John M. Rogers

Significance of Supernumerary Ribs in Rodent Developmental Toxicity Studies: Postnatal Persistence in Rats and Mice

Pregnant Sprague-Dawley rats and Swiss-Webster mice were gavaged with bromoxynil at 15 and 96.4 mg/kg/day, respectively, on Days 6-15 of gestation. The frequency of supernumerary ribs (SNR), which are here defined as any degree of ossification lateral to the first lumbar vertebrae, was determined in fetuses at term and offspring on Postnatal Days 6, 20, and 40. Bromoxynil induced significant increases in the incidence of SNR in fetuses of both species. In rats, SNR occurred in 62% of treated fetuses as compared to 14% in controls; in mice these values were 45% and 11%, respectively. The postnatal incidence and persistence of SNR was species dependent. In the rat, postnatal SNR incidence in treated animals did not differ significantly from controls. In contrast, in mice the bromoxynil-induced elevated incidence of SNR persisted through Day 40 (42.3% in treated vs 0% in controls). Analysis of SNR was also done on the basis of their length (greater or less than 1/2 the length of the 13th rib). In the mouse, the incidence of smaller SNR was much lower on Day 40 as compared to Day 20; in contrast the incidence of larger SNR persisted through Day 40. In the rat, the incidence of larger SNR was too small to draw conclusions as to the postnatal fate of these structures. As in the mouse, however, the incidence of smaller SNR was significantly lower by Day 40. The significance of SNR in developmental toxicity remains problematic. The impact of this anomaly on animals is difficult to assess.(ABSTRACT TRUNCATED AT 250 WORDS)

Interspecies Comparisons of A/D Ratios: A/D Ratios Are Not Constant across Species

The hypothesis that the ratio of the adult (A) and developmental (D) toxicity of a chemical is constant across animal species has been proposed as the basis for identifying developmental hazards, both from traditional developmental toxicity screens using laboratory mammals and from alternative systems such as the coelenterate Hydra attenuata. The purpose of this study was to determine whether A/D ratios are constant across species. The developmental and adult toxicity of 14 chemicals was assessed in four phylogenetically different species. The chemicals tested were aminopterin, bromodeoxyuridine, cadmium chloride, caffeine, congo red, dinocap, dinoseb, diphenylhydantoin, epinephrine, ethylenethiourea, 2-methoxyethanol, mirex, all-trans-retinoic acid, and trypan blue. These chemicals are representative of a variety of toxic mechanisms and a range of potencies. Species used were the CD-1 mouse (Mus musculus), South African clawed frog (Xenopus laevis), fathead minnow (Pimephales promelas), and fruit fly (Drosophila melanogaster). The mouse is a commonly used model for developmental toxicity. The other species are known to be sensitive to mammalian toxicants and have well-studied embryologies. Mice were exposed to chemicals either po or by sc injection using a standard Segment II protocol in which pregnant mice are administered the test agent on a daily basis from Gestation Days 6 to 15, adult toxicity is evaluated during and after treatment, and developmental toxicity is evaluated in fetuses at term. The exposure duration spans the period of organ formation in the embryo. The other species were exposed to test agents for a developmentally comparable period. This was from blastulation (shortly after fertilization) to the free-swimming tadpole stage in Xenopus (4 days); from blastulation to the free-swimming fry stage in Pimephales (7 days); and for the entire larval period, the period of development of the imaginal discs, in Drosophila (6 days). Adults of each species were exposed to test agents for 4, 7, and 6 days, respectively. The route of exposure was via the water column in the two aquatic species and via the diet in Drosophila. Statistical lowest observed effect level (LOEL) and no observed effect level (NOEL) values were generated for adult and developmental toxicity in each species. A/D ratios were calculated using both LOEL and NOEL values.(ABSTRACT TRUNCATED AT 400 WORDS)

Plasma glucose and protein concentrations in rat fetuses and neonates exposed to cataractogenic doses of mirex

Mirex was administered to rats during gestation or the early postnatal period and the effects on blood chemistry were studied, especially with regard to changes which might play a role in the known cataractogenicity of mirex. In the prenatal study dams were intubated with 6 mg/kg/day mirex on Days 8 through 15 of gestation, and fetal blood samples were obtained on Days 18 and 20. For postnatal studies, litters were culled to eight pups at birth. Dams were intubated with 10 mg/kg/day mirex on Days 1 through 4 postpartum, and blood was drawn from pups at ages 6 through 14 days. Glucose determinations were done on a Beckman ASTRA 8 autoanalyzer. Protein determinations were done by the method of Lowry et al. (O.H. Lowry, N. J. Rosebrough , A. L. Farr, and R. J. Randall (1951). J. Biol. Chem. 193, 165-175.) Plasma glucose levels were decreased by over 40% in mirex-treated fetuses which developed cataracts. Postnatal exposure to mirex did not alter plasma glucose. Mean plasma protein concentrations were significantly lower in treated litters on Days 12 and 14 postpartum, and treated pups with cataracts on Day 14 were hypoproteinemic compared to treated pups without cataracts. Hypoproteinemia is a common factor related to cataractogenesis induced by either prenatal or postnatal mirex exposure, and may possibly be a causative factor. Although hypoglycemia may be a contributing factor in prenatal cataractogenesis, it does not seem to be implicated in postnatal cataractogenesis.

Postnatal mirex cataractogenesis in rats: lens cation balance, growth and histology.

The effects of lactational mirex exposure on postnatal lens cation balance, lens growth and histology were examined in this study. Pregnant Long Evans rats were allowed to give birth, and litters were culled to eight individuals. Dams were intubated with 10 mg kg-1 day-1 mirex on days 1-4 postpartum. Pups were killed at 6-14 days of age, and lenses were removed, examined for opacities, weighed, and assayed for K+ and Na+ concentrations. Cataracts occurred in 71.0% of all treated litters and 58.1% of all treated individuals. Histologically, lenses from treated pups showed anterior and posterior cortical vacuoles by 8 days of age, greatly swollen lens fibers by 10 days of age, and areas of cell degeneration by 12-14 days of age. Lens K+:Na+ ratios were lower in treated groups on days 8-14 after birth, in both clear and cataractous lenses, and cataractous lenses were significantly higher in water content. Lens:eye weight ratios were not affected. The results of this study indicate that mirex-induced postnatal cataracts are due to lens degeneration subsequent to fiber cells swelling.

Lower Court Application of the “Overruling Law” of Higher Courts

The obligation of a court to follow the law of a superior court is commonly taken to be stronger than the obligation of the higher court to respect its own precedent. The Supreme Court has recently asserted this stronger obligation in the most forceful terms. What follows is an attempt to demonstrate that this is wrong as a matter of policy and as a matter of law.

The effects of mirex on the neonatal rat lens in vitro, with a comparison to kepone

Lenses were removed from 12-day-old rats and incubated in medium containing 20 microM mirex. Lenses were removed from the medium at 2, 4, 6, and 8 h and assayed for K+ and Na+ concentrations and hydration. Mirex had no in vitro effect on these lens parameters compared to controls, and produced no changes in lens clarity. In contrast, incubation for 8 h in medium containing 20 microM kepone resulted in elevated lens Na+ and lower lens K+. Thus, cataracts induced by mirex in vivo are probably not due to a direct toxic effect on the lens.

Casimer T. Grabowski: 1927–2007

Casimer T. Grabowski, a Past President and Charter Member of the Teratology Society, passed away December 16, 2007, in Miami. He was 80 years old. Cas was responsible for pioneering research on the teratogenic effects of hypoxia. This work led him to describe a mode of action for abnormal development in which osmotic imbalances lead to vascular changes that are the underlying cause of malformations related to hypoxia or vasoconstriction, or both. This seminal work is still being built upon by others to this day: two of the symposia planned for the 2008 Teratology Society annual meeting are on subjects related to his early research and insights, The March of Dimes (which long supported Cas’s work) Symposium on Hypoxia and the Rare Diseases Symposium on Vascular Disruptive Defects. Cas was born and raised in the Polish-American community in the Fleet Avenue neighborhood of Cleveland, Ohio. He received his B.S. from Western Reserve University in 1950, and his Ph.D. from Johns Hopkins University in 1954, where he learned experimental embryology. He was an Instructor at the University of Pittsburgh from 1954–1960, when he became an Assistant Professor in the Department of Biology at the University of Miami. Cas spent the rest of his career at Miami, rising quickly to the rank of Full Professor, and served a stint as department chair. While Cas spent virtually all of his scientific career as a developmental biologist and teratologist, his first research project at Johns Hopkins was to describe the tarsal chemoreceptor of the blowfly. Cas did this work while on rotation in the lab of Vincent Dethier, who was interested in the feeding behavior of insects. Dethier had observed that a fly could be induced to extrude its proboscis when its feet were placed in contact with something edible. Cas’s description of the tarsal chemoreceptor provided a morphological basis for Dethier’s contention that flies tasted with their feet. The resulting paper by Grabowski and Dethier in the Journal of Morphology in 1954 has been cited hundreds of times and is considered foundational work in insect chemoreception. Cas’s research is described in Dethier’s book To Know a Fly, a wonderful popular account of how scientific research is done. Cas’s dissertation work was also groundbreaking. He transplanted Hensen’s node onto the blastoderm of chick embryos and demonstrated that the node is the avian equivalent of Spemann’s organizer region in amphibians. As any embryologist knows, Hensen’s node is extremely small and delicate, and it is not an easy feat to remove it, and only it, and then successfully transplant it. But Cas was good with his hands, an attribute that he put to good use not only in his research but in his hobbies, especially woodworking. Cas’s dexterity was especially noteworthy because one of his hands had been badly injured in an industrial accident in an aluminum foundry where he had worked while in school. His ability to do fine,