Fetal toxicity of aminoguanidine in mice and rats

It has been reported that aminoguanidine (AG) sulfate injected into hen's eggs at the earlier incubation stages induced retardation of development of body and liver, aplasia of gallbladder and enlarged spleen in high frequency without high mortality. The present study deals with the influence of AG against rodent animals and their fetuses. AG sulfate showed low lethality at the intraperitoneal administration against young mice and rats. When the maximum tolerated dose of AG sulfate (750 mg/kg in mice and 500 mg/kg in rats) was intraperitoneally injected into pregnant mice and rats on day 0 to 6 or day 7 to 13 of gestation, the incidence of resorption was comparatively high in the day 7 to 13 group of both species, but severe abnormalities as observed in the chick embryos and other external abnormalities were not induced in rodent fetuses. 14C-AG hydrochloride intraperitoneally injected to pregnant mice on day 12 was widely distributed in the maternal body and fetuses within 1 hr and rapidly excreted. Furthermore, AG might be hardly metabolized in mice, while a metabolite clearly appeared in chick embryos. Therefore, the lower toxicity of AG on rodent animals as compared with chick embryos may be due to the fact that the agent is not metabolized and rapidly excreted from maternal body and fetuses.     

Influence of hypothyroidism induced by thiamazole on the toxicity of amitriptyline in chick embryos

The effect of hypothyroidism induced by thiamazole on the toxicity of amitriptyline was studied in chick embryos. Fertilized eggs of White Leghorns were incubated and investigated. 1.2 mg/0.2 ml/egg of thiamazole was injected into the albumen of fertilized eggs on the 9th day of incubation. The control group was given 0.2 ml/egg of physiological saline in the same manner. Amitriptyline at 1 mg/egg was injected into the air sac of fertilized eggs on the 16th day of incubation. Electrocardiograms were recorded 0 to 60 min after the injection. After the injection of amitriptyline into the thiamazole-treated eggs, the heart rate was significantly decreased compared with the untreated eggs. These findings indicate that hypothyroidism induced by thiamazole has a marked influence on the toxicity of amitriptyline in chick embryos.     

Influence of hypothyroidism induced by thiamazole on the toxic interaction between propranolol and disopyramide in chick embryos

The effect of the hypothyroidism induced by thiamazole on toxic interactions between propranolol and disopyramide were studied in chick embryos. Fertilized eggs of White Leghorns were incubated and investigated. 1.2 mg/0.2 ml/egg of thiamazole was injected into the albumen of fertilized eggs on the 9th day of incubation. The control group was given 0.2 ml/egg of physiological saline in the same manner. Propranolol at 0.1 mg/egg and disopyramide at 0.3 mg/egg were injected into the air sac of fertilized eggs on the 16th day of incubation. Electrocardiograms were recorded 0 to 60 min after the injection. After the injection of propranolol and disopyramide into the thiamazole treated eggs, the heart rate was significantly decreased compared with the thiamazole untreated eggs. These findings indicate that hypothyroidism induced by thiamazole has a marked influence on the toxic interaction between propranolol and disopyramide in chick embryos.     

Toxicity and distribution of 2-methyl-4-chlorophenoxyacetic acid (MCPA) in developing chick embryos

The toxicity of the herbicide Erbitox E30, a commercial formulation of 2-methyl-4-chlorophenoxyacetic acid (MCPA) containing 28% MCPA as sodium-potassium salt and 72% of unknown ingredients, was tested on chick embryos. Sterile aqueous solutions of MCPA were injected into the air chamber at doses of 0, 1.5, 3.0, 6.0, 9.0, or 10.5 mg/egg on day 0 or on day 4 of incubation. The mortality rate for the embryos treated on day 0 of incubation was high in the first 5 days, low from 5-12 days and again increased by 15 days. The 15-day LD50 was 4.4 mg/egg (95% C.I. 3.7-5.3 mg/egg). HPLC analysis of albumen and yolk showed that concentrations of MCPA in the albumen were detectable at 5 min, highest at 7 days and markedly diminished by 14 days of incubation; a significantly lower concentration of MCPA was found in the yolk throughout the incubation period, except at 14 days when the yolk concentration was 4 times higher than the albumen concentration. At 15 days of incubation, MCPA was evenly distributed in the tissues of the embryo. MCPA was more toxic to 4-day embryos; concentrations above 6.0 mg/egg were lethal to all embryos within the first week of incubation. The 15-day LD50 for treatment on day 4 of incubation was 2.8 mg/egg (95% C.I. 2.5-3.2 mg/egg). The liver was affected by treatment with MCPA, being green in treated embryos. However, histological examination revealed few changes in the liver parenchyma.     

Experimental diabetes model in chick embryos treated with streptozotocin

The aim of the present study was to investigate whether diabetes model can be made by treatment of streptozotocin (STZ) in chick embryos and this model can be used to predict the effect of drug. When STZ (0.3 mg/egg) was injected into the albumen of fertile eggs on the 14th day of incubation, level of blood glucose significantly increased than that of the control on the 17th day of incubation, and level of serum insulin significantly decreased. In addition, the enhanced level of blood glucose in STZ-treated embryos reduced by injection of human insulin. In conclusion, STZ-treated embryos may be applicable to evaluate human insulin and anti-diabetes drugs as an experimental diabetes model.     

Development and validation of a herring gull embryo toxicokinetic model for PCBs

A toxicokinetic model was developed to describe polychlorinated biphenyl (PCB) accumulation by herring gull (Larus argentatus) embryos during development. The model consists of two components, a bioenergetics model that predicts the lipid mass balance of embryo and yolk compartments with time and an empirical toxicokinetic model that describes PCB partitioning between lipid compartments in the egg. The model was calibrated using data on PCB and lipid partitioning between embryo and yolk+albumen at four time points during incubation in herring gull eggs injected with a PCB mixture, combined with data sets on herring gull embryo growth rates and bioenergetic demands with time. The model was validated using independent data consisting of maternally exposed, field-incubated Lake Superior herring gull eggs that varied in incubation ages over the range of 8.5 d to pipping age (26–28 days). PCB concentrations in 6–9 d embryos were nearly an order of magnitude less than predicted by equilibrium lipid partitioning between the embryo and yolk+albumen compartments of the eggs. PCB concentrations in embryos were adequately predicted by equilibrium partitioning, however, for eggs incubated for 23–28 d. An empirical relationship was developed to account for the apparent non-equilibrium behaviour of PCBs during early development. The model was sensitive to the mass of yolk lipids and the mass of PCBs deposited to fresh eggs and much of the variability in embryo PCB concentrations could by explained by accounting for variability in these input parameters. Consistent with experimental data for other avian species, the model predicts that the highest PCB concentrations in the embryo/chick occur during pipping or soon after when yolk lipids have been completely resorbed by the embryo.     

Effect of pineal indoles on the chick embryo

In a study on the embryotoxicity of pineal indoles on developing chick embryos in vivo, the pineal indoles--namely, melatonin (MEL), methoxytryptamine (MTA) and methoxytryptophol (MTP)--were injected into the yolk sacs of the chick embryos through the air chambers of the eggs on the 4th day of incubation. The eggs were opened and the embryos examined after 6, 10 or 14/15 days of incubation. Abnormalities were found to occur mainly in the 6- and 10-day-old embryos, which exhibited external malformations such as twisted vertebral column, abdominal hernia, exteriorization of heart and viscera, defects of eye, beak and limb. From the results obtained from embryos on the 14th or 15th day of incubation, MEL was found to be the most toxic indole in regard to the mortality induced, whereas MTA had the highest teratogenicity because of the frequent incidence of abnormal embryos. Effect of MTP treatment on the development of chick embryos varied greatly between doses, and there were no abnormal embryos found on the 14th or 15th day of incubation.     

Histological studies on developing chick embryos treated with aminoguanidine sulfate

Fertile eggs were injected 10 or 20 mg/egg of aminoguanidine sulfate (AGS) into the albumen on the 5th day of incubation and examined macroscopically and histopathologically. The marked abnormalities were observed in liver, gallbladder and spleen while other organs were not different from the control. The earliest change was an increase of mitotic cells in the liver which reached to the peak at 2 days after the AGS injection. The increase of the ratio of cells in metaphase was characteristic. Decrease of weight, retardation of the development, fatty degeneration, necrosis and increase of connective tissues in the liver were followed. The abnormality of gallbladder such as aplasia and hypoplasia was observed with high frequency. Spleens of the AGS-treated group were enlarged from the 16th day of incubation. The clearest change of spleen was fatty degeneration.     

Effect of simultaneous exposure to nickel chloride and benzo(a)pyrene on developing chick embryos

In the present investigation the effect of Benzo(a)pyrene (BP) and Nickel chloride, quite often identified in crude and refined or waste oil, when injected in combination, was investigated on developing chick embryos after exposing them through yolk sac route on 6th day of incubation. Exposure to this combination in different doses resulted in no new deformities other than those experienced in BP treated chick embryos. The mortality and malformations experienced in the chick embryos exposed to BP alone and NiCl2 alone were seen to be added when both the chemicals were simultaneously inoculated into the same chick eggs.     

Cardiotoxicity of trastuzumab (herceptin) in chick embryos

The cardiac toxicity of trastuzumab was studied in chick embryos. Fertilized eggs of White Leghorns were incubated and investigated. Trastuzumab 5 mg/egg (low dose) or 15 mg/egg (high dose) was injected into the air sac of a fertilized egg on the 16th day of incubation. Electrocardiograms (ECGs) were recorded 0 to 60 min after the injection. After low dosing of trastuzumab, the heart rate was not different compared with the control. However, the heart rate was significantly decreased by high dosing of trastuzumab. In addition, arrhythmia was produced by high dosing of trastuzumab. These findings indicate that trastuzumab has a marked dose- and time-dependent influence on the heart rate in chick embryos.     

Influence of the light schedule on the toxic interaction between propranolol and disopyramide in chick embryos

The effect of the light schedule on toxic interactions between propranolol and disopyramide were studied in chick embryos. Fertilized eggs of White Leghorns were incubated under dark conditions and investigated, on two occasions, under light conditions or under dark conditions. Propranolol, with and without disopyramide, was injected into the air sac of fertilized eggs on the 16th day of incubation. Electrocardiograms (ECGs) were recorded 0 to 60 min after the injection. After the injection of propranolol with disopyramide, the heart rate was significantly decreased compared with the injection of propranolol alone under light conditions. In addition, this toxic interaction between propranolol and disopyramide was more severe under dark conditions than under light conditions. These findings indicate that manipulation of the light schedule has a marked influence on the toxic interaction between propranolol and disopyramide in chick embryos.     

Drug withdrawal prior to hatch in the morphine tolerant chick embryo

Morphine tolerance and dependence were produced in chick embryos by injecting 20 mg/kg into the air space of the egg daily from incubation day 12. Starting on day 16, two groups of eggs were withdrawn from morphine by either substituting water or by treating with naloxone. Chick embryo activities were monitored on incubation day 18. Activities of the embryos withdrawn from morphine did not differ from control, but naloxone injection produced a substantial increase in activity. Neither regimen of morphine withdrawal affected the hatchability of the eggs. When these chicks were 1 day old, the effect of 1 mg morphine/kg on the rate of distress vocalization was measured. The withdrawn chicks vocalized at a rate that was intermediate between that of controls and that of nonwithdrawn chicks indicating that tolerance was still present one week after the last morphine injection.     

Influence of the light schedule on the toxicity of amitriptyline in chick embryos

The cardiac toxicity of amitriptyline and the effect of the light schedule on it were studied in chick embryos. Fertilized eggs of White Leghorns were incubated under dark conditions and investigated, on two occasions, in the light phase and in the dark phase. Amitriptyline was injected into the air sac of fertilized eggs on the 16th day of incubation. Electrocardiograms were recorded 0 to 60 min after the injection. After the administration of amitriptyline 1 mg/egg in the light phase, the heart rate did not differ compared with that in controls. However, the heart rate was significantly decreased by the administration of amitriptyline 2.5 mg/egg and 5 mg/egg in the light phase. The heart rate was significantly decreased by the administration of amitriptyline 1 mg/egg under dark conditions. In addition, arrhythmia was produced by the administration of amitriptyline under dark conditions. These findings indicate that manipulation of the light schedule has a marked influence on the toxicity of amitriptyline in chick embryos.     

Antagonistic effect of zinc in lead treated developing chick embryos

Lead (50 micrograms) and zinc (100 & 200 micrograms) alone and in combination, were injected on 7th day of incubation into the yolk sac of developing chick embryos. The deformities induced by Pb alone included reduced hatchability, growth retardation, hydrocephalus, defective beak and legs, microphthalmia, anophthalmia and curling of hair. The concurrent exposure with Zn (200 micrograms), however, provided a remarkable protection against lead induced deformities, reflected in terms of almost normal development of chick embryos with increased hatchability and lower incidences of malformations, and mortality.     

Development of opiate tolerance in the chick embyro

Tolerance to morphine was produced in the chick embryo. Eggs were injected with morphine sulfate (MS) (20 mg/kg egg) or H₂O daily starting on incubation day 12. On day 16, embryo activities were recorded and eggs were injected with either MS or naloxone. Activity of H₂O-pretreated controls decreased after both MS and naloxone. Embryos treated with MS from incubation days 12–15 showed no activity change after morphine and responded to naloxone with increases in activity. Baseline rates of distress vocalizations (DV) of 1–2 day old chicks were not affected by MS pretreatment during incubation days 12–19. However, 1 mg/kg MS decreased the rate of DV of control chicks by 90% whereas MS-pretreated chicks were unaffected. At age 4–5 days, the baseline rate of DV and rate after MS were higher in MS-pretreated chicks. However, all chicks showed significant decreases in rate of DV after MS injection. Naloxone increased the rate of DV of paired 1–2 day old chicks, but response of MS-pretreated chicks was significantly greater than controls.     

Abnormal embryogenesis induced by thiopental

The effects of thiopental on chick embryos were analyzed in the present study. Thiopental was dissolved in saline and injected into embryonating chicken eggs at doses ranging from 0.2 to 4.0 mg per egg. The injections were made into the air sacs of eggs after two to four days of incubation. Control eggs were injected with an equivalent volume of saline (0.1 ml per egg). In all 1080 chicken eggs were used for this study. All embryos were examined on day 7. The LD50 for eggs injected on days 2, 3 and 4 was 2.1, 1.9, and 4.1 mg per egg, respectively. The principal malformations observed were exencephaly, anencephaly, twisted limbs, twisted neck, microphthalmia, everted viscera, and hemorrhage above the left eye and in both cerebral hemispheres. The results of the present study indicate that thiopental has a tendency to cause malformations in the chick embryos tested.     

Protective role of calcium in lead toxicity during development of chick embryo

In the present study the interaction of lead (Pb) and calcium (Ca) during embryogenesis was investigated. Lead (50 micrograms) alone and in combination with Ca (100 micrograms, 500 micrograms and 1000 micrograms) was injected on seventh day of incubation into the yolk sac of developing chick embryos. Treatment with lead alone caused stunted growth in chick embryos with several deformities e.g. defective beak and legs, hydrocephalus, microphthalmia and anophthalmia. The combined treatment of Pb with Ca showed a remarkable protective effect against lead induced deformities characterized by development of chick embryos with a low incidence of deformities.     

Teratism induced in the developing chick by RPR-V, an organophosphate.

Fertile White Leghorn chicken eggs were injected on day 4 of incubation with 0, 0.25, 0.50 or 1.00 mg RPR-V/egg and opened on day 20 or allowed to hatch. The following parameters were examined: hatchability, growth and development, external malformations and skeletal deformities. As the dose was increased, the hatchability decreased, and the incidence of deformities increased. Staining of the skeleton with Alizarin red showed clear deformities in the embryos. The results suggest that RPR-V has teratogenic effects on chick embryos when injected on day 4 of incubation.     

Activity of aldrinepoxidase, 7-ethoxycoumarin-O-deethylase and 7-ethoxyresorufