Studies on implantation traces in rats. II. Staining of cleared uteri, formation and distribution of implantation traces

We made an investigation of implantation traces in delivered rats. 1. Non-fixed rat uteri were immersed in 2%-NaOH solution for over one hour. The uteri were then cleared and the implantation traces were seen to be stained yellowish-brown. This staining method was convenient for observation of the implantation traces. 2. Stillbirth was induced by stabbing or crushing, either in some embryos on one day between the 6th and 12th day of gestation, or in all embryos after the 13th day of gestation. Also, abortion was induced by stabbing or crushing all embryos before the 10th day of gestation. 3. When live embryos existed in the uterine horns, abortion traces were not detected. 4. In cleared uteri stained with 2%-NaOH solution, abortion traces were observed as small globes, reddish-brown. Normal delivery traces were observed as large globes, yellowish-brown, covered with yellowish-white of agglomerate cells, while stillbirth traces appeared as middle-sized, orange or yellowish-brown masses. 5. Though the implantation traces in rats which had been delivered four times were arranged like beads, they were recognizable by NaOH stain as either old or new traces, the former appearing smaller without agglomerate cells and a little more brown in comparison with the latter. 6. As the implantation traces increased in the uterine horns they became distributed from the central region to the cervical end, and the mean space between them narrowed. The implantation traces occupying the ovarian end or central region of the uterine horns trendes smallest size or largest size. 7. The implantation traces were composed of cicatrical tissue, and the area they occupied did not show adhesion of the placenta. On the other hand, placenta adhering to the uterine wall proliferated, and formed new implantation traces which did not overlap the old traces after decollement of the placenta.     

Diazepam effect upon the microscopic structure of the mouse placenta.

CD-1 strain, female mice, aged 5 to 7 months, were mated with males of the same age. Females presenting vaginal plug were separated and randomly distributed in two groups to be treated from the 6th to 17th day of gestation. One group received single daily diazepam doses (2.7 mg/kg i.p.), the other, 0.9% saline in equivalent volumes. Females were killed on 18th day, the placentas removed and fixed in 10% formaldehyde, pH 7.3, dehydrated and embedded in paraplast; 3 microns thick sections were stained with hematoxylin-eosin and Weigert hematoxylin and analyzed under light microscopy. Placentas of the diazepam-treated females presented dilated chorion vessels and intervillous spaces. Trophoblastic cell nuclei presented chromatin in coarse granules, atypically distributed in the karyolymph, which had lesser staining affinity. Giant cells showed vacuolized cytoplasm and coarsely granulated chromatin. Results indicate that diazepam causes structural changes, possibly placental and fetal physiology.     

Paternal cyclophosphamide treatment causes postimplantation loss via inner cell mass-specific cell death.

Treatment of the father with the anticancer alkylating agent cyclophosphamide has negative effects on embryonic development in the rat. Four-week treatment of male rats with a low dose of cyclophosphamide causes a dramatic, dose-dependent increase in postimplantation death of the progeny. Several recent studies have indicated that the paternal genome is required for the development of the extraembryonic tissues. Thus, the purpose of this study was to determine which tissues of the implanting embryo were affected by paternal exposure to cyclophosphamide. Male Sprague-Dawley rats were given cyclophosphamide (6 mg/kg/day) or saline by gavage and bred to untreated female rats after 4 weeks of treatment. Pregnant female rats were killed on day 7 of gestation, and implantation sites were dissected from the uterus, fixed, embedded in Epon for semithin serial sectioning, and stained for subsequent light microscopy. Strikingly, many of the implantation sites of affected embryos sired by treated males displayed an apparently normal trophectoderm enclosing a region of dying cells, containing dark-stained pyknotic nuclei. Very few or no inner cell mass-derived embryonic cells were present in these implantation sites. Therefore, there is a selective death of inner cell mass-derived cells in day 7 implantation sites obtained from the progeny of cyclophosphamide-treated males. The results of this study suggest that treatment of the male with cyclophosphamide can affect paternal genes specifically required for development of the inner cell mass cells of the embryo, without an apparent effect on those genes required for normal trophectoderm.     

