bluuebunny/crossref_metadata_2025 huggingface dataset:
Cadmium exposure on day 12 of gestation in the Wistar rat: Distribution, uteroplacental blood flow, and fetal viability
Authors: Saltzman Risa A. , Miller Richard K. , di Sant'Agnese P. A.
Published: June 2005

The effects of cadmium exposure (40 μmole CdCl2/kg, s.c.) on day 12 of gestation were evaluated in the Wistar rat. At 16–18 hours following such cadmium exposure, blood flow (as determined by radiolabeled microspheres) to the chorioallantoic placenta (CAP) was significantly reduced by 35%; at 24–26 hours, blood flow to the CAP had returned to control levels and was still unaffected at 38–43 hours. Uterine blood flow was not significantly altered at any of these timepoints. Between 16–18 and 24–26 hours after cadmium exposure, the concentration of cadmium in the placenta decreased significantly, while total cadmium content did not change. By 38–43 hours after cadmium exposure, total cadmium content of the placenta had increased significantly, although cadmium concentration was unchanged. There were no adverse effects on fetal viability or growth, as determined on day 20 of gestation. In sharp contrast, near‐term (day 18) exposure to 40 or 50 μmole CdCl2/kg (s.c.) resulted in 53% and 82% mean incidences of fetolethality, respectively, within 24 hours. Administration of 50 μmole CdCl2/kg (sc) on day 12 also had no effect on fetal growth but resulted in increased fetolethality (12%). Thus midgestational cadmium exposure and its accompanying alterations in placental blood flow do not compromise fetal viability or growth. The differential response to cadmium at mid‐ and late gestation, in terms of fetolethality, is not due to maternal cadmium dose.
Developmental study of neural tube closure in a mouse stock with a high incidence of exencephaly
Authors: Macdonald K. B. , Juriloff D. M. , Harris M. J.
Published: June 2005

About 17% of embryos and fetuses in the SELH/Bc mouse stock have the anterior neural tube defect, excncephaly. No other malformations are seen. The genetic liability to exencephaly was shown to be probably genetically fixed in the SELH/Bc stock. This means that SELH/Bc embryos with successful neural tube closure are genetically the same as exencephalics. Females were significantly more likely to be affected than males (66% females).

The pattern of morphological development events during anterior neural tube closure on days 8 and 9 of gestation was compared among 322 ICR/Bc (normal), 304 SWV/Bc (normal), and 265 SELH/Bc embryos. Anterior neural tube closure was found to follow a strikingly different pattern in almost all SELH/Bc embryos than in either of the normal strains or in previous published studies. SELH/Bc embryos lack the initial contact between the anterior folds in the posterior prosencephalon/anterior mesencephalon region (Closure 2). In spite of this, all but 17% manage to close the anterior neural tube by extending caudally the later occurring normal anterior zone of contact and fusion at the most rostral aspect of the prosencephalon (Closure 3) through the region of Closure 2 to meet the zone of closure of the rhombencephalon, Closure 4. Anterior neural tube closure was completed late, and in some SELH/Bc embryos, elevation and fusion in the mesencephalon did not occur at all.

In histological sections of six‐ and eight‐somite embryos, elevated numbers of pyknotic cells in the neuroepithelium and mesenchyme, and elevated numbers of unstained inclusions in the neuroepithelium were found; but their relationship, if any, to the abnormal pattern of neural tube closure is not clear.
Effects of preconceptional and gestational exposure to tordon 202c on fetal growth and development in CD‐1 mice
Authors: Blakley Patricia M. , Kim Jin Suk , Firneisz Gay D.
Published: June 2005

Female CD‐1 mice were exposed to Tordon 202c (a picloram and 2,4‐D combination herbicide) in the drinking water at concentrations of 0.21, 0.42, and 0.84% for 60 days prior to mating with untreated males. One‐half of the pregnant females subsequently continued treatment throughout gestation while the remaining females were maintained on distilled water. Fetal weight, crown–rump length, placental weight, and maternal gestational weight gain were reduced in a dose‐dependent manner following combined preconceptional and gestational exposure. The incidence of malformed fetuses (cleft palate, renal agenesis, hydronephrosis, unilateral testicular agenesis, and umbilical hernia) and fetuses with variants (especially incomplete ossification of the skeleton) were increased in a dose‐dependent manner following combined exposure. Increased maternal mortality and decreased preconception weight gain were observed in the highest‐dosage group. Relative maternal liver weight was increased in a dose‐dependent manner. The results suggest that combined preconceptional and gestational exposure to Tordon 202c is required for teratogenesis and fetal growth depression. Preconceptional exposure alone is not effective in increasing the risk for embryotoxicity.
Effects of human diabetic serum on the in vitro development of mouse preimplantation embryos
Authors: Zusman Igor , Yaffe Perhija , Ornoy Asher
Published: June 2005

