Standardizing Extracted Data Using Automated Application of Controlled Vocabularies.

BACKGROUND: Extraction of toxicological end points from primary sources is a central component of systematic reviews and human health risk assessments. To ensure optimal use of these data, consistent language should be used for end point descriptions. However, primary source language describing treatment-related end points can vary greatly, resulting in large labor efforts to manually standardize extractions before data are fit for use. OBJECTIVES: To minimize these labor efforts, we applied an augmented intelligence approach and developed automated tools to support standardization of extracted information via application of preexisting controlled vocabularies. METHODS: We created and applied a harmonized controlled vocabulary crosswalk, consisting of Unified Medical Language System (UMLS) codes, German Federal Institute for Risk Assessment (BfR) DevTox harmonized terms, and The Organization for Economic Co-operation and Development (OECD) end point vocabularies, to roughly 34,000 extractions from prenatal developmental toxicology studies conducted by the National Toxicology Program (NTP) and 6,400 extractions from European Chemicals Agency (ECHA) prenatal developmental toxicology studies, all recorded based on the original study report language. RESULTS: We automatically applied standardized controlled vocabulary terms to 75% of the NTP extracted end points and 57% of the ECHA extracted end points. Of all the standardized extracted end points, about half (51%) required manual review for potential extraneous matches or inaccuracies. Extracted end points that were not mapped to standardized terms tended to be too general or required human logic to find a good match. We estimate that this augmented intelligence approach saved >350 hours of manual effort and yielded valuable resources including a controlled vocabulary crosswalk, organized related terms lists, code for implementing an automated mapping workflow, and a computationally accessible dataset. DISCUSSION: Augmenting manual efforts with automation tools increased the efficiency of producing a findable, accessible, interoperable, and reusable (FAIR) dataset of regulatory guideline studies. This open-source approach can be readily applied to other legacy developmental toxicology datasets, and the code design is customizable for other study types. https://doi.org/10.1289/EHP13215.

Early Developmental Exposure to Triclosan Impacts Fecal Microbial Populations, IgA and Functional Activities of the Rat Microbiome.

Triclosan (TCS), a broad-spectrum antibacterial chemical, is detected in human urine, breast milk, amniotic fluid, and feces; however, little is known about its impact on the intestinal microbiome and host mucosal immunity during pregnancy and early development. Pregnant female rats were orally gavaged with TCS from gestation day (GD) 6 to postpartum (PP) day 28. Offspring were administered TCS from postnatal day (PND) 12 to 28. Studies were conducted to assess changes in the intestinal microbial population (16S-rRNA sequencing) and functional analysis of microbial genes in animals exposed to TCS during pregnancy (GD18), and at PP7, PP28 and PND28. Microbial abundance was compared with the amounts of TCS excreted in feces and IgA levels in feces. The results reveal that TCS decreases the abundance of Bacteroidetes and Firmicutes with a significant increase in Proteobacteria. At PND28, total Operational Taxonomic Units (OTUs) were higher in females and showed correlation with the levels of TCS and unbound IgA in feces. The significant increase in Proteobacteria in all TCS-treated rats along with the increased abundance in OTUs that belong to pathogenic bacterial communities could serve as a signature of TCS-induced dysbiosis. In conclusion, TCS can perturb the microbiome, the functional activities of the microbiome, and activate mucosal immunity during pregnancy and early development.

Broadening the Environmental Lens to Include Social and Structural Determinants of Women's Health Disparities.

BACKGROUND: Due to the physical, metabolic, and hormonal changes before, during, and after pregnancy, women-defined here as people assigned female at birth-are particularly susceptible to environmental insults. Racism, a driving force of social determinants of health, exacerbates this susceptibility by affecting exposure to both chemical and nonchemical stressors to create women's health disparities. OBJECTIVES: To better understand and address social and structural determinants of women's health disparities, the National Institute of Environmental Health Sciences (NIEHS) hosted a workshop focused on the environmental impacts on women's health disparities and reproductive health in April 2022. This commentary summarizes foundational research and unique insights shared by workshop participants, who emphasized the need to broaden the definition of the environment to include upstream social and structural determinants of health. We also summarize current challenges and recommendations, as discussed by workshop participants, to address women's environmental and reproductive health disparities. DISCUSSION: The challenges related to women's health equity, as identified by workshop attendees, included developing research approaches to better capture the social and structural environment in both human and animal studies, integrating environmental health principles into clinical care, and implementing more inclusive publishing and funding approaches. Workshop participants discussed recommendations in each of these areas that encourage interdisciplinary collaboration among researchers, clinicians, funders, publishers, and community members. https://doi.org/10.1289/EHP12996.

Reproductive and developmental toxicity following exposure to organophosphate ester flame retardants and plasticizers, triphenyl phosphate and isopropylated phenyl phosphate, in Sprague Dawley rats.

Two organophosphate esters used as flame retardants and plasticizers, triphenyl phosphate (TPHP) and isopropylated phenyl phosphate (IPP), have been detected in environmental samples around the world. Human exposure primarily occurs via oral ingestion with reported higher concentrations in children. Currently, there are no data to evaluate potential risk from exposure to either TPHP or IPP during fetal development. These short-term perinatal studies in rats provide preliminary toxicity data for TPHP and IPP, including information on transfer to fetus/offspring and across the pup blood-brain barrier. In separate experiments, TPHP or IPP were administered via dosed feed at concentrations 0, 1000, 3000, 10 000, 15 000, or 30 000 ppm to time-mated Hsd:Sprague Dawley SD rats from gestation day (GD) 6 through postnatal day (PND) 28; offspring were provided dosed feed at the same concentration as their dam (PND 28-PND 56). TPHP- and IPP-related toxicity resulted in removal of both 30 000 ppm groups on GD 12 and 15 000 ppm IPP group after parturition. Body weight and organ weights were impacted with exposure in remaining dams. Reproductive performance was perturbed at ≥10 000 ppm TPHP and all IPP exposure groups. In offspring, both TPHP- and IPP-related toxicity was noted in pups at ≥10 000 ppm as well as reduction in bodyweights, delays in pubertal endpoints, and/or reduced cholinesterase enzyme activity starting at 1000 ppm TPHP or IPP. Preliminary internal dose assessment indicated gestational and lactational transfer following exposure to TPHP or IPP. These findings demonstrate that offspring development is sensitive to 1000 ppm TPHP or IPP exposure.

Comparative toxicokinetics of bisphenol S and bisphenol AF in male rats and mice following repeated exposure via feed.

We investigated the plasma toxicokinetic behavior of free (parent) and total (parent and conjugated forms) of bisphenol S (BPS) and bisphenol AF (BPAF) in plasma of adult male rats and mice following exposure via feed for 7 days to BPS (338, 1125, and 3375 ppm) or BPAF (338, 1125, and 3750 ppm). In rats, the exposure concentration-normalized maximum concentration [Cmax/D (ng/mL)/(ppm)] and area under the concentration time curve [AUC/D (h × ng/mL)/(ppm)] for free was higher for BPS (Cmax/D: 0.476-1.02; AUC/D: 3.58-8.26) than for BPAF (Cmax/D: 0.017-0.037; AUC/D:0.196-0.436). In mice, the difference in systemic exposure parameters between free BPS (Cmax/D: 0.376-0.459; AUC/D: 1.52-2.54) and free BPAF (Cmax/D: 0.111-0.165; AUC/D:0.846-1.09) was marginal. Elimination half-lives for free analytes (4.41-10.4 h) were comparable between species and analogues. When systemic exposure to free analyte was compared between species, in rats, BPS exposure was slightly higher but BPAF exposure was much lower than in mice. BPS and BPAF were highly conjugated; total BPS AUC values (rats ≥18-fold, mice ≥17-fold) and BPAF (rats ≥127-fold, mice ≥16-fold) were higher than corresponding free values. Data demonstrated that there are analogue and species differences in the kinetics of BPS and BPAF.

Differential Toxicological Outcome of Corn Oil Exposure in Rats and Mice as Assessed by Microbial Composition, Epithelial Permeability, and Ileal Mucosa-Associated Immune Status.

Studies to evaluate the toxicity of xenobiotics on the human gut microbiome and related health effects require a diligent selection of (1) an appropriate animal model to facilitate toxicity assessment in predicting human exposure, and (2) an appropriate non-interfering vehicle for the administration of water insoluble compounds. In biomedical studies with water insoluble xenobiotics, corn oil is one of the most commonly used nonaqueous vehicles. This study evaluated the suitability of corn oil as a vehicle in adult female Sprague Dawley rats and adult CD-1 mice; the rodent models that are often utilized in toxicological studies. We studied the host response in terms of change in the intestinal microbiome and mRNA expression of intestinal permeability and immune response-related genes when water (control) and corn oil (2 ml/kg) were administered as a vehicle through oral gavage. The results showed that the use of corn oil as a vehicle has no adverse impact in rats for either the immune response or the intestinal microbial population. On the other hand, mice treated with corn oil showed changes in bacterial community adhered to the ileum, as well as changes in the mRNA expression of intestinal permeability-related and ileal mucosa-associated immune response genes. Overall, results of this study suggest that the type of rodent species and vehicle used in toxicological risk assessments of xenobiotics studies should be taken into consideration in the experimental setup and study design.

An investigation of systemic exposure to bisphenol AF during critical periods of development in the rat.

Due to structural similarity to bisphenol A and lack of safety data, the National Toxicology Program (NTP) is evaluating the potential toxicity of bisphenol AF (BPAF) in rodent models. The current investigation reports the internal exposure data for free (unconjugated BPAF) and total (free and conjugated forms) BPAF during critical stages of development following perinatal dietary exposure in Hsd:Sprague Dawley®SD® rats to 0 (vehicle control), 338, 1125, and 3750 ppm BPAF from gestation day (GD) 6 to postnatal day (PND) 28. Free and total BPAF concentrations in maternal plasma at GD 18, PND 4, and PND 28 increased with the exposure concentration; free BPAF concentrations were ≤ 1.61% those of total BPAF demonstrating extensive first pass metabolism of BPAF following dietary exposure in adults. Free and total BPAF were quantified in GD 18 fetuses and PND 4 pups with free concentrations 11.7-53.4% that of corresponding total concentrations. In addition, free concentrations were higher (130-571%) and total concentrations were lower (1.71-7.23%) than corresponding concentrations in dams, demonstrating either preferential transfer of free BPAF and/or inability of fetuses and pups to conjugate BPAF. Free and total concentrations in PND 28 pups were similar to maternal concentrations demonstrating direct exposure of pups via feed and that conjugating enzymes are developed in PND 28 pups. In conclusion, these data demonstrate considerable gestational and lactational transfer of parent aglycone from the mother to offspring. Since the ontogeny of conjugating enzymes in humans is similar to that of rodents, the data from rodent BPAF studies may be useful in predicting human risk from exposure to BPAF.

Environmental Factors Involved in Maternal Morbidity and Mortality.

Nongenetic, environmental factors contribute to maternal morbidity and mortality through chemical exposures via air, water, soil, food, and consumer products. Pregnancy represents a particularly sensitive window of susceptibility during which physiological changes to every major organ system increase sensitivity to chemicals that can impact a woman's long-term health. Nonchemical stressors, such as low socioeconomic status, may exacerbate the effects of chemical exposures on maternal health. Racial/ethnic minorities are exposed disproportionately to both chemicals and nonchemical stressors, which likely contribute to the observed health disparities for maternal morbidities and mortality. Epidemiological studies linking exposures to adverse maternal health outcomes underscore the importance of environmental health impacts, and mechanistic studies in model systems reveal how chemicals perturb biological pathways and processes. Environmental stressors are associated with a variety of immediate maternal health impacts, including hypertensive disorders of pregnancy, fibroids, and infertility, as well as long-term maternal health impacts, such as higher risk of breast cancer and metabolic disorders. Identifying and reducing a pregnant woman's environmental exposures is not only beneficial to her offspring but also important to preserve her short- and long-term health.

Comparative toxicokinetics of bisphenol S in rats and mice following gavage administration.

Bisphenol S (BPS) is a component of polyether sulfone used in a variety of industrial applications and consumer products. We investigated the plasma toxicokinetic (TK) behavior of free (unconjugated parent) and total (parent and conjugated) BPS in rats and mice following a single gavage administration (34, 110, or 340 mg/kg). In male rats, BPS was rapidly absorbed with free BPS maximum concentration (Cmax) reached at ≤2.27 h. Elimination of free BPS in male rats was dose-dependent with estimated half-lives of 5.77-11.9 h. Cmax and area under the concentration versus time curve (AUC) increased with dose although the increase in AUC was more than dose proportional. In male rats, total BPS Cmax was reached ≤2.77 h with both Cmax (≥ 10-fold) and AUC (≥ 15-fold) higher than free BPS demonstrating rapid and extensive conjugation of BPS. In male mice, the increase in Cmax and AUC of free BPS was dose-proportional; Cmax was higher and AUC was lower than in male rats. BPS was cleared more rapidly in male mice (half-life 2.86-4.21 h) compared to male rats (half-life 5.77-11.9 h). Similar to rats, total BPS Cmax (≥ 6-fold) and AUC (≥ 12-fold) were higher than corresponding free BPS. Oral bioavailability of free BPS was low to moderate (rats, ≤ 21%; mice, ≤ 19%). There were some species differences in TK parameters of free and total BPS and limited sex difference in rats and mice. In addition, there were dose-related effects of plasma TK parameters in rats.

The Gut Microbiome and Xenobiotics: Identifying Knowledge Gaps.

There is an increasing awareness that the gut microbiome plays a critical role in human health and disease, but mechanistic insights are often lacking. In June 2018, the Health and Environmental Sciences Institute (HESI) held a workshop, "The Gut Microbiome: Markers of Human Health, Drug Efficacy and Xenobiotic Toxicity" (https://hesiglobal.org/event/the-gut-microbiome-workshop) to identify data gaps in determining how gut microbiome alterations may affect human health. Speakers and stakeholders from academia, government, and industry addressed multiple topics including the current science on the gut microbiome, endogenous and exogenous metabolites, biomarkers, and model systems. The workshop presentations and breakout group discussions formed the basis for identifying data gaps and research needs. Two critical issues that emerged were defining the microbial composition and function related to health and developing standards for models, methods and analysis in order to increase the ability to compare and replicate studies. A series of key recommendations were formulated to focus efforts to further understand host-microbiome interactions and the consequences of exposure to xenobiotics as well as identifying biomarkers of microbiome-associated disease and toxicity.

Disposition and metabolism of antibacterial agent, triclocarban, in rodents; a species and route comparison.

Triclocarban is a residue-producing antibacterial agent used in a variety of consumer products. These studies investigated the disposition and metabolism of [14C]triclocarban. In male rats following a single gavage administration of 50, 150, and 500 mg/kg, excretion was primarily via feces (feces, 85-86%; urine, 3-6%) with no apparent dose-related effect. In male rats, 29% of the administered dose was excreted in bile suggesting some of the fecal excretion is from the absorbed dose which was excreted to the intestine via bile. The tissue retention of radioactivity was low in male rats (24 h, 3.9%; 72 h, 0.1%). Disposition pattern following gavage administration of 50 mg/kg in female rats and male and female mice were similar to male rats. Plasma elimination half-life of triclocarban in rats following gavage administration was shorter (∼2 h) compared to that based on total radioactivity (≥9 h) which included all products of triclocarban. Absorption following a single dermal application of 1.5 or 3% was low (≤3%) in rodents. Hydroxylated and conjugated metabolites of triclocarban predominated in bile. In hepatocytes, clearance of triclocarban in mouse and human was similar and was faster than in rat.

Postnatal Effects of Gestational and Lactational Gavage Exposure to Boric Acid in the Developing Sprague Dawley Rat.

Human exposure to boron occurs primarily through diet and drinking water sources. Animal studies have found that reduced fetal weight following gestational exposure to boron (as boric acid) is the most sensitive toxicological effect. However, recent studies suggest that newborns in areas with elevated boron in drinking water may receive levels of exposure that exceed the U.S. EPA oral reference dose for B. Currently, there are no data to inform a boron risk assessment accounting for this developmental window. To address this knowledge gap, the National Toxicology Program evaluated developmental toxicity following pre- and postnatal boron exposure. Time-mated female Sprague Dawley (Hsd: Sprague Dawley SD) rats were administered 0-20 mg B/kg/day (as boric acid) via gavage from gestation day 6 to 21; offspring were dosed via gavage at the same respective dose level from postnatal day (PND) 1 to 28. There were no dose-related effects on dam bodyweight, bodyweight gain, or feed consumption. Clinical findings were limited to low incidences of umbilical hernia in the 20 mg B/kg pups which resolved by study completion. Pup plasma boron concentrations increased in dose-proportional manner and were similar between PND 4 and PND 28. Postnatal weight gain was significantly reduced at 20 mg B/kg, with male and female pups weighing 23% less than the controls on PND 28. These findings demonstrate that postnatal growth in the Sprague Dawley rat is sensitive to boron exposure and highlights the importance of evaluating the potential toxicity of agents with known human exposures during early life stages.

Toxicokinetics and bioavailability of bisphenol AF following oral administration in rodents: A dose, species, and sex comparison.

We investigated the toxicokinetics and bioavailability of bisphenol AF (BPAF) in male and female Harlan Sprague Dawley rats and B6C3F1/N mice following a single gavage administration of 34, 110, or 340 mg/kg. A validated analytical method was used to quantitate free (unconjugated parent) and total (unconjugated and conjugated) BPAF in plasma. BPAF was rapidly absorbed in rats with the maximum plasma concentration, Cmax, of free BPAF reached at ≤2.20 h. BPAF was cleared rapidly with a plasma elimination half-life of ≤3.35 h. Cmax and the area under the concentration versus time curve, AUC0-∞, increased proportionally to the dose. Total BPAF Cmax was reached ≤1.07 h in rats with both Cmax (≥27-fold) and AUC0-∞ (≥52-fold) much higher than corresponding free values demonstrating rapid and extensive conjugation of BPAF following oral administration. Absorption of BPAF following a 34 mg/kg gavage dose in mice was more rapid than in rats with free BPAF Cmax reached ≤0.455 h. Free BPAF was cleared rapidly in mice with an elimination half-life of ≤4.22 h. Similar to rats, total BPAF was much higher than corresponding free BPAF. There was no apparent sex-related effect in plasma toxicokinetic parameters of free or total BPAF in mice and rats. Bioavailability in rats was ~ 1% with no apparent dose-related effect. Bioavailability in mice was slightly higher than in rats (male ~ 6%, female 3%). These data demonstrate that BPAF was rapidly absorbed following gavage administration in rodents, rapidly and extensively conjugated with low bioavailability.

Disposition and metabolism of the bisphenol analogue, bisphenol S, in Harlan Sprague Dawley rats and B6C3F1/N mice and in vitro in hepatocytes from rats, mice, and humans.

With the removal of bisphenol A (BPA) from many consumer products, the potential use of alternatives such as bisphenol S (BPS) and its derivatives is causing some concerns. These studies investigated the comparative in vitro hepatic clearance and metabolism of BPS and derivatives and the disposition and metabolism of BPS in rats and mice following gavage and intravenous administration. The clearance of BPS and its derivatives was slower in human hepatocytes than in rodents. In male rats following gavage administration of 50, 150, and 500 mg/kg [14C]BPS the main route of excretion was via urine; the urinary excretion decreased (72 to 48%) and the fecal excretion increased (16 to 30%) with increasing dose. The disposition was similar in female rats and male and female mice following gavage administration. Radioactivity remaining in tissues at 72 h in both species and sexes was ≤2.4%. In bile duct cannulated rats 53% of a gavage dose was secreted in bile suggesting extensive enterohepatic recirculation of [14C]BPS. Following an intravenous dose in rats and mice, the pattern of excretion was similar to gavage. These data suggest that the dose excreted in feces folowing gavage administration is likely the absorbed dose. Urinary metabolites included the glucuronide and sulfate conjugates with a moderate amount of parent. The pattern of in vitro hepatic metabolsim was similar to in vivo with some difference among derivatives. These data suggest that similar to other bisphenol analogues, BPS was well absorbed following oral expsosure and extensively excreted with minimal tissue retention.

Embryo-fetal development studies with the dietary supplement vinpocetine in the rat and rabbit.

Dietary supplement and natural product use is increasing within the United States, resulting in growing concern for exposure in vulnerable populations, including young adults and women of child-bearing potential. Vinpocetine is a semisynthetic derivative of the Vinca minor extract, vincamine. Human exposure to vinpocetine occurs through its use as a dietary supplement for its purported nootropic and neuroprotective effects. To investigate the effects of vinpocetine on embryo-fetal development, groups of 25 pregnant Sprague-Dawley rats and 8 pregnant New Zealand White rabbits were orally administered 0, 5, 20, or 60 mg vinpocetine/kg and 0, 25, 75, 150, or 300 mg/kg daily from gestational day (GD) 6-20 and GD 7-28, respectively. Pregnant rats dosed with vinpocetine demonstrated dose-dependent increases in postimplantation loss, higher frequency of early and total resorptions, lower fetal body weights, and fewer live fetuses following administration of 60 mg/kg, in the absence of maternal toxicity. Additionally, the rat fetuses displayed dose-dependent increases in the incidences of ventricular septum defects and full supernumerary thoracolumbar ribs. Similarly, albeit at higher doses than the rats, pregnant rabbits administered vinpocetine displayed an increase in postimplantation loss and fewer live fetuses (300 mg/kg), in addition to significantly lower fetal body weights (≥75 mg/kg). In conclusion, vinpocetine exposure resulted in similar effects on embryo-fetal development in the rat and rabbit. The species differences in sensitivity and magnitude of response is likely attributable to a species difference in metabolism. Taken together, these data suggest a potential hazard for pregnant women who may be taking vinpocetine.

Disposition of [14C]hydroquinone in Harlan Sprague-Dawley rats and B6C3F1/N mice: species and route comparison.

1. Hydroquinone (HQ) is present in some foods and has varied industrial, medical and consumer uses. These studies were undertaken to investigate the disposition of HQ in rats and mice following gavage, intravenous (IV) and dermal exposure. 2. [14 C]HQ administered (0.5, 5 or 50 mg/kg) by gavage or IV routes to male and female Harlan Sprague-Dawley (HSD) rats and B6C3F1/N mice was well absorbed and rapidly excreted primarily in urine. Radioactivity remaining in tissues at 72 h was <1% for both species at all dose levels and routes. No sex, species or route related differences in disposition were found. 3. With dermal application of 2, 10 or 20% [14 C]HQ, mice absorbed higher percentages of the dose than rats (37, 12, 12% versus 18.6, 4.43 and 1.79%, respectively). The HQ mass absorbed by mice increased with dose, while in rats it was more constant over the dose range. Absorbed HQ was rapidly excreted in urine of both species and urinary excretion indicated continued absorption over the exposure period. No sex differences in disposition were found. 4. The oral bioavailability of HQ at 5 mg/kg was low in both rats (1.6%) and mice (3.9%) demonstrating significant first pass metabolism. Dermal bioavailability in mice was 9.4% following application of 2% formulation. 5. Urinary metabolites for both species and all routes included the glucuronide and sulfate conjugates; no parent was found in urine.

Scientific and Regulatory Policy Committee Points to Consider Review: Inclusion of Reproductive and Pathology End Points for Assessment of Reproductive and Developmental Toxicity in Pharmaceutical Drug Development.

Standard components of nonclinical toxicity testing for novel pharmaceuticals include clinical and anatomic pathology, as well as separate evaluation of effects on reproduction and development to inform clinical development and labeling. General study designs in regulatory guidances do not specifically mandate use of pathology or reproductive end points across all study types; thus, inclusion and use of these end points are variable. The Scientific and Regulatory Policy Committee of the Society of Toxicologic Pathology (STP) formed a Working Group to assess the current guidelines and practices on the use of reproductive, anatomic pathology, and clinical pathology end points in general, reproductive, and developmental toxicology studies. The Working Group constructed a survey sent to pathologists and reproductive toxicologists, and responses from participating organizations were collected through the STP for evaluation by the Working Group. The regulatory context, relevant survey results, and collective experience of the Working Group are discussed and provide the basis of each assessment by study type. Overall, the current practice of including specific end points on a case-by-case basis is considered appropriate. Points to consider are summarized for inclusion of reproductive end points in general toxicity studies and for the informed use of pathology end points in reproductive and developmental toxicity studies.

Micro-CT imaging: Developing criteria for examining fetal skeletons in regulatory developmental toxicology studies - A workshop report.

During the past two decades the use and refinements of imaging modalities have markedly increased making it possible to image embryos and fetuses used in pivotal nonclinical studies submitted to regulatory agencies. Implementing these technologies into the Good Laboratory Practice environment requires rigorous testing, validation, and documentation to ensure the reproducibility of data. A workshop on current practices and regulatory requirements was held with the goal of defining minimal criteria for the proper implementation of these technologies and subsequent submission to regulatory agencies. Micro-computed tomography (micro-CT) is especially well suited for high-throughput evaluations, and is gaining popularity to evaluate fetal skeletons to assess the potential developmental toxicity of test agents. This workshop was convened to help scientists in the developmental toxicology field understand and apply micro-CT technology to nonclinical toxicology studies and facilitate the regulatory acceptance of imaging data. Presentations and workshop discussions covered: (1) principles of micro-CT fetal imaging; (2) concordance of findings with conventional skeletal evaluations; and (3) regulatory requirements for validating the system. Establishing these requirements for micro-CT examination can provide a path forward for laboratories considering implementing this technology and provide regulatory agencies with a basis to consider the acceptability of data generated via this technology.

Spatial behavior and seizure-induced changes in c-fos mRNA expression in young and old rats.

The subcellular processes of gene induction and expression in the hippocampus are likely to underlie some of the known age-related impairments in spatial learning and memory. It is well established that immediate-early genes are rapidly and transiently induced in response to neuronal activity and this expression is required for stabilization of durable memories. To examine whether age-related memory impairment might be caused, in part, by differences in the level of cellular activation or subcellular processing, c-fos expression in CA1 pyramidal and dentate gyrus granule cells in the dorsal hippocampus of young and old rats was determined using fluorescence in situ hybridization and reverse transcription polymerase chain reaction. No significant age differences were found in the numbers of pyramidal or granule cells that show c-fos expression; however, c-fos mRNA transcripts were altered in these 2 cell types in aged animals. These findings suggest that though the networks of cells that participate in behavior or seizure-induced activity are largely maintained in aged rats, their RNA transcript levels are altered. This might, in part, contribute to cognitive deficits frequently observed with advancing age.