Studies of implantation traces in rats. III. Histological examination

We carried out a histological examination of the implantation traces in delivered rats. The implantation traces could be identified more than 500 days after delivery on the mesometrial side as black and brown spots. The implantation traces were recognizable as a cicatrix remaining in the parametrium, mesometrial triangle, which was formed by repair of injury caused by placental desquamation. In this area, metrial gland cells which were laid down through pregnancy were recognized for about two months after delivery. The implantation traces consisted of cicatrix tissue associated with collagen production and hemosiderin. It was possible to distinguish old and new traces by the size of the siderophile cells and by the degree of hemosiderin present. It was also possible to discriminate new traces as yellowish-brown areas, covered with a yellowish-white mass of degenerated metrial gland, in cleared uteri stained with 2% NaOH solution. Siderophile cells on the implantation traces were derived from giant cells which persisted around the peripheral region of the placental desquamation site, and these giant cells were considered to be identifiable with metrial gland cells. It was considered that formation of the cicatrix is essentially the same in abortion, stillbirth and normal delivery. However, it was found that the implantation traces had different histological appearances depending on the degree of injury to the endometrium and myometrium and time of placental desquamation. The iron content of the implantation traces corresponded quantitatively with the hemosiderin observed in the histological investigations. The iron content decreased rapidly up to 21 days after delivery, decreasing gradually thereafter. The iron in the implantation traces could, however, be analyzed quantitatively by atomic absorption spectroscopy until day 365 after delivery.     

Action of the phenothiazine derivative methophenazine on prenatal development in rats.

Single doses of 100-400 mg/kg or multiple doses of 10 or 50 mg/kg of the phenothiazine derivative methophenzaine were given per osto Wistar rats at various times on the 7th-14th days fo gestation and the fetuses examined near term. Results indicated that methophenazine was mainly embryolethal when administered on the 8th-11th days, and was teratogenic at later times, producing types of malformations that depended on the day of treatment, the most susceptible period being the 13th and 14th days of gestation. Teratogenicity occurred only when the dosages were highly toxic to the pregnant rats. Ribovlavin given ip on the 14th day significantly reduced the embryolethal but not the teratogenic action of methophenazine.     

Cyclophosphamide teratogenesis: evidence for compensatory responses to induced cellular toxicity

Cyclophosphamide (CP) administered ip to pregnant mice on day 10 of gestation (day of plug = day 0) is teratogenic (exencephaly, cleft palate, and limb malformations) at 20 mg/kg and embryolethal at higher doses. In the present study, CP was administered at 1, 5, 10, or 20 mg/kg on day 10 of gestation. Embryos were removed at 8 and 28 hr postdosing, and two embryos from each litter were immediately stained with Nile blue sulfate (NBS) to identify areas of cell death. The remaining embryos were frozen and forelimb buds subsequently removed for flow cytometric (FCM) analysis of the cellular DNA synthetic cycle. Additional litters were examined near term (day 17) for morphological abnormalities; these data were correlated with embryonic toxicity as detected by NBS staining and FCM analysis. Only the highest dose produced malformations. In marked contrast, a dose-related increase in the percentage of limb bud cells in the S (DNA synthetic) phase of the cell cycle was detectable at all doses. Inhibition of DNA synthesis was detected at all doses 8 hr post exposure and persisted through 28 hr for doses greater than or equal to 10 mg/kg. NBS staining indicated increased cell death in the alar plate of the neural tube 28 hr after exposure to 10 mg/kg CP and generally increased cell death in areas of rapid cell proliferation throughout the embryo at 20 mg/kg. The absence of an overt teratogenic response at dose levels that produced significant perturbation of the cell cycle indicates that a measure of embryonic damage can be compensated for or repaired. The implications of these findings for the existence of thresholds in developmental toxicity are discussed.     