The effects of sera from different types of human diabetes (type I with and without ketoacidosis; type II treated with insulin or Daonil or untreated) on the in vitro development of early preimplantation mouse embryos were studied. In controls, 20% of blastocysts failed to develop successfully when grown for 72 h in RPMI medium supplemented with 10% fetal bovine serum and 50% nondiabetic human serum. In experiments using 50% diabetic serum, the highest embryotoxic effect was found in type‐I diabetes with and without ketoacidosis: The percents of undeveloped embryos were 66 and 58, respectively. In type‐II diabetes, embryotoxic effects were found among all studied types: The percent of undeveloped blastocysts varied from 36% in insulin‐treated type‐II diabetes to 44% in untreated type‐II diabetes. A high correlation was found between the number of undeveloped embryos and the blood concentrations of metabolic diabetic factors: glucose (r = .53–.64 in type‐I diabetes), B‐HOB (r = .7–.77 in type‐II diabetes untreated or treated with Daonil), acetoacetate (r = .66 in insulin‐treated type‐II diabetes), and HbA1c (r = .89 in insulin‐treated type‐II diabetes or .99 in Daonil‐treated type‐II diabetes). A concentration of 80% serum was embryotoxic when obtained from nondiabetic or from diabetic human. The possible role of diabetic metabolic factors in causing increased risk of spontaneous abortions and infertility among diabetic women is discussed.
Test of six chemicals for embryotoxicity using fetal mouse salivary glands in culture
Authors: Lyng R. Douglas
Published: June 2005

Many new chemicals come into use each year, and the need for rapid and cost‐effective methods for testing developmental toxicity is apparent. Establishing reliable in vitro techniques is important to a tier approach to testing for developmental toxicity. The fetal mouse salivary gland was selected as a possible test system because several interacting developmental processes occur in gland growth, and development is quantifiable by counting lobes. For each chemical tested, 20 glands from 13‐day embryos were treated in a control media and in three concentrations of the test chemicals. The number of lobes present after 48 hours is dependent on the number of lobes at explanation. Glands with different numbers of lobes at explantation were compared by dividing the number of lobes present after 48 hours by the number present at explantation to determine a growth ratio. Mean growth ratios were used to construct dose‐response curves, and from these curves the concentration that reduced growth by 50% (TP50) was determined. Comparisons with in vivo data were made by calculating three ratios; the TP50 was divided into the lowest teratogenic dose, the lowest maternal toxic dose, and the dose that was lethal to 50%. Four in vivo teratogens, 6‐aminonicotinamide, cytochalasin B, hydroxyurea, and 3‐acetylpyridine, all had ratios much higher than 1, indicating a very sensitive response by the glands. One in vivo teratogen, dexamethasone, had much lower ratios, indicating less sensitivity. Acetaminophen, a nonteratogen in vivo, actually stimulated growth of the glands at 10−5 M and had very low ratios indicating a minimal response by the glands.
Developmental alterations in offspring of female rats orally intoxicated by aluminum chloride or lactate during gestation
Authors: Bernuzzi Viviane , Desor Didier , Lehr Paul R.
Published: June 2005

The oral treatment of pregnant rats by aluminum chloride or lactate at various doses was applied from day 1 to day 21 of gestation to determine its influence on mortality, weight evolution, and neuromotor maturation of their pups. No effect of treatment on litter size was detected, but an increased mortality appeared during the first week: treatment by aluminum lactate was less active than was an equivalent treatment by aluminum chloride. Weight was transitorly delayed, but the reversal of this effect could be attributed to the decrease of litter size. The neuromotor maturation of surviving pups treated with the two aluminum salts showed an important impairment during the first 2 weeks of postnatal life.
Biomechanical basis of diazepam‐induced neural tube defects in early chick embryos: A morphometric study
Authors: Nagele R. G. , Bush K. T. , Hunter E. T. , Kosciuk M. C. , Lee H.
Published: June 2005

The biomechanical basis of diazepam (Valium/Roche)‐induced neural tube defects in the chick was investigated using a combination of electron microscopy and morphometry. Embryos at stage 8 (four‐somite stage) of development were explanted and grown for 6 hr in nutrient medium containing 400 μg/ml diazepam. Nearly 80% of these embryos exhibited neural tube defects that were most pronounced in the forming midbrain region and typified by a “relaxation” or “collapse” of neural folds. The hindbrain and spinal cord regions were less affected. Electron microscopy revealed that neuroepithelial cells in diazepam‐treated embryos had smoother apical surfaces and broader apical widths than did controls. Morphometric measurements supported this observation and further showed that these effects were focused at sites within the wall of the forming neural tube that typically exhibit the greatest degree of bending and apical constriction (i.e., the floor and midlateral walls). Overall results indicate that neural tube defects associated with exposure to diazepam are due largely to a general inhibition of the contractile activity of apical microfilament bundles in neuroepithelial cells. These findings 1) emphasize the important contribution of microfilament‐mediated apical constriction of neuroepithelial cells in providing the driving forces for bending of the neuroepithelium during neural tube formation and 2) suggest that agents or conditions that impair their contractile activity could play a role in the pathogenesis of certain types of neural tube defects.
Determinants of recall and recall bias in studying drug and chemical exposure in pregnancy
Authors: Feldman Yaacov , Koren Gideon , Mattice David , Shear Heather , Pellegrini Elizabeth , MacLeod Stuart M.
Published: June 2005

