Alzheimer's disease (AD)-like pathology in aged monkeys after infantile exposure to environmental metal lead (Pb): evidence for a developmental origin and environmental link for AD.

The sporadic nature of Alzheimer's disease (AD) argues for an environmental link that may drive AD pathogenesis; however, the triggering factors and the period of their action are unknown. Recent studies in rodents have shown that exposure to lead (Pb) during brain development predetermined the expression and regulation of the amyloid precursor protein (APP) and its amyloidogenic beta-amyloid (Abeta) product in old age. Here, we report that the expression of AD-related genes [APP, BACE1 (beta-site APP cleaving enzyme 1)] as well as their transcriptional regulator (Sp1) were elevated in aged (23-year-old) monkeys exposed to Pb as infants. Furthermore, developmental exposure to Pb altered the levels, characteristics, and intracellular distribution of Abeta staining and amyloid plaques in the frontal association cortex. These latent effects were accompanied by a decrease in DNA methyltransferase activity and higher levels of oxidative damage to DNA, indicating that epigenetic imprinting in early life influenced the expression of AD-related genes and promoted DNA damage and pathogenesis. These data suggest that AD pathogenesis is influenced by early life exposures and argue for both an environmental trigger and a developmental origin of AD.

Co-localization and distribution of cerebral APP and SP1 and its relationship to amyloidogenesis.

Alzheimer's disease is characterized by amyloid-beta peptide (Abeta)-loaded plaques in the brain. Abeta is a cleavage fragment of amyloid-beta protein precursor (APP) and over production of APP may lead to amyloidogenesis. The regulatory region of the APP gene contains consensus sites recognized by the transcription factor, specificity protein 1 (SP1), which has been shown to be required for the regulation of APP and Abeta. To understand the role of SP1 in APP biogenesis, herein we have characterized the relative distribution and localization of SP1, APP, and Abeta in various brain regions of rodent and primate models using immunohistochemistry. We observed that overall distribution and cellular localization of SP1, APP, and Abeta are similar and neuronal in origin. Their distribution is abundant in various layers of neocortex, but restricted to the Purkinje cell layer of the cerebellum, and the pyramidal cell layer of hippocampus. These findings suggest that overproduction of Abeta in vivo may be associated with transcriptional pathways involving SP1 and the APP gene.

Meeting report: moving upstream-evaluating adverse upstream end points for improved risk assessment and decision-making.

BACKGROUND: Assessing adverse effects from environmental chemical exposure is integral to public health policies. Toxicology assays identifying early biological changes from chemical exposure are increasing our ability to evaluate links between early biological disturbances and subsequent overt downstream effects. A workshop was held to consider how the resulting data inform consideration of an "adverse effect" in the context of hazard identification and risk assessment. OBJECTIVES: Our objective here is to review what is known about the relationships between chemical exposure, early biological effects (upstream events), and later overt effects (downstream events) through three case studies (thyroid hormone disruption, antiandrogen effects, immune system disruption) and to consider how to evaluate hazard and risk when early biological effect data are available. DISCUSSION: Each case study presents data on the toxicity pathways linking early biological perturbations with downstream overt effects. Case studies also emphasize several factors that can influence risk of overt disease as a result from early biological perturbations, including background chemical exposures, underlying individual biological processes, and disease susceptibility. Certain effects resulting from exposure during periods of sensitivity may be irreversible. A chemical can act through multiple modes of action, resulting in similar or different overt effects. CONCLUSIONS: For certain classes of early perturbations, sufficient information on the disease process is known, so hazard and quantitative risk assessment can proceed using information on upstream biological perturbations. Upstream data will support improved approaches for considering developmental stage, background exposures, disease status, and other factors important to assessing hazard and risk for the whole population.

Overview of modifiers of methylmercury neurotoxicity: chemicals, nutrients, and the social environment.