Early postimplantation embryolethality in mice following in utero inhibition of adenosine deaminase with 2'-deoxycoformycin

Adenosine deaminase (ADA) catalyzes the hydrolytic deamination of adenosine (or 2'-deoxyadenosine) to inosine (or 2'-deoxyinosine). Previously, we have shown that ADA activity is subject to strong cell-specific developmental regulation in placental tissues of mice between days 6 and 11 of gestation (Knudsen et al.:Biology of Reproduction 39:937-951, 1988). In the present study, we examined the effects of intrauterine exposure to 2'-deoxycoformycin (dCF; pentostatin), a potent irreversible inhibitor of ADA, on early postimplantation development. Deoxycoformycin was administered to pregnant ICR mice as a single intraperitoneal injection at a dose of 5 mg/kg on one of days 6 through 11 of gestation (plug day 0). A marked increase in the incidence of implantation site resorptions was observed following treatment specifically on days 7 (61% resorbed) or 8 (78% resorbed). No effect was observed following treatment on days 6, 9, 10, or 11. ADA-immunoreactive protein was shown, by ABC-immunoperoxidase staining on days 7 or 8 of gestation, to be present at high levels in decidual cells of the antimesometrial region but at below-detectable levels in the embryo. Treatment of pregnant dams with dCF on day 7 produced a complete (greater than 99%) inhibition of ADA activity in the antimesometrial decidua by 30 min, induced excessive cell death in the prospective neural plate and primary mesenchyme of the trilaminar disc by 6 h, and arrested embryonic development at an early somite stage. These results suggest that the antimesometrial decidua plays a protective role in preventing an inappropriate accumulation of endogenous ADA substrates in the implantation site.     

Acute-phase protein profile during gestation and diestrous: proposal for an early pregnancy test in bitches

Early pregnancy diagnosis in bitches has special relevance for adequate medical assistance to assure normal gestation, diagnosis of abortion or embryonic resorption, undesirable pregnancy interruption and dog owners wishing to assure medical assistance during parturition and to program participation of females in dog shows. The aim of this study was to verify acute-phase protein profile variation as a consequence of generalized inflammatory reaction due to embryonic endometrial invasion and use alterations as a method for early pregnancy diagnosis. Also the relationship between hormonal status and acute-phase proteins concentrations was assessed. Weekly serum samples were collected from 20 non-pregnant (NP) bitches and 20 pregnant females (P) to determine levels of fibrinogen, haptoglobin, ceruloplasmin, seromucoid, glycoprotein, alpha(2) globulin, progesterone and estradiol-17beta. No correlation was found between the implantation sites formed (number of pups born) and the hepatic stimulus for the acute-phase protein production. The conclusions are that acute-phase proteins can be used as an early pregnancy test for bitches from the 3rd week of gestation (14th-21st day post LH peak) for haptoglobin assay (values above 112.42 mg/dl of HbCN binding capacity), from the 4th to the 6th week (21st-42nd day post LH peak) for ceruloplasmin (values above 12.76+/-5.29 U/l), from the 4th week (21st-28th day post LH peak) for glycoprotein (values above 13.67%) and from the 4th week of gestation (21st-28th day post LH peak) for alpha(2) globulin (values above 0.61 g/dl). Fibrinogen and seromucoid increased in the P group from the 5th to the 6th week, respectively, thus not being suitable as parameters for an early pregnancy diagnosis. Relationship between ceruloplasmin and estradiol-17beta and seromucoid and progesterone were verified. For the acute-phase protein test it is important to verify bitches' healthy condition and to assure the precise mating dates to avoid false-positive and -negative results, respectively.     

Demonstration of dominant lethal mutations in early mouse embryogenesis

Male mice of C57Bl/6Y strain were injected intraperitoneally with 2.5 and 5 mg/kg doses of thioTEPA. Males were mated to tetrahybrid CBWA females during the second week after the treatment. Embryonic mortality was studied by two methods: by standard dominant lethal method on the 15-17th day of pregnancy and cytologically on the 4th day. The rate of fertilization was not affected by thioTEPA. After treatment with 2.5 mg/kg of thioTEPA the frequency of induced dominant lethals was 89.8%; preimplantation losses were 78,5% in treated and 13,8% in control group. The cytological analysis revealed that preimplantation embryonic death is equal to 63,9%. The death of embryos before implantation occurred at 2-20 blastomere stages. After treatment with 5 mg/kg of thioTEPA all embryos died before implantation at 2-16 blastomere stages. It was demonstrated that dominant lethal method gave more complete estimation of dominant lethal frequency, and that cytological analysis is the correct estimation of preimplantation death. Thus the methods used supplement each other.     