Case‐control studies on effects of durgs in pregnancy rely heavily on maternal recall. At the Motherisk Program in Toronto we counsel women during early pregnancy on the risk of drug and chemical exposure; subsequently, we follow up the outcome of pregnancy after birth. This cohort has given us an opportunity to assess the magnitude of recall of early pregnancy exposure and determinants likely to affect it in 145 consecutive cases. The mean recall of exposure identity was 62%, while accurate recall of timing of exposure was 37% and of dosage 24%. Exposures that prompted the clinic visit, chronic therapeutic exposures, environmental agents, and known teratogens were recalled significantly better than were other exposures. Accurate report of smoking was significantly higher than of alcohol use (79.4% vs. 59%, respectively, P = .0002). The number of agents consumed by the pregnant woman negatively correlated with her recall; mean recall of 1 agent was 85% vs. only 40% recall of 4 agents. Women ≥ 30 years of age recalled significantly worse (mean ± SEM, 52 ± 4%) than women younger than 30 (70 ± 4%), P = .002) despite a similar mean number of exposures. No difference in mean recall was found between women having normal (n = 112) or adverse pregnancy outcome (n = 33). There was a recall bias in reporting alcohol consumption; postnatally, women with adverse outcome tended to report significantly less than the amount initially reported by them.
Laminin immunized monkeys develop sera toxic to cultured rat embryos and fail to reproduce
Authors: Weeks Benjamin S. , Klein Norman W. , Kleinman Hynda , Fredrickson Timm , Sackett Gene P.
Published: June 2005

In a previous study antilaminin antibodies in a monkey with a poor reproductive history were found to be the cause of serum toxicity to cultured rat embryos. In the present study four monkeys were immunized with murine tumor laminin and a fifth with bovine serum albumin. Subsequently, sera from only the laminin immunized monkeys became toxic to cultured rat embryos. This serum toxicity was not mediated by complement but did require the antibody to have a divalent structure. Finally, mating trials were conducted with two of the laminin immunized monkeys ‐ that previously had excellent reproductive histories. Based on progesterone levels and observation the monkeys continued to have normal menstrual cycles but failed to initiate a successful pregnancy following immunization in over 2 years of mating trials. These data demonstrated that antibodies against laminin could have prevented conception or could have interrupted pregnancy because of embryotoxicity or failure of implantation.
Spinal ganglia reduction in the splotch‐delayed mouse neural tube defect mutant
Authors: Moase Connie E. , Trasler Daphne G.
Published: June 2005

Splotch and splotch‐delayed mutants have anomalies in certain neural crest cell derivatives as well as neural tube defects. A genetic marker was used to identify mutant, heterozygote, and wild‐type embryos within a litter, which enabled us to make intergenotypic comparisons. Histological studies of the lumbosacral region of day 15 and day 16 embryos indicated that the splotch‐delayed mutant had similar but less severe defects in spinal ganglion development than those reported for splotch (Auerbach: Journal of Experimental Zoology 127:305–329, 1954). The ganglia were extensively reduced in size, residual, or missing in the splotch‐delayed mutant, whereas in the splotch mutant, they were virtually nonexistent. Paired comparison analyses showed that all mutant embryos had a significant reduction in their volume of lumbosacral spinal ganglia when compared to their heterozygous and/or wild‐type littermates. Also, some heterozygotes were found to have spinal ganglia volumes that were significantly reduced when compared to wild‐type embryos. The volume of spinal ganglia was not related to the severity of the neural tube defect. In fact, three mutant embryos, which did not exhibit a neural tube defect, had spinal ganglia volumes comparable to or less than those mutants with open neural tube lesions or curly tails. This shows that the formation of abnormal neural crest cell derivatives is not a result of the neural tube closure defect. We hypothesize that the two anomalies observed in these mutants have a common etiological basis.
Morphological and physiological effects of β‐hydroxybutyrate on rat embryos grown in vitro at different stages
Authors: Moore D. C. P. , Stanisstreet M. , Clarke C. A.
Published: June 2005

Diabetic women are more likely to give birth to infants with congenital malformations than are nondiabetic women. Rodent embryos have been used as a model for the study of abnormal fetal development associated with maternal diabetes, and some of the metabolic factors which are altered in diabetes, such as raised glucose and ketones, have been shown to cause abnormal development of rodent embryos in vitro. The present work explores further the teratogenicity of β‐hydroxybutyrate to rat embryos. To determine the sensitivities of rat embryos at different stages of their development, rat embryos at 9.5 days of gestation have been cultured in vitro for 24 or 48 h, with or without 4 × 10−2 M β‐hydroxybutyrate for all or part of the culture period. The ebryos have been examined by scanning electron microscopy, and a detailed morphometric analysis of one tissue, the neuroepithelium, has been undertaken. The results confirm that β‐hydroxybutyrate causes abnormal development of rat embryos. The results of experiments in which embryos were exposed to β‐hydroxybutyrate for only part of a 48 h culture show that embryos expoosed to β‐hydroxybutyrate for a complete 48 h culture are more severely affected than embryos exposed to β‐hydroxybutyrate for only part of the culture and that embryos are more vulnerable to β‐hydroxybutyrate during the first half of a 48 h culture (equivalent to 9.5 to 10.5 days of gestation) than during the second half of a 48 h culture (10.5 to 11.5 days of gestation). The results of experiments in which embryos were cultured with β‐hydroxybutyrate from 9.5 days of gestation for 24 h (equivalent to 9.5 to 10.5 days of gestation) showed that some effects of β‐hydroxybutyrate are already apparent after 24 hours in culture. Many of the abnormalities produced by β‐hydroxybutyrate can be calssified as embryonic retardations rather than malformations—that is, embryos show features characteristic of normal, but younger, embryos. Embryos exposed to β‐hydroxybutyrate for the complete 48 h culture period consume less glucose and produce less lactate than control embryos on a per embryo basis, but not on a per μg protein basis, suggesting that the reduced metabolism is an effect of β‐hydroxybutyrate‐induced developmental delay rathen than a cause of it. Morphometric analyses of the shapes of the cells of the neuroepithelium show that the cells are less elongated/tapered in β‐hydroxybutyrate‐treated embryos than in control embryosd; these differences are characteristic of embryos in which the elevation of the neural fold has been inhibited. The mechanisms by which developmental retardation might lead to fetal malformation are discussed, with special reference to the possible of neural tube defects.
Anatomic findings in dicephalic conjoined twins: Implications for morphogenesis
Authors: Siebert Joseph R. , Machin Geoffrey A. , Sperber Geoffrey H.
Published: June 2005