It has been known for decades that methylmercury is a potent neurotoxicant, and that the developing brain is more susceptible to impairment as a result of methylmercury exposure than is the adult. Exposure to methylmercury is exclusively through consumption of fish and marine mammals. In recent years, the potential for protection against methylmercury toxicity by nutrients present in fish, particularly omega-3 fatty acids and selenium, has been explored in both epidemiological and experimental studies. There is evidence from several studies that fish consumption per se and methylmercury body burden act in opposition with regard to neuropsychological outcomes, whereas the evidence for a protective effect of specific nutrients is contradictory in both epidemiological and experimental studies published to date. The potential for methylmercury to interact with other chemicals present in marine food, particularly PCBs, has been explored in both animal models and human studies. Results may be both exposure- and endpoint-dependent. The Seychelles Islands study has explored the potential for the social environment to modify the effects of developmental methylmercury exposure. An understanding of the interactions of the multiple factors that determine the final behavioral outcome of exposure to methylmercury is crucial to risk assessment and risk management decisions.

Blood lead concentration and delayed puberty in girls.

BACKGROUND: Environmental lead exposure has been linked to alterations in growth and endocrine function. It is not known whether such exposure affects pubertal development. METHODS: We analyzed the relations between blood lead concentration and pubertal development among girls (defined as females 8 to 18 years of age) who were enrolled in a cross-sectional study (the third National Health and Nutrition Examination Survey) in which race was self-reported or proxy-reported: 600 were non-Hispanic white, 805 were non-Hispanic African-American, and 781 were Mexican-American girls. Puberty was measured on the basis of the age at menarche and Tanner stage for pubic-hair and breast development. RESULTS: Geometric mean lead concentrations were less than 3 microg per deciliter (0.144 micromol per liter) in all three groups. As compared with concentrations of 1 microg per deciliter (0.048 micromol per liter), lead concentrations of 3 microg per deciliter were associated with decreased height (P<0.001), after adjustment for age, race, and other factors, but not with body-mass index or weight. Blood lead concentrations of 3 microg per deciliter were associated with significant delays in breast and pubic-hair development in African-American and Mexican-American girls. The delays were most marked among African-American girls; in this group, the delays in reaching Tanner stages 2, 3, 4, and 5 associated with a lead concentration of 3 microg per deciliter as compared with 1 microg per deciliter were 3.8, 5.3, 5.8, and 2.1 months, respectively, for breast development and 4.0, 5.5, 6.0, and 2.2 months, respectively, for pubic-hair development; the associated delay in age at menarche was 3.6 months. In white girls, there were nonsignificant delays in all pubertal measures in association with a lead concentration of 3 microg per deciliter. CONCLUSIONS: These data suggest that environmental exposure to lead may delay growth and pubertal development in girls, although confirmation is warranted in prospective studies.

Evaluation of the U.S. EPA/OSWER preliminary remediation goal for perchlorate in groundwater: focus on exposure to nursing infants.

BACKGROUND: Perchlorate is a common contaminant of drinking water and food. It competes with iodide for uptake into the thyroid, thus interfering with thyroid hormone production. The U.S. Environmental Protection Agency's Office of Solid Waste and Emergency Response (OSWER) set a groundwater preliminary remediation goal (PRG) of 24.5 microg/L to prevent exposure of pregnant women that would affect the fetus. This does not account for the greater exposure that is possible in nursing infants or for the relative source contribution (RSC), a factor normally used to lower the PRG due to nonwater exposures. OBJECTIVES: Our goal was to assess whether the OSWER PRG protects infants against exposures from breast-feeding, and to evaluate the perchlorate RSC. METHODS: We used Monte Carlo analysis to simulate nursing infant exposures associated with the OSWER PRG when combined with background perchlorate. RESULTS: The PRG can lead to a 7-fold increase in breast milk concentration, causing 90% of nursing infants to exceed the reference dose (RfD) (average exceedance, 2.8-fold). Drinking-water perchlorate must be < 6.9 microg/L to keep the median, and < 1.3 microg/L to keep the 90th-percentile nursing infant exposure below the RfD. This is 3.6- to 19-fold below the PRG. Analysis of biomonitoring data suggests an RSC of 0.7 for pregnant women and of 0.2 for nursing infants. Recent data from the Centers for Disease Control and Prevention (CDC) suggest that the RfD itself needs to be reevaluated because of hormonal effects in the general population. CONCLUSIONS: The OSWER PRG for perchlorate can be improved by considering infant exposures, by incorporating an RSC, and by being responsive to any changes in the RfD resulting from the new CDC data.

