of adolescent cognitive and behavioral characteristics were also conducted. Levels of total and free [T.sub.4], [T.sub.3], and TSH were then measured in the adolescents' blood.

Preliminary analysis of data from 117 adolescents in this study has found that the level of highly chlorinated congeners is significantly related to thyroid function, although it is important to note that the Akwesasne have been exposed to a large number of harmful contaminants through living on the St. Lawrence River adjacent to a number of hazardous waste sites. The researchers emphasize that because the PCB levels in the sample are very low, their results may pertain to many populations with low exposure. According to Schell, these results support the theory that PCB exposure at levels commonly found in human populations alters thyroid function.

Other Efforts

An important part of both Sweeney's and Schell's population studies is an intensive effort to inform and involve communities in the research. This effort to involve the public is an important part of the NIEHS approach, according to Jerrold Heindel, a program administrator in the NIEHS Division of Extramural Research and Training. Researchers try to involve the community by explaining their work and requesting comments and input from the people involved. In some cases, such as the Fox River study, community, outreach also involves showing people how to avoid PCB exposure, for example by avoiding certain fish and eating only skinned fillets.

The National Toxicology Program Center for the Evaluation of Risks to Human Reproduction is interested in developing tests to detect adverse reproductive and developmental effects resulting from chemical-induced thyroid dysfunction, says center director Michael Shelby. Such tests could be used to screen chemicals for thyroid hormone effects. Developmental toxicity tests do not routinely monitor thyroid status because there is no definitive, easily measurable biological end point whose consequences are well understood, he says.

Last September, the NIEHS co-sponsored an international conference on thyroid hormone and brain development. Meeting participants agreed that thyroid hormone action plays multiple roles in both early and late brain development, and biomarkers to detect subtle effects are elusive. But new studies, conceived to pinpoint these effects by using genetically modified animals, are expected to make great strides soon. Together, all of these advances mean that there soon may be screening tests developed to determine whether environmental contaminants affect thyroid hormone signaling in the developing brain. Such knowledge could lead to a reduction in associated learning deficiencies.