The morphogenesis of conjoined twins is incompletely understood. We therefore conducted a postmortem study of dicephalus dibrachii dipus conjoined twins. The twins were born without pertinent history or prenatal diagnosis at 38 weeks and lived for several hours. External genitalia were female and partly duplicated; a caudal appendage was present in the thoracolumbar region. The heart and liver were shared and exhibited major abnormalities in configuration. Four lungs, three kidneys and adrenal glands, and two spleens were identified; biliary and upper gastrointestinal tracts appeared as mirror images. From these findings, we postulate three major sets of consequences arising from the anatomical disposition of the twin notochords (“paleoaxes”). 1) The degree of convergence/divergence of craniocaudal paleoaxes is variable. Convergences are maximal in the upper thoracic and sacral regions, where duplication of organs is minimal because of interaction aplasia. 2) In the horizontal plane, paleoaxes are sufficiently divergent to produce a degree of twin expression posteriorly, whereas anteriorly they converge to form a single, anterior, midline “neoaxis.” Interposed between these zones of paleoaxial and neoaxial expression are areas of variable interaction aplasia. 3) The left twin was in situs solitus; the right twin was in situs inversus in a manner resembling polysplenia.
Phenytoin embryotoxicity: Role of enzymatic bioactivation in a murine embryo culture model
Authors: Shanks Marion J. , Wiley Michael J. , Kubow Stanley , Wells Peter G.
Published: June 2005

A murine embryo culture model was developed to study the potential contribution of enzymatic bioactivation to the teratogenicity of phenytoin. To assess the relative embryonic and maternal contributions to bioactivation, embryos were cultured respectively alone or in the presence of an exogenous source of cytochromes P‐450 (P‐450), which are thought to bioactivate phenytoin to a teratogenic reactive intermediate. Embryological development from gestational day 9 to day 10 was assessed, and bioactivation was quantified by the irreversible binding of radiolabeled phenytoin to embryonic protein. Embryos cultured with phenytoin and an exogenous P‐450 bioactivating system showed a significant decrease in the incidence of turning and closure of the anterior neuropore, yolk sac diameter, and protein content as well as growth retardation. In the absence of an exogenous P‐450 system, phenytoin did not decrease the incidence of turning or anterior neuropore closure but did cause growth retardation and a lesser but significant reduction in yolk sac diameter and embryonic protein content. An exogenous P‐450 system enhanced the bioactivation of phenytoin, although significant activity also was detectable in embryos cultured without an exogenous bioactivating system. These results suggest that the embryo itself can enzymatically bioactivate embryotoxically significant amounts of phenytoin, and that bioactivation and embryotoxicity can be further enhanced, qualitatively and quantitatively, by an exogenous P‐450 system, implicating a possible maternal contribution to phenytoin teratogenicity.
Genetic analysis of the cause of exencephaly in the SELH/Bc mouse stock
Authors: Juriloff D. M. , Macdonald K. B , Harris M. J.
Published: June 2005

A new mouse stock, SELH/Bc, having a high liability to exencephaly has been developed. About 17% of SELH fetuses are exencephalic. The genetic cause of this exencephaly was investigated in a cross to a normal related ICR/Bc strain and in subsequent classical genetic crosses (F2, first and second backcrosses). The data were compared with a number of genetic models, including that of a single recessive mutation with 17% penetrance. The data did not fit single‐locus inheritance. The expectations from the multifactorial threshold model based on an underlying quantitative liability trait with additive inheritance were found to fit the data very well. The number of loci involved was estimated to be about two or three. About 70% of exencephalic SELH fetuses are female, and there is no overall deficiency of males. The relatively higher risk in females was constant across the genetic backgrounds in the experiment. In summary, the liability to exencephaly in SELH mice appears to be a multifactorial threshold trait, and it therefore resembles human neural tube defects in type of genetic etiology. SELH therefore may be a valuable animal model in the study of neural tube defects.
Piperidine alkaloid composition and relation to crooked calf disease‐inducing potential of Lupinus formosus
Authors: Keeler Richard F. , Panter Kip E.
Published: June 2005

A congenital deformity condition called crooked calf disease, of widespread occurrence in western North America, is known to be induced by maternal ingestion during gestation of certain members of the Lupinus genus containing the quinolizidine alkaloid teratogen anagyrine. Because some piperidine alkaloids from other sources induce a similar condition, we have investigated the alkaloid composition and teratogenicity of Lupinus formousus, reported by others to be low in quinolizidines but rich in the type of piperidine alkaloids that we have speculated would be teratogenic. GC/MS analysis of L. formosus showed seven major and nine minor components in the total alkaloid fraction. All seven major and nine minor components in the total alkaloid fraction. All seven major and five of the nine minor components, representing all but 3% of the fraction, were identified by mass spectrometric fragmentation patterns and GC retention times. They included several potentially teratogenic piperidine alkaloids (including a very large amount of ammodendrine), as well as several nonteratogenic quinolizidine alkaloids plus a trace (at nonteratogenic levels) of the known quinolizidine teratogen anagyrine. The plant induced severe crooked calf disease with limb, spinal, and palate involvement in experimental calves. The deformities are believed to have been induced by ammodendrine.
Preparation and developmental toxicity of monoclonal antibodies against rat visceral yolk sac antigens
Authors: Jensen Marcela , Lloyd John B. , Koszalka Thomas R. , Beckman David A. , Brent Robert L.
Published: June 2005