Evaluation of developmental toxicity in rats exposed to the environmental estrogen bisphenol A during pregnancy.

Bisphenol A (BPA) is an essential component of epoxy resins used in the lacquer lining of metal food cans, as a component of polycarbonates, and in dental sealants. The present study was conducted in an attempt to evaluate the adverse effects of the environmental estrogen BPA on initiation and maintenance of pregnancy and embryofetal development after maternal exposure during the entire period of pregnancy in Sprague-Dawley rats. The test chemical was administered by gavage to mated females from days 1 to 20 of gestation (sperm in varginal lavage = day 0) at dose levels of 0, 100, 300, and 1000 mg/kg. All females were subjected to caesarean section on day 21 of gestation and their fetuses were examined for external, visceral and skeletal abnormalities. In the 1000 mg/kg group, significant toxic effects including abnormal clinical signs, decreased maternal body weight and body weight gain, and reduced food consumption were observed in pregnant rats. An increase in pregnancy failure was also found in the successfully mated females. In addition, increased number of embryonal deaths, increased postimplantation loss, reduced litter size and fetal body weight, and decreased number of fetal ossification centers of several skeletal districts were seen. On the contrary, no significant changes induced by BPA were detected in the number of corpora lutea and implantation sites and by fetal morphological examinations. In the 300 mg/kg group, suppressed maternal body weight and body weight gain, decreased food intake and reduced body weight of male fetuses were seen. There were no adverse signs of either maternal toxicity or developmental toxicity in the 100 mg/kg group. It was concluded that BPA administration during the entire period of pregnancy in rats produced pregnancy failure, pre- and postimplantation loss, fetal developmental delay and severe maternal toxicity, but no embryo-fetal dysmorphogenesis at an oral exposure level of 1000 mg/kg.     

Effects of (R)-deoxycoformycin (pentostatin) on intrauterine nucleoside catabolism and embryo viability in the pregnant mouse.

The viability of early mouse embryos is acutely sensitive to (R)-deoxycoformycin (pentostatin), a tight-binding inhibitor of adenosine deaminase (ADA). Previous studies have shown that a single 5-mg/kg dose on day 7 (plug = day 0) of gestation fully inhibits uteroplacental ADA activity within 0.5 h; causes massive cell death in the neural plate and primary mesenchyme by 6 h, major craniofacial anomalies by day 10, and resorption by day 12 (Knudsen et al., '89; Airhart et al., '91). The present study has examined further the developmental toxicity and early effects of this inhibitor on ADA metabolism. (R)-Deoxycoformycin was administered to pregnant CD-1 (ICR) mice as a single intraperitoneal dose of 0.5-10 mg/kg total body weight on days 6-11 of gestation. The major adverse effect, early resorption, was dose dependent and specific to day 7-8 exposure. Treatment with 5 mg/kg on day 7 resulted in 85% resorptions, 15% malformations, and a 24% reduction in mean fetal weight, whereas the same dose of (S)-deoxycoformycin had no effect. Levels of adenosine and 2'-deoxyadenosine, which are the endogenous substrates of ADA, were monitored in the embryo/decidual unit (E/D) by reversed-phase high-performance liquid chromatography (RP-HPLC). In response to the inhibitor, both nucleosides increased transiently in the antimesometrial compartment (antimesometrial decidua + embryo). Peak levels (Cmax) of adenosine and 2'-deoxyadenosine were dose dependent over the range tested (0.05-10 mg/kg). Exposure to 5 mg/kg on day 7 raised adenosine levels within 0.5 h to 42-fold over the basal level of 0.06 nmol/mg protein. There was an even stronger effect on 2'-deoxyadenosine levels, which were elevated 674-fold over the detection limit of 0.0005 nmol/mg protein. Direct exposure to the inhibitor in serum-free E/D culture produced similar results: 50 microM (R)-deoxycoformycin within 1 h raised adenosine levels 26-fold and 2'-deoxyadenosine levels 410-fold. In vivo studies also showed a general correlation between embryolethality and the length of adenine nucleoside pool expansion, apparent for exposure on day 7, 8, or 9 but not on day 6, suggesting that the embryo becomes sensitive to adenosine or 2'-deoxyadenosine once the neural plate has formed.     

Developmental toxicity evaluation of Bendectin in CD rats

Bendectin, composed of doxylamine succinate and pyridoxine HCl (1:1), is an antinauseant previously prescribed for nausea and vomiting during pregnancy. The present study examined the maternal and developmental effects of Bendectin (0, 200, 500, or 800 mg/kg/day, po) administered to timed-pregnant CD rats (36-41/group) during organogenesis (gestational days [gd] 6-15). At death (gd 20), all live fetuses were examined for external, visceral, and skeletal abnormalities. At 500 and 800 mg/kg/day, maternal toxicity included reduced food consumption during treatment and for the gestation period, increased water consumption in the posttreatment period, reduced weight gain during treatment, and sedation; water consumption was reduced during treatment and for the gestation period, and maternal mortality (17.1%) was observed only at the high dose. Developmental toxicity included reduced prenatal viability (800 mg/kg/day) and reduced fetal body weight/litter (500 and 800 mg/kg/day). In addition, reduced ossification of metacarpals (800 mg/kg/day), phalanges of the forelimbs (500 and 800 mg/kg/day), and of caudal vertebral centra (all doses) was observed. No increase in percent malformed live fetuses/litter was observed. The proportion of litters with one or more malformed fetuses was higher than vehicle controls only at 800 mg/kg/day, with short 13th rib (to which the test species is predisposed) as the predominant observation. By contrast, a positive control agent (nitrofen, 50 mg/kg/day, po, 14 dams) produced 85% malformed fetuses/litter with the predominant malformation being diaphragmatic hernia. In conclusion, the incidence of litters with one or more malformed fetuses was increased only at a dose of Bendectin which produced maternal mortality (17.1%) and other indices of maternal and developmental toxicity (see Discussion).     

Developmental toxicity of dichloroacetate in the rat.

Dichloroacetic acid (DCA) is a principal by-product of the chlorine disinfection of water containing humic and fulvic acids, and is also a drug of interest in the therapeutic management of metabolic disorders. The developmental effects of DCA were evaluated in the pregnant Long-Evans rat. In two separate studies, animals were dosed by oral intubation on gestation days 6-15 (plug = 0) with 0, 900, 1,400, 1,900 or 2,400 mg/kg/day and 0, 14, 140, or 400 mg/kg/day. The vehicle control was distilled water. Maternal observations included clinical signs, weight change, and gross evaluation of organ weights and uterine contents at necropsy (day 20). Corpora lutea were counted and uteri stained for implantation sites. Live fetuses were examined for external, skeletal, and soft tissue malformations. Seven dams died during treatment (1,400 mg 1/19, 1,900 mg 2/19, 2,400 mg 4/21), and maternal weight gain was reduced at all except the lowest treatment levels. Liver, spleen, and kidney weights increased in a dose-related manner. The mean percentage of resorbed implants per litter was significantly elevated at greater than or equal to 900 mg/kg/day. Live fetuses showed dose-dependent reductions in weight and length at doses above 140 mg/kg. Statistically significant frequencies of soft tissue malformations ranged from 2.6% (140 mg/kg) to 73% (2,400 mg/kg). These were principally in the cardiovascular system and predominantly comprised defects between the ascending aorta and the right ventricle. Skeletal malformations were not observed in significant numbers in any dose group.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of dexamethasone on implantation and pregnancy in albino rats

Administration of 3 mg/kg body weight of dexamethasone from day 1 or 3 to 7 of pregnancy did not prevent implantation in albino rats. But the same dose when administered from day 8 to 11 resulted in complete abortion / resorption in all rats. Administration of 2 mg / kg body weight of dexamethasone from day 8 to 11 of pregnancy held no effect on the foetal survival. The results indicate that a high dose of dexamethasone does not affect implantation but the same dose affects the more advanced stages of pregnancy.     

Analysis of the abortive action induced by prostaglandin F2 alpha in the mouse. Study of ova and their transplantation]

In mice PGF2 alpha injections (120 mg/kg from first to the 4th day) determines a high proportion of early abortions. The mechanism of this action has been investigated by transplantation of blastocysts removed from PGF2 alpha treated mice to pseudopregnant mice. The results of transplantation experiments indicate that the percentage of implantation is the same whether the eggs are taken from PFG2alpha treated or control mice. The number of eggs present in the uterine horns 84 hours after fertilization is 40 % lower than in control animals. At an earlier stage, 60 hours after fertilization, the uterus of PGF2 alpha treated females contains 85 % of eggs as compared to 20 % in controls. This proportion is reversed in the fallopian tubes. It can therefore be concluded that the high abortion rate of PGF 2 alpha treated mice is not due to an embryotoxic effect but to a faster transport of the fertilized eggs which reach therefore the uterus before implantation can take place.     

Immunocytochemical studies of relaxin in ovaries of pregnant and cycling mice

Immunocytochemical staining for relaxin in ovarian sections of pregnant mice from day 11 through day 18 of gestation revealed that only corpora lutea (CL) of pregnancy are stained. Evaluation of serial sections of ovaries from a day 16 pregnant mouse revealed that the only luteal structures present are CL of pregnancy. The number of CL present in each ovary equaled the number of implantation sites in each related horn (7 on the right side and 8 on the left side). These large CL varied in shape, being round in some profiles to very elongate in others. All CL were immunochemically stained for relaxin using the peroxidase-antiperoxidase method of L. Sternberger (Immunocytochemistry, 2nd ed. Wiley, New York, 1979). The intensity of the strain varied from cell to cell within each CL. Small luteal structures that were observed to be immunochemically stained for relaxin were demonstrated to represent the periphery of CL of pregnancy. No luteinized follicles were observed and interstitial cells and follicles were not immunochemically stained in any of the day 16 serial ovarian sections or in any of the ovarian sections from pregnant mice on the other days of gestation studied. CL of previous cycles were not observed to be present in the ovaries at days 15, 16, or 18 of gestation. However on day 14 and before, CL of previous cycles were observed and they did not exhibit any relaxin immunostaining. Immunocytochemical studies using the biotin-avidin system revealed that no relaxin immunostaining could be demonstrated in the ovaries of cycling mice at any stage of the estrous cycle. In conclusion, this study revealed that the only ovarian structures demonstrating relaxin immunocytochemical staining in the mouse were CL of pregnancy.     

Perinatal death and respiratory apparatus dysgenesis due to a bis (dichloroacetyl) diamine

N, N1-bis (dichloroacetyl) diamine 1, 8-octomethylenediamine (WIN 18,446) is an experimental drug which was first investigated as a male contraceptive. It is soluble in lipid solvents but not in water. The administration of 1,200 to 1,600 mg/kg to pregnant rats on the tenth day of gestation produced multiorgan fetal malformations. Smaller doses, 400 to 800 mg/kg, especially if divided over 2 or 3 days, caused perinatal death. Thus, 60 to 100% of offspring of rats given WIN 18,446 on the tenth and 11th days of gestation died at birth or within 4 days (Taleporos et al., 78). The present study investigated such deaths. At doses of 200 mg/kg on day 10 or 50 mg/kg on days 10 and 11, 67% of offspring had defective or absent diaphragms, 48% had tracheobronchiomegaly with cystic lungs, and 67% had pleural hemorrhage. At doses of 100 mg/kg given on 1 day or 25 mg/kg each day for 2 days, 50% had tracheobronchiomegaly with cystic lungs and rudimentary acini. At lower doses (18.8 mg/kg X 2 or 12.4 mg/kg X 3), a majority of fetal lungs had rudimentary acini, thick septa, few capillaries, and wide cuffs of perivascular connective tissue. Thus, a chemical given during organogenesis produced dysgenesis of the respiratory apparatus. Varying the dose produced malformed lungs with persistently deficient acini which model such human lung faults as tracheobronchiomegaly (Mournier-Kuhn Syndrome; Mounier-Kuhn, '32), bronchiolar dysplasia (Wilson-Mikity Syndrome), and perinatal death with acinar failure resembling neonatal hyaline membrane disease.     

Salicylate-induced teratogenesis in the ferret

A single subcutaneous injection of 400 mg/kg sodium salicylate produced a high resorption rate on day 13 (91%) and on day 18 (66%) of gestation. Malformations were seen in the surviving fetuses. Pregnant ferrets injected with 250 mg/kg salicylate produced a lower resorption rate of between 31% and 43%. Malformations were seen in the surviving fetuses of animals injected with lower doses of sodium salicylate both at 13 and 18 days of gestation.Salicylate-induced teratogenicity at 400 mg/kg was compared with that produced in a closed colony of Wistar rats. The concentration of salicylate in whole blood (and serum) was determined after a single injection of 125 mg/kg or 400 mg/kg sodium salicylate. Although salicylate concentration in the blood in both species showed remarkable similarity at the doses tested and the times of sampling, the results indicated that the drug was far more embryo-toxic in ferrets than in rats. The inter-order variation in the embryotoxicity of sodium salicylate is such that it would be unwise to ignore its possible teratogenic activity in man.     

Developmental toxicity of bropirimine in rats after oral administration

Timed-pregnant Upj:TUC(SD)spf (Sprague-Dawley) rats were orally (gastric intubation) dosed with bropirimine (an immunomodulator and inducer of interferon with antiviral and antitumor activities against experimental models) at 100, 200 or 400 mg/kg/day (first experiment), or at 25, 50, or 100 mg/kg/day (second experiment), on days 7-15 of gestation. In the first experiment, maternal toxicity occurred in all bropirimine-treated groups as evidenced primarily by significant decreases in weight gain, as compared to the vehicle control group. Embryotoxicity also occurred as evidenced by a dose-related increase in the number of dams with early implantation sites only. This pronounced effect on early embryonic development led to an insufficient number of offspring to access the developmental toxicity of bropirimine. This effect and the fact that all three doses were toxic to the dams dictated that a second experiment be carried out at lower doses. Significant effects on maternal weight gain also were observed in the second experiment, at least in the first 4 days of dosing, although only one dam in the 100 mg/kg/day group had early implantation sites only, in contrast to 11 such dams at this dosage in the first experiment. However, the fact that there were significant dose-related increases in the incidence of several variations in fetuses in this group indicated that there also was embryotoxicity at 100 mg/kg/day in the second experiment. Thus, although no biologically significant increases in the incidence of any malformation or major variation were found in this study, the results did indicate that bropirimine was embryotoxic at dosages which also produced significant maternal toxicity.     

Development of spontaneous motility in chick embryos. Interaction of strychnine, glycine and GABA.

The interaction of strychnine (1 mg/kg egg weight), glycine (100 mg/kg egg weight) and GABA (103 mg/kg egg weight) on spontaneous motor activity recorded by the method of Kovach (1970) in intact eggs was studied in chick embryos from the 11th to 21st day of incubation. In 11- and 13-day embryos, neither of the amino acids influenced strychnine activation of spontaneous motility. From the 15th incubation day, strychnine activation was distinctly affected by both amino acids, but the maximum effect was observed on the 19th day. Glycine had a stronger inhibitory effect, since it prevented strychnine convulsions from developing, whereas GABA only modified them. It can be concluded from the results that glycine-sensitive and GABA-sensitive mechanisms of embryonal spontaneous motility do not begin to take effect in chick embryos until the 15th day of incubation.