The Toxicologist - Society of Toxicology

were collected from TK animals at designated time points on GD 6 and 17 in rats
and GD 6 and 18 in rabbits. On GD 20 or GD 29 maternal and developmental parameters
were evaluated and in rats, fetal exposure was assessed on GD 20. No maternal
and developmental toxicity was observed at any of the dose levels used in the
rat study. Evidence of fetal exposure was determined in fetal plasma with mean fetal
concentrations of PPD10558 and the metabolite (PPD11901) found to be between
1% and 6% of the mean maternal concentrations. In rabbits, marked maternal
toxicity including mortality (8 deaths; 1 doe at 25 and 7 at 50 mg/kg/day),
abortions (2 at 25 mg/kg/day and 6 at 50 mg/kg/day) and reduction in gestation
body weight, gestation body weight changes and decreased food consumption were
observed. In addition, fetal body weights of the combined sexes were significantly
reduced at 50 mg/kg/day in comparison to the controls. Based on a human dose of
80 mg BID the margins of exposure for teratogenic effects are at least 19x and 6x
for rats and rabbits, respectively, based on the top doses of 320 mg/kg/day for rats
and 50 mg/kg/day for rabbits. In conclusion, these studies show no evidence of teratogenicity
of PPD10558 in both rats and rabbits, though overt maternal toxicity
occurred in rabbits.
1631 FOXQ1: A BRIDGE BETWEEN THE AHR AND
HEDGEHOG SIGNALING PATHWAYS.
A. Planchart. Mount Desert Island Biological Laboratory, Salisbury Cove, ME.
The aryl hydrocarbon receptor (AHR) pathway has been of particular interest in
studying the effects of environmental exposures on development because this pathway
functions as an “antennae”, receiving input from the external environment that
leads to changes in gene expression. Toxicant activation of the AHR pathway leads
to craniofacial defects in model organisms including the zebrafish and mouse that
are similar in pathology to some human craniofacial abnormalities. The hedgehog
signaling pathway, a critical pathway required for the development of the vertebrate
head and face and implicated in several human craniofacial disorders, is misregulated
as a result of AHR pathway activation yet how this occurs is currently unknown.
Our lab was the first to identify foxq1l, an evolutionarily conserved forkhead
box transcription factor in zebrafish, as a target of the dioxin-activated AHR
pathway. Foxq1l is activated by dioxin exposure in the zebrafish head, particularly
in embryonic structures that give rise to the upper and lower jaw. We hypothesized
that the AHR pathway links environmental exposures to craniofacial birth defects
by misregulating the hedgehog signaling pathway via an intermediary signaling
molecule, possibly foxq1l. We now demonstrate that misexpressing foxq1l leads to
misregulation of the hedgehog pathway and abnormal craniofacial chondrogenesis,
and consequently to abnormal head development in affected embryos, supporting
the hypothesis that foxq1l functions as a bridge between the AHR and hedgehog
signaling pathways. We propose that this mechanism is conserved in all vertebrates.
1632 CLEFT PALATE IN RATS TREATED WITH A
COMPOUND THAT HAS STEROL ANTISYNTHESIS
ACTION.
N. Ikemi 1 , T. Oota 2 and K. Kawashima 2 . 1 Otsuka AgriTechno Co., Ltd.,
Tokushima, Japan and 2 Mitsubishi Chemical Medience Co., Ltd., Kumamoto, Japan.
[Objective] We have been conducting experiments to investigate the mode of occurrence
of cleft palate. The present experiments were designed to specify the period
of sensitivity to a compound with a sterol synthesis inhibition action in induction
of cleft palate in SD rats and to determine whether induction of cleft palate
was due to the parent compound or its metabolite following amniotic fluid administration
of this compound.
[Methods] (Experiment 1) To identify its cleft palate induction action, forced oral
administration of the compound to the animals was conducted once daily for 10
days from the 6th day to 15th day of pregnancy (vaginal plug confirmation day: day
0 of pregnancy). The doses were set at 15, 20 and 25 mg/kg. The animals underwent
cesarean section on day 20 of pregnancy. (Experiment 2) To identify the critical
period for the occurrence of cleft palate, a single dose (100 mg/kg) of the compound
was administered to dams, and the animals underwent cesarean section on
day 20 of pregnancy. (Experiment 3) To investigate whether the induction of cleft
palate following administration of the compound was caused by the compound itself
or by its metabolite, the abdomens of the dams were incised, and the compound
was administered into the amniotic fluid through the uterine wall. Results]
(Experiment 1) The incidences of cleft palate were 11%, 43% and 76% in the 15,
20 and 25 mg/kg groups, respectively. (Experiment 2) The occurrence of cleft
palate was observed following administration of every single dose on days 10, 11,
12 and 13 of gestation. Based on a comparison of the incidences of cleft palate, the
critical period for the occurrence of cleft palate was considered to be the day 13 of
gestation. (Experiment 3) No cleft palate was observed. [Conclusion] It was possible
that metabolites of the compound induced cleft palate as cleft palate occurred
frequently in fetuses of dams receiving an oral dose of the compound, while it was
not observed in fetuses given the compound via the intraamniotic route.
1633 CHEMICAL FORM MATTERS: DIFFERENTIAL
ACCUMULATION OF ORGANIC AND INORGANIC
MERCURY IN ZEBRAFISH LARVAE.
T. C. MacDonald, M. Korbas, I. J. Pickering, G. N. George and P. H. Krone.
University of Saskatchewan, Saskatoon, SK, Canada.
Mercury is found in both organic and inorganic forms in the atmosphere, in sediment
and in water bodies due to natural and anthropogenic sources. Limited information
is available on the uptake and accumulation of Hg in developing organisms,
particularly as it relates to chemical form of the metal. To address this problem we
utilized synchrotron x-ray fluorescence imaging in zebrafish (Danio rerio), an increasingly
well studied model vertebrate for investigating mechanisms of chemical
toxicity. Zebrafish larvae were exposed to one of four forms of mercury (methyl
mercury chloride, methyl mercury L-cysteine, mercuric chloride, and mercury bis-
L-cysteineate). Adjacent serial sections were utilized for synchrotron imaging and
histological staining, respectively. Sections were imaged using the X-ray fluorescence
imaging technique on beamline 20-ID at the Advanced Photon Source (APS)
in Argonne, IL. Significant variations in mercury accumulation were found in fish
exposed to organic mercury as compared to fish exposed to inorganic mercury.
Larvae exposed to inorganic mercury exhibited accumulation in the ventricular region
of the brain. However, only the fish exposed to organic mercury exhibited accumulation
in the lens epithelium, gut tube, and skeletal muscles. These variations
demonstrate the importance of considering chemical form when considering the
uptake and accumulation of mercury. Ongoing research will investigate the efficacy
of chelating agents. Although dimercaptosuccinic acid (DMSA) is not a true chelator
it is currently used to treat mercury exposure. As DMSA is poorly optimized for
this role it is not always effective. Other sequestration agents of interest are Nacetylcysteine
(NAC) and alpha lipoic acid (ALA). Eventually, we hope to design
and test a custom chelator that can bind and excrete different forms of mercury
more effectively.
1634 ABERRANT LIGAND-INDUCED ACTIVATION OF G-
PROTEIN-COUPLED ESTROGEN RECEPTOR 1 (GPER)
RESULTS IN DEVELOPMENTAL MALFORMATIONS
DURING VERTEBRATE EMBRYOGENESIS.
B. S. Jayasinghe and D. C. Volz. Department of Environmental Health Sciences,
Arnold School of Public Health, University of South Carolina, Columbia, SC.
G-protein-coupled estrogen receptor 1 (GPER) – a GPCR unrelated to nuclear estrogen
receptors (ERs) but strongly activated by estradiol in both mammals and fish
– is expressed and functional in numerous vertebrate organ systems and has been
implicated as a potential target for xenoestrogen-mediated signaling in reproductive
organs and tumors. To date, the distribution and functional characterization of
GPER within vertebrate organs have been restricted to juvenile and adult animals.
In contrast, virtually nothing is known about the spatiotemporal distribution and
function of GPER during vertebrate embryogenesis. Using zebrafish as an animal
model, the aim of this study was to assess the functional role of GPER during embryogenesis.
RT-PCR and whole mount in situ hybridization data suggest that
GPER mRNA is expressed as early as 1-hr post-fertilization (hpf) and is strongly localized
in the central nervous system. Continuous exposure to a selective GPER agonist
(G-1) from 8 to 96 hpf resulted in dose-dependent effects on survival and
gross larval morphology, including decreased body length, tail abnormalities, and
axial curvature. However, exposure to a selective GPER antagonist (G-15) resulted
in no discernible effects on survival or gross larval morphology. Importantly, based
on co-exposure studies with G-1 and G-15, G-15 fully blocked G-1-induced morphologic
effects, suggesting that G-1-induced effects are mediated via aberrant activation
of GPER. Using reverse genetics (morpholino-based) strategies, we are currently
testing whether (1) GPER knockdown alone results in developmental
abnormalities and (2) translation of GPER (or other ERs) is required for G-1-induced
effects on gross larval morphology. Overall, our findings suggest that aberrant
ligand-induced GPER activation represents a potentially novel and understudied
mode-of-action for environmentally relevant chemicals that affect vertebrate
embryogenesis.
1635 PRENATAL BISPHENOL-A ALTERS LUNG EPITHELIAL
SECRETORY CELL MATURATION.
S. Murphy, M. Boetticher, C. VandeVoort and L. Van Winkle. California
National Primate Research Center, UC Davis, Davis, CA.
Bisphenol-A (BPA) is used in the production of polycarbonate plastics. We hypothesized
that exposure to BPA can disrupt fetal lung development because both airway
and alveolar development occur in the prenatal period. To test this hypothesis
we exposed timed pregnant Rhesus macaques to BPA. There were 3 exposure
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