Thirty clones producing monoclonal antibodies (MCAs) to rat visceral yolk sac (VYS) antigens have been prepared. These MCAs localized by immunofluorescence in the VYS endoderm in vitro and were tested for developmental toxicity by intraperitioneal injection of ascites fluid into pregnant rats on day 9 of gestation. Five of the hybridomas produced MCAs that induced embryonic death, malformation, and growth retardation; the other MCAs had no developmental toxicity. Five MCAs, three teratogenic and two nonteratogenic, were tested for their ability to inhibit pinocytosis in the isolated day 17‐VYS. Only the teratogenic MCAs were inhibitory, providing further evidence for the hypothesis that teratogenic antibodies interfere with the nutritional supply to the embryo.
Teratogen susceptibility of XO mothers and XO embryos in mice
Authors: Endo Akira , Watanabe Toshiaki
Published: June 2005

We examined whether the chromosomal imbalance inherent in an XO constitution in mice is more susceptible to teratogenic influence of biotin deficiency using a newly established mouse colony with pure X monosomy. We hypothesized that XO mothers or XO embryos might be more susceptible to certain teratogens. Contrary to our expectation, the incidence of external malformations induced by biotin deficiency did not differ either between XX dams and XO dams or between XX fetuses and XO fetuses.
Etiology of retinoic acid‐induced cleft palate varies with the embryonic stage
Authors: Abbott B. D. , Harris M. W. , Birnbaum L. S.
Published: June 2005

Retinoic acid (RA) has been shown to be teratogenic in many species, and 13‐cis‐RA is teratogenic in humans. Exposure to RA during embryonic morphogenesis produced a variety of malformations including limb defects and cleft palate. The type and severity of malformation depended on the stage of development exposed. The purpose of this study was to compare the effects of RA exposure in vivo on different stages of palate development. These results were compared to effects observed after exposure in organ culture. The vehicle used in RA dosing was also shown to be a major factor in the incidence of RA‐induced cleft palate. For the in vivo studies, RA (100 mg/kg) in 10 ml corn oil/kg was given p.o. on gestation day (GD) 10 or 12, and the embryos were examined on GD 14 and 16. Exposure to RA in an oil:DMSO vehicle resulted in much higher incidences of cleft palate than were observed after dosing with RA in oil only. After exposure on GD 10, to RA, small palatal shelves formed which did not make contact and fuse on GD 14. The medial cells did not undergo programmed cell death. Instead, the medial cells differentiated into a stratified, squamous, oral‐like epithelium. The RA‐exposed medial cells did not incorporate 3H‐TdR on GD 14 or 16, but the cells expressed EGF receptors and bound 125I‐EGF. In contrast, RA‐induced clefting after exposure on GD 12 did not involve growth inhibition. Shelves of normal size formed and made contact, but because of altered medial cell differentiation did not fuse. Medial cells differentiated into a pseudostratified, ciliated, nasal‐like epithelium. This response was produced in vivo at exposure levels which produced cleft palate, and after exposure of palatal shelves to RA in vitro from GD 12–15. The medial cells exposed on GD 12 incorporated 3H‐TdR on GD 14, expressed EGF receptors, and bound 125I‐EGF. The responses to RA which lead to cleft palate differed after exposure on GD 10 or 12, and the pathways of differentiation which the medial cells followed depended on the developmental stage exposed.
Chloride transport in embryonic cells: Effect of ethanol and GABA
Authors: Zimmerman Ernest F. , Collins Michael
Published: June 2005

Ethanol and GABA (γ‐aminobutyric acid) and their interaction on 36Ci−influx were analyzed in cultured embryonic palate and limb mesenchymal cells in order to determine whether ethanol exerts its teratogenic action through a GABA receptor involved in embryogenesis. Cl−transport in secondary cultures of C57BL/6 palate mesenchymal cells was shown to consist of three systems including the electroneutral Cl−HCO3− exchange (50%) and Na+ /K+/Cl− cotransport (30%) pathways and the voltage‐dependent Cl− channel (20%). Treatment with DIDS (4,4′‐diisothiocyanostilbene 2,2′‐disulfonic acid) or SITS (4‐acetamido‐4′‐isocyano‐stilbene‐2,2′ disulfonic acid) in SWV palate cells inhibited the Cl−HCO3− exchange pathway, while treatment with DIDS and bumetanide inhibited both the exchange and cation cotransport pathways, the residual Cl− influx inferred to be the electrogenic pathway. Inhibition of Cl− transport by anthracene‐9‐carboxylic acid confirmed the presence of the electrogenic Cl− pathway. It was observed that the rate of Cl− transport was significantly greater in palate cells of C57BL/6 mice than those of SWV mice. Also the rate of Cl− transport was significantly greater in secondary cultures of palate cells from C57BL/6 mice than from primary cultures of limb cells from the same strain. No evidence could be obtained that ethanol (10 to 100 mM) or GABA (3 × 10−5 M) or their combination stimulated total Cl− influx in either C57BL/6 or SWV palate mesenchymal cells, putative voltage‐dependent Cl− influx in C57BL/6 palate cells, or total Cl− influx in primary cultures of C57BL/6 limb mesenchymal cells.
Maternal diabetes and cardiovascular malformations: Predominance of double outlet right ventricle and truncus arteriosus
Authors: Ferencz Charlotte , Rubin Judith D. , McCarter Robert J. , Clark Edward B.
Published: June 2005

Most studies on the relationship of maternal diabetes to cardiovascular malformations (CVM) have been prospective investigations of pregnancy outcome and therefore could not identify associations with rare cardiac lesions. The results of a retrospective study shed new light on the risks of specific cardiac defects in diabetic pregnancies.

The Baltimore‐Washington Infant Study, a population‐based case‐control investigation of CVM, provides information on maternal diabetes reported in personal interviews. Among 2259 mothers of cases, 35 (1.5%) reported diabetes present before pregnancy (called “overt”) and 95 (4.2%) reported diabetes only during pregnancy (called “gestational”). Among 2,801 mothers of controls, 14 (0.5%) had overt diabetes and 83 (3.0%) had gestational diabetes. Malformationspecific risks were expressed as odds ratios (OR) with 99.5% confidence intervals (CI). The strongest associations with overt maternal diabetes were found with double outlet right ventricle (OR 21.33; 99.5% CI 3.34, 136.26), and truncus arteriosus (OR 12.81; 99.5% CI 1.43, 114.64). No significant diagnosis‐specific associations were found with gestational diabetes. Non‐cardiac malformations were present in 23% of infants with CVM whose mothers had overt diabetes and in 26% of infants with CVM whose mother had gestational diabetes, in 32% of infants with CVM whose mothers did not have diabetes, and in 4% of controls. Double outlet right ventricle and truncus arteriosus are malformations dependent upon neural‐crest‐cell‐derived ectomesenchymal tissues; these are precisely the conotruncal abnormalities that result from experimental ablation of the neural crest in chick embryos. The association with diabetes suggests a further etiologic link between these two lesions.
Examination of a rodent limb bud micromass assay as a prescreen for developmental toxicity
Authors: Wise L. D. , Clark R. L. , Rundell J. O. , Robertson R. T.
Published: June 2005

The mouse limb bud micromass assay is one of many short‐term tests proposed as preliminary screens for potential developmental toxicity. Previous efforts to validate this assay have used too few “nonteratogens.” The purpose of this study was to examine additional compounds, most of which, based on the literature, were perceived to have low potential for developmental toxicity in vivo. In addition, a method of data analysis was sought that would identify selective developmental toxins in the micromass assay, i.e., those that are effective at dosages not maternally toxic. The concentration of each of 23 compounds that produced a 50% inhibition (IC50) of radiolabeled thymidine (T) and sulfate (S) incorporation was determined and used to calculate a T/S ratio. The T/S ratio may be a useful measure of developmental hazard, since T incorporation measures toxicity toward a general cell function (DNA synthesis) and S incorporation measures mainly toxicity toward a developmentally specific cell activity (chondroitin sulfate synthesis). All compounds tested produced T/S ratios of <2.0. Since 22 of these 23 compounds are classified as “nonteratogens” or nonselective developmental toxins in vivo, a low T/S ratio in this in vitro assay system may be capable of discriminating potential for developmental hazard in vivo.
Risk factors for birth defects: Data from the Atlanta Birth Defects Case‐Control Study
Authors: Erickson J. David
Published: June 2005

The Atlanta Birth Defects Case‐Control Study data comprises information obtained from interviews with parents of 4,900 babies born with major birth defects and with the parents of 3,000 babies born without defects. The source of cases is the Centers for Disease Control's Metropolitan Atlanta Congenital Defects Program; the case‐control study is population‐based. Birth defects are classified into 92 groups and cross‐tabulated by 105 exposure/risk factor variables; data from selected cross‐tabulations are presented. The associations of each of the 105 exposure variables with all types of defects combined are presented, as are the associations of each of the 92 defect groups with the specific exposure variable, maternal diabetes. These data can be used to evaluate hypotheses arising from other sources, and for the purpose of “generating” hypotheses. The data describing all 92 × 105 cross‐tabulations are available to other investigators on floppy disk; write to Chief, Birth Defects and Genetic Diseases Branch, Centers for Disease Control, Atlanta, Georgia 30333.
Exposure to ionizing radiation during pregnancy: Perception of teratogenic risk and outcome
Authors: Bentur Y. , Horlatsch N. , Koren G.
Published: June 2005

We quantified the perception of teratogenic risk in women attending the Motherisk program for counseling about diagnostic radiation in pregnancy (n = 50) and compared it with a control group of women exposed to nonteratogenic drugs and chemicals (n = 48). Before receiving known information about the specific exposure, women exposed to radiation assigned themselves a significantly higher teratogenic risk compared with the control group (25.5 ± 4.3% versus 15.7 ± 3.0% for major malformations, P < 0.01). The post‐consultation perception of teratogenic risk did not differ between the two groups. Special consideration and attention should be given when counseling pregnant women exposed to low‐dose ionizing radiation, as their misperception of teratogenic risk may lead them to unnecessary termination of their pregnancy.
Morphological differences elicited by two weak acids, retinoic and valproic, in rat embryos grown in vitro
Authors: Seegmiller Robert E. , Harris Craig , Luchtel Daniel L. , Juchau Mont R.
Published: June 2005

We compared in rat whole‐embryo culture the morphological changes elicited by valproic acid (VPA) with those elicited by trans‐retinoic acid (RA). Rat embryos explanted on day 9.5 of gestation were treated on day 10 with RA or VPA at concentrations producing equivalent reductions in embryonic protein. The concentrations selected for morphological assessment by scanning and transmission electron microscopy, 2.3 and 800 μM, respectively, for RA and VPA, produced approximately a 50% incidence of abnormally open anterior neuropores in initial range‐finding experiments in the culture system. Protein and DNA analyses were also performed on corresponding groups of embryos at three different doses. With concurrent control groups used as reference standards, the two treatment groups were compared for differences in external and internal morphology, protein and DNA contents, and growth indices. While certain variables responded similarly in the two treatment groups, e.g., the growth variables, protein and DNA contents, each drug produced selective morphological effects. Whereas treatment with RA produced underdeveloped branchial arches, symmetrically cleft cranial defects resulting in openings in rhombencephalic and prosencephalic regions, and exteriorized neural tissue in the caudal neuropore region, VPA produced irregular clefts with wavy margins along the entire length of the neural tube, and an open caudal neuropore without eversion of the neuroepithelium, while producing no detectable effect on the branchial arches. The similar effects of these two drugs on protein and DNA contents suggest comparable degrees of overall toxicity; however, the dissimilar effects on neural tube and branchial arches, coupled with the large difference in concentration of the drug required to produce the effects, add to the evidence that their mechanisms for elicitation of abnormal development are qualitatively different.
Effects of lysosomal proteinase inhibition on the development of the rat embryo in vitro
Authors: Daston George P. , Baines Donald , Yonker Julie E. , Lehman‐McKeeman Lois D.
Published: June 2005

Altered lysosomal function in the visceral yolk sac can result in abnormal development. As proteolysis is an important function of the rodent visceral yolk sac during early and mid‐gestation, we characterized the lysosomal proteolytic enzyme activity of this extraembryonic membrane and determined the effects of inhibitors of protein degradation on embryonic development. Constituent activities of cysteine and aspartic acid proteinases were measured in rat visceral yolk sac on gestation day 12, and the effects of the cysteine proteinase inhibitors leupeptin, E‐64 [trans‐epoxysuccinyl‐l‐leucylamido(4‐guanido)butane] and N‐ethylmaleimide and the aspartic acid proteinase inhibitor pepstatin were determined in Sprague–Dawley rat embryos cultured in vitro from gestation days 10–12. It was determined that only cysteine proteinases, primarily cathepsins B and L, are active in the mid‐gestation visceral yolk sac. The cysteine proteinase inhibitors leupeptin and E‐64 both produced a concentration‐related decrease in embryonic growth, as measured by crown–rump length, somite number, and embryonic protein content, and a concentration‐related increase in incidence of abnormalities. A characteristic pattern of abnormalities was produced which involved a decrease in neural tube volume and the formation of a subectodermal blister opposite the point of attachment of the vitelline vessels. At high concentrations, anophthalmia was also observed. The decreased neural tube volume was associated with increased osmolality of the exocoelomic fluid, the major extraembryonic fluid compartment. It is possible that the osmotic change decreased neural tube volume by causing water to move to the compartment with a higher solute concentration, out of the embryo. Leupeptin and E‐64 treatment increased yolk sac protein content, and produced histologically observable protein droplets (probably secondary lysosomes), indicating that lysosomal protein degration was inhibited. N‐Ethylmaleimide was a potent embryotoxin, but had none of the effects of the other cysteine proteinase inhibitors at concentrations which were not embryolethal. However, it should be noted that N‐ethylmaleimide was embryolethal at a concentration which is 1,000‐fold lower than that which inhibits cysteine proteinase activity in a lysosomal lysate. Pepstatin, an aspartic acid proteinase inhibitor, was without effect at any concentration tested (up to 300 μg/ml). These results indicate that inhibition of lysosomal proteolysis in the visceral yolk sac, specifically by cysteine proteinase inhibitors, is developmentally toxic to rats. The effects produced may be attributable both to lack of substrates (amino acids) for growth, and altered osmolality in the extraembryonic fluid compartment.
Effects of lithocholic acid exposure throughout pregnancy on late prenatal and early postnatal development in rats
Authors: Zimber A. , Zusman I. , Bentor R. , Pinus H.
Published: June 2005

The effects of lithocholic acid (LCA) on late prenatal and early postnatal development were studied. LCA was given in the diet (0.8% by weight) or administered in the drinking water (0.8%) throughout the entire period of pregnancy. The addition of LCA to the drinking water caused a significant reduction in the weight of 20‐day‐old fetuses as well as their placentas and livers. Feeding LCA to pregnant rats affected postnatal growth of the offspring: their weight was significantly lower than control values until 6 weeks of age. The pattern of postnatal growth was similar in the experimental and control rats. The retardation in growth and development of the offspring was accompanied by microscopic and biochemical changes in their livers.
Sensorineural hearing loss as evidenced by the auditory brainstem response following prenatal cocaine exposure in the long‐evans rat
Authors: Church M. W. , Overbeck G. W.
Published: June 2005

Prenatal cocaine exposure has been associated with a variety of adverse neurological effects. Three recent studies found evidence that prenatal cocaine exposure is associated with abnormal auditory electrophysiology, suggesting abnormal processing of auditory information. The present study used the auditory brainstem response to evaluate the effects of prenatal cocaine exposure on hearing in an animal model (Long‐Evans rat). We report that prenatal cocaine exposure can cause elevated ABR thresholds and latency‐intensity curves consistent with a recruitment‐type sensorineural hearing loss.
Selective staining methods for cartilage of rat fetal specimens previously treated with alizarin red S
Authors: Yamada Takashi
Published: June 2005

A convenient method for staining cartilage with several basic stains after alizarin red S staining of bone was investigated in rat fetuses. It was found that bromophenol blue was useful and effective for staining of the margin and center areas of cartilage, even in specimens stored in glycerin for over 10 years. The specimens were washed in running tap water for 1 hr, and subsequently were immersed in water or in 70% ethanol at pH 4 for 1 hr or longer. The specimens were then stained with 0.005% bromophenol blue in 40% ethanol adjusted to pH 4 for 2 hr, or with 0.001% bromophenol blue in 40% ethanol adjusted to pH 4 for 24 hr. Furthermore, the bromophenol blue stain color actually faded when the specimens were immersed in water or in 70% ethanol at pH 8. Descending order of the stain‐effective action on fetal rat cartilage for the basic stains tested was bromophenol blue, aniline blue, Evans blue, methyl violet, trypan blue, and water blue.
Marked increase in the teratogenicity of the combined administration of the industrial solvent 2‐methoxyethanol and radiofrequency radiation in rats
Authors: Nelson B. K. , Conover David L. , Brightwell W. Stephen , Shaw Peter B. , Werren Dwight , Edwards Richard M. , Lary Joseph M.
Published: June 2005

Limited published animal research reports synergistic teratogenic effects following combined hyperthermia (induced by elevated ambient temperature) and administration of chemical teratogens. Radiofrequency (RF) radiation is widely used in occupational environments. Since RF radiation also elevates the body temperature of, and is teratogenic to, exposed animals, concurrent RF radiation and chemical agent administration may enhance teratogenicity. The present exploratory study, consisting of preliminary dose‐finding studies and the primary study, was designed to investigate whether concurrent exposure of rats to RF radiation and the industrial solvent 2‐methoxyethanol (2ME) can enhance the developmental toxicity of either agent acting alone. Preliminary dose‐finding studies using small numbers of rats investigated the ability of various RF radiation conditions and doses of 2ME to produce external malformations (primarily of the paws) when administered on gestation day 13. Based on these preliminary studies, RF radiation exposure [sufficient to elevate rectal temperature to 42.0°C (4°C above normal for rats) for 30 min] and 2ME administration (150 mg/kg) were selected for the primary study. In the primary study, groups of 18 to 27 pregnant rats were administered RF radiation exposure and distilled water gavage, 2ME gavage and sham RF exposure, RF radiation exposure and 2ME gavage concurrently, or sham RF exposure and distilled water gavage. Pregnant rats were sacrificed on gestation day 20, and the offspring were examined for external malformations. Combined exposures enhanced the adverse effects produced by either experimental agent alone (no malformations were detected in the double sham group). Mean fetal malformations/litter increased from 14% after 2ME and sham RF (15/26 litters affected, with an average of 2 fetuses/litter malformed) and 30% after RF radiation and water gavage (10/18 litters affected, with an average of 4 fetuses/litter malformed), to 76% after the combined treatment (18/18 litters affected, with an average of 12 fetuses/litter malformed). In addition to a significant increase in the frequency of malformations, the severity of malformations also was enhanced by the combination treatment (on a relative severity ranking scale, the 2ME severity score was less than 1, the RF score was 3, and the combination score was 6). This study provided evidence of synergism between RF radiation and 2ME administration, but additional research will be required to characterize the extent of synergism between these two agents. Potential interactive effects between chemical and physical agents need to be investigated to determine the extent to which such interactions should impact occupational exposure standards.
Hypothermia: Teratogenic and protective effects on the development of mouse embryos in vitro
Authors: Smoak Ida W. , Sadler Thomas W.
Published: June 2005

Hypothermia often occurs in association with clinical conditions involving severe hypoglycemia, but its effect on embryonic development has not been well evaluated. Thus, the whole embryo culture method was used to expose day 9 (neurulating) and day 10 (early limb bud stage) mouse embryos to physiologic levels of hypothermia (35°C and 32°C) for 4 and 24 hr. Embryos were evaluated after 24 hours for growth and malformations and compared with controls grown at 37°C. Lactate production was measured in embryos cultured for 4 hr at 32°C and compared with those cultured at 37°C. A 4‐hr exposure to hypothermia produced little effect morphologically but reduced the rate of lactate production at both embryonic stages. A 24‐hr exposure to hypothermia at 35°C or 32°C produced growth retardation and dysmorphogenesis in embryos undergoing neurulation. Early limb bud stage embryos were less sensitive to this treatment, with growth retardation produced only at the lower temperature. Since hypothermia is commonly associated with severe hypoglycemia in cases of diabetic insulin overdose, day 9 (neurulating) mouse embryos were exposed concurrently to short periods of hypothermia and hypoglycemia and compared with embryos cultured in hypoglycemic medium at normal temperature. The results demonstrated that hypothermia partially protects embryos against the dysmorphogenic effects of hypoglycemia. A balance of metabolic rate and available substrate is discussed as a possible mechanism for this protective effect.