Developmental delays and locomotor activity in the C57BL6/J mouse following neonatal exposure to the fully-brominated PBDE, decabromodiphenyl ether.

After several decades of commercial use, the flame retardant chemicals polybrominated diphenyl ethers (PBDEs) and their metabolites have become pervasive environmental contaminants with a global distribution. PBDEs have entered the food chain and increasing levels can be detected in the human body. Decabrominated diphenyl ether (decaBDE) is currently the most widely used of the PBDEs in the United States. Despite its widespread use, little is known about the health effects of decaBDE. The current study examined the effects of neonatal exposure to decaBDE in the inbred C57BL6/J mouse. Neonatal male and female mice were exposed to a daily oral dose of 0, 6, or 20 mg/kg decaBDE from postnatal day 2 to 15. Three groups of endpoints were examined: the ontogeny of sensorimotor responses and serum thyroxine levels in immature animals, and locomotor activity in adult animals. In immature animals, 20 mg/kg/day produced developmental delays in the acquisition of the palpebral reflex. At this age, exposed males also showed a dose-related reduction of serum thyroxine levels. As adults, decaBDE exposure altered the normal sex- and age-specific characteristics of spontaneous locomotor activity. The most striking effect was an increase of activity during the first 1.5 h of the 2 h assessment in males exposed to 20 mg/kg/day decaBDE. These effects suggest that decaBDE is a developmental neurotoxicant that can produce long-term behavioral changes following a discrete period of neonatal exposure.

Assessing the effects of environmental toxicant exposure in developmental epidemiological studies: issues for risk assessment.

Epidemiological studies have been of critical importance to the understanding of the effects of environmental chemical exposure during development on the behavior of infants and children. The ultimate goal of these studies should be to provide information that may be used directly for the protection of public health. The strategies for the assessment endpoints include development of domain-specific tests based on knowledge concerning effects of the chemicals being assessed, or use of standard clinical instruments that sample a range of functions. Discussion of an overall strategy for choice of endpoints would allow more straightforward comparisons across studies. There is increasing recognition of the importance of measuring a number of chemicals relevant to the population under study; however, different investigators make different decisions concerning which and how many chemicals to measure, as well as how to include them in the statistical analysis, particularly when there is a high degree of collinearity. Chemicals that are highly correlated with the "chemical of interest" are sometimes not included in the statistical analysis, resulting in missed opportunity to derive important information from the study. In addition, the shape of the relationship between exposure and effect is usually not explored in epidemiological studies, even though such information is critical for risk assessment. Opportunity for discussion among investigators, statisticians, and risk assessors potentially would result in human developmental toxicity studies being maximally useful for public health decisions.

Effects of PCB exposure on neuropsychological function in children.

In the last decade advances in the analytic methods for quantification of polychlorinated biphenyls (PCBs) have resulted in widespread availability of congener-specific analysis procedures, and large amounts of data on PCB congener profiles in soil, air, water, sediments, foodstuffs, and human tissues have become available. These data have revealed that the PCB residues in environmental media and human tissues may not closely resemble any of the commercial PCB mixtures, depending on source of exposure, bioaccumulation through the food chain, and weathering of PCBs in the environment. At the same time, toxicological research has led to a growing awareness that different classes of PCB congeners have different profiles of toxicity. These advances in analytic techniques and toxicological knowledge are beginning to influence the risk assessment process. As the data from ongoing PCB studies assessing the mediators of neurobehavioral outcomes in children are published, the weight of evidence for PCB effects on neurodevelopment is growing. Studies in Taiwan, Michigan (USA), New York (USA), Holland, Germany, and the Faroe Islands have all reported negative associations between prenatal PCB exposure and measures of cognitive functioning in infancy or childhood. The German study also reported a negative association between postnatal PCB exposure and cognitive function in early childhood--a result that had not been found in previous studies. Only one published study in North Carolina (USA) has failed to find an association between PCB exposure and cognitive outcomes. Despite the fact that several more recent studies have used congener-specific analytic techniques, there have been only limited attempts to assess the role of specific PCB congeners or classes of congeners in mediating neurodevelopmental outcomes. From a statistical standpoint, attempts to determine the role of individual congeners in mediating outcomes are hampered by the fact that concentrations of most individual congeners are highly correlated with each other and with total PCBs. From a toxicological standpoint, these efforts are hampered by the fact that many of the PCB congeners present in human tissues have never been studied in the laboratory, and their relative potency to produce nervous system effects is unknown. More complete information on the health effects of various congeners or congener classes would allow more informed scientific and risk assessment decisions.

Concentration of metals in blood of Maine children 1-6 years old.

Blood lead concentrations are higher in young children than in other age groups, whereas little is known regarding concentrations of other metals in young children. We measured the concentrations of a suite of metals in the blood of children 1-6 years of age, and assessed potential differences by age, season, or region of Maine. We used blood submitted to the Maine State Health and Environmental Testing Laboratory for blood lead analysis to determine the concentrations of arsenic (As), antimony (Sb), cadmium (Cd), manganese (Mn), mercury (Hg), selenium (Se), tin (Sn), and uranium (U) in 1350 children 1-6 years of age. The essential metals Mn and Se were detected in all samples, and As and Sb were detected in >90% of samples. Hg was detected in approximately 60% of samples. U and Cd were less often detected in blood samples, at approximately 30% and 10% of samples, respectively. Sn was not detected in any sample. Concentrations of As, Hg, and Se increased with age, whereas Sb decreased with age. Concentrations also varied by season and region for some though not all metals. Significant pairwise correlations were observed for a number of metals. Blood is a reasonable compartment for measurement of most of these metals in young children. The use of convenience samples provided a cost-effective mechanism for assessing exposure of young children in Maine.

Behavioral toxicology in the 21st century: challenges and opportunities for behavioral scientists. Summary of a symposium presented at the annual meeting of the neurobehavioral teratology society, June, 2009.

The National Research Council (NRC) of the National Academies of Science recently published a report of its vision of toxicity testing in the 21st century. The report proposes that the current toxicity testing paradigm that depends upon whole-animal tests be replaced with a strategy based upon in vitro tests, in silico models and evaluations of toxicity at the human population level. These goals are intended to set in motion changes that will transform risk assessment into a process in which adverse effects on public health are predicted by quantitative structure-activity relationship (QSAR) models and data from suites of high-throughput in vitro tests. The potential roles for whole-animal testing in this futuristic vision are both various and undefined. A symposium was convened at the annual meeting of the Neurobehavioral Teratology Society in Rio Grande, Puerto Rico in June, 2009 to discuss the potential challenges and opportunities for behavioral scientists in developing and/or altering this strategy toward the ultimate goal of protecting public health from hazardous chemicals. R. Kavlock described the NRC vision, introduced the concept of the 'toxicity pathway' (a central guiding principle of the NRC vision), and described the current status of an initial implementation this approach with the EPA's ToxCast(R) program. K. Crofton described a pathway based upon disruption of thyroid hormone metabolism during development, including agents, targets, and outcomes linked by this mode of action. P. Bushnell proposed a pathway linking the neural targets and cellular to behavioral effects of acute exposure to organic solvents, whose predictive power is limited by our incomplete understanding of the complex CNS circuitry that mediates the behavioral responses to solvents. B. Weiss cautioned the audience regarding a pathway approach to toxicity testing, using the example of the developmental toxicity of phthalates, whose effects on mammalian sexual differentiation would be difficult to identify based on screening tests in vitro. Finally, D. Rice raised concerns regarding the use of data derived from toxicity screening tests to human health risk assessments. Discussion centered around opportunities and challenges for behavioral toxicologists regarding this impending paradigm shift. Opportunities include: identifying and characterizing toxicity pathways; informing the conditions and limits of extrapolation; addressing issues of susceptibility and variability; providing reality-checks on selected positives and negatives from screens; and performing targeted testing and dose-response assessments of chemicals flagged during screening. Challenges include: predicting behavior using models of complex neurobiological pathways; standardizing study designs and dependent variables to facilitate creation of databases; and managing the cost and efficiency of behavioral assessments. Thus, while progress is being made in approaching the vision of 21st century toxicology, we remain a long way from replacing whole-animal tests; indeed, some animal testing will be essential for the foreseeable future at least. Initial advances will likely provide better prioritization tools so that animal resources are used more efficiently and effectively.

Does rapid metabolism ensure negligible risk from bisphenol A?

BACKGROUND: Bisphenol A (BPA) risks are being evaluated by many regulatory bodies because exposure is widespread and the potential exists for toxicity at low doses. OBJECTIVE: We evaluated evidence that BPA is cleared more rapidly in humans than in rats in relation to BPA risk assessment. DISCUSSION: The European Food Safety Authority (EFSA) relied on pharmacokinetic evidence to conclude that rodent toxicity data are not directly relevant to human risk assessment. Further, the EFSA argues that rapid metabolism will result in negligible exposure during the perinatal period because of BPA glucuronidation in pregnant women or sulfation in newborns. These arguments fail to consider the deconjugation of BPA glucuronide in utero by beta-glucuronidase, an enzyme that is present in high concentrations in placenta and various other tissues. Further, arylsulfatase C, which reactivates endogenous sulfated estrogens, develops early in life and so may deconjugate BPA sulfate in newborns. Biomonitoring studies and laboratory experiments document free BPA in rat and human maternal, placental, and fetal tissues, indicating that human BPA exposure is not negligible. The pattern of these detections is consistent with deconjugation in the placenta, resulting in fetal exposure. The tolerable daily intake set by the EFSA (0.05 mg/kg/day) is well above effect levels reported in some animal studies. CONCLUSION: This potential risk should not be dismissed on the basis of an uncertain pharmacokinetic argument. Rather, risk assessors need to decipher the BPA dose response and apply it to humans with comprehensive pharmacokinetic models that account for metabolite deconjugation.

Thyroid-disrupting chemicals: interpreting upstream biomarkers of adverse outcomes.

BACKGROUND: There is increasing evidence in humans and in experimental animals for a relationship between exposure to specific environmental chemicals and perturbations in levels of critically important thyroid hormones (THs). Identification and proper interpretation of these relationships are required for accurate assessment of risk to public health. OBJECTIVES: We review the role of TH in nervous system development and specific outcomes in adults, the impact of xenobiotics on thyroid signaling, the relationship between adverse outcomes of thyroid disruption and upstream causal biomarkers, and the societal implications of perturbations in thyroid signaling by xenobiotic chemicals. DATA SOURCES: We drew on an extensive body of epidemiologic, toxicologic, and mechanistic studies. DATA SYNTHESIS: THs are critical for normal nervous system development, and decreased maternal TH levels are associated with adverse neuropsychological development in children. In adult humans, increased thyroid-stimulating hormone is associated with increased blood pressure and poorer blood lipid profiles, both risk factors for cardiovascular disease and death. These effects of thyroid suppression are observed even within the "normal" range for the population. Environmental chemicals may affect thyroid homeostasis by a number of mechanisms, and multiple chemicals have been identified that interfere with thyroid function by each of the identified mechanisms. CONCLUSIONS: Individuals are potentially vulnerable to adverse effects as a consequence of exposure to thyroid-disrupting chemicals. Any degree of thyroid disruption that affects TH levels on a population basis should be considered a biomarker of adverse outcomes, which may have important societal outcomes.

Behavioral changes in aging but not young mice after neonatal exposure to the polybrominated flame retardant decaBDE.

BACKGROUND: After several decades of commercial use, the flame-retardant chemicals polybrominated diphenyl ethers (PBDEs) and their metabolites are pervasive environmental contaminants and are detected in the human body. Decabrominated diphenyl ether (decaBDE) is currently the only PBDE in production in the United States. OBJECTIVES: Little is known about the health effects of decaBDE. In the present study we examined the effects of neonatal decaBDE exposure on behavior in mice at two ages. METHODS: Neonatal male and female C57BL6/J mice were exposed to a daily oral dose of 0, 6, or 20 mg/kg decaBDE from postnatal days 2 through 15. Two age groups were examined: a cohort that began training during young adulthood and an aging cohort of littermates that began training at 16 months of age. Both cohorts were tested on a series of operant procedures that included a fixed-ratio 1 schedule of reinforcement, a fixed-interval (FI) 2-min schedule, and a light-dark visual discrimination. RESULTS: We observed minimal effects on the light-dark discrimination in the young cohort, with no effects on the other tasks. The performance of the aging cohort was significantly affected by decaBDE. On the FI schedule, decaBDE exposure increased the overall response rate. On the light-dark discrimination, older treated mice learned the task more slowly, made fewer errors on the first-response choice of a trial but more perseverative errors after an initial error, and had lower latencies to respond compared with controls. Effects were observed in both dose groups and sexes on various measures. CONCLUSIONS: These findings suggest that neonatal decaBDE exposure produces effects on behavioral tasks in older but not younger animals. The behavioral mechanisms responsible for the pattern of observed effects may include increased impulsivity, although further research is required.

Lifespan profiles of Alzheimer's disease-associated genes and products in monkeys and mice.

Alzheimer's disease (AD) is characterized by plaques of amyloid-beta (Abeta) peptide, cleaved from amyloid-beta protein precursor (AbetaPP). Our hypothesis is that lifespan profiles of AD-associated mRNA and protein levels in monkeys would differ from mice and that differential lifespan expression profiles would be useful to understand human AD pathogenesis. We compared profiles of AbetaPP mRNA, AbetaPP protein, and Abeta levels in rodents and primates. We also tracked a transcriptional regulator of the AbetaPP gene, specificity protein 1 (SP1), and the beta amyloid precursor cleaving enzyme (BACE1). In mice, AbetaPP and SP1 mRNA and their protein products were elevated late in life; Abeta levels declined in old age. In monkeys, SP1, AbetaPP, and BACE1 mRNA declined in old age, while protein products and Abeta levels rose. Proteolytic processing in both species did not match production of Abeta. In primates, AbetaPP and SP1 mRNA levels coordinate, but an inverse relationship exists with corresponding protein products as well as Abeta levels. Comparison of human DNA and mRNA sequences to monkey and mouse counterparts revealed structural features that may explain differences in transcriptional and translational processing. These findings are important for selecting appropriate models for AD and other age-related diseases.

Prenatal methylmercury exposure affects spatial vision in adult monkeys.

Decades of research have demonstrated that exposure to methylmercury (MeHg), a ubiquitous environmental pollutant, can have both early and long-term neurobehavioral consequences in exposed offspring. The present study assessed visual functioning in adult macaque monkeys (Macaca fascicularis) exposed in utero to 0, 50, 70, or 90 microg/kg/day of MeHg hydroxide. Twenty-one full-term, normal birth weight offspring (9 controls, 12 exposed) were tested at approximately 11-14.5 years of age on a visual contrast sensitivity task. A forced-choice tracking procedure was utilized with spatial frequencies of 1, 4, 10, and 20 cycles per degree of visual angle. On each test session, a single spatial frequency was presented across five levels of contrast, each differing by 3 dB. Methylmercury-exposed monkeys exhibited reduced contrast sensitivity thresholds, particularly at the higher spatial frequencies. The degree of visual impairment was not related to MeHg body burden or clearance and almost half of the exposed animals were unimpaired. The results from this study demonstrate that chronic in utero MeHg exposure, at subclinical levels, is associated with permanent adverse effects on spatial vision in adult monkeys.

The US EPA reference dose for methylmercury: sources of uncertainty.

The US Environmental Protection Agency (EPA) derived a reference dose for methylmercury in 2001, based on an extensive analysis by the National Research Council (NRC) of the National Academy of Sciences. The NRC performed benchmark dose analysis on a number of endpoints from three longitudinal prospective studies: the Seychelles Islands, the Faroe Islands, and the New Zealand studies. Adverse effects were reported in the latter two studies, but not in the Seychelles study. The NRC also performed an integrative analysis of all three studies. Dose conversion from cord blood or maternal hair mercury concentration was performed by EPA using a one-compartment pharmacokinetic model. A total uncertainty factor of 10 was applied for intrahuman pharmacokinetic and pharmacodynamic variability. There are numerous decisions made by the NRC/EPA that could greatly affect the value of the reference dose (RfD). Some of these include the choice of a linear model for the relationship between mercury body burden and neuropsychological performance, the choice of values of P0 and the benchmark response, the use of the "critical study/critical endpoint" approach in the interpretation of the maternal body burden that corresponds to the RfD, the use of central tendencies in a one-compartment pharmacokinetic model rather than the inclusion of the distributions of variables for the population of reproductive-age women, the assumption of unity for the ratio of fetal cord blood to maternal blood methylmercury concentrations, the choice of a total of 10 as an uncertainty factor, and the lack of dose-response analysis for other health effects such as cardiovascular disease. In addition, it may be argued that derivation of a RfD for methylmercury is inappropriate, given that there does not appear to be a threshold for adverse neuropsychological effects based on available data.

Methods and rationale for derivation of a reference dose for methylmercury by the U.S. EPA.

In 2001, the U.S. Environmental Protection Agency derived a reference dose (RfD) for methylmercury, which is a daily intake that is likely to be without appreciable risk of deleterious effects during a lifetime. This derivation used a series of benchmark dose (BMD) analyses provided by a National Research Council (NRC) panel convened to assess the health effects of methylmercury. Analyses were performed for a number of endpoints from three large longitudinal cohort studies of the neuropsychological consequences of in utero exposure to methylmercury: the Faroe Islands, Seychelles Islands, and New Zealand studies. Adverse effects were identified in the Faroe Islands and New Zealand studies, but not in the Seychelles Islands. The NRC also performed an integrative analysis of all three studies. The EPA applied a total uncertainty factor (UF) of 10 for intrahuman toxicokinetic and toxicodynamic variability and uncertainty. Dose conversion from cord blood mercury concentrations to maternal methylmercury intake was performed using a one-compartment model. Derivation of potential RfDs from a number of endpoints from the Faroe Islands study converged on 0.1 microg/kg/day, as did the integrative analysis of all three studies. EPA identified several areas for which further information or analyses is needed. Perhaps the most immediately relevant is the ratio of cord:maternal blood mercury concentration, as well as the variability around this ratio. EPA assumed in its dose conversion that the ratio was 1.0; however, available data suggest it is perhaps 1.5-2.0. Verification of a deviation from unity presumably would be translated directly into comparable reduction in the RfD. Other areas that EPA identified as significant areas requiring further attention are cardiovascular consequences of methylmercury exposure and delayed neurotoxicity during aging as a result of previous developmental or adult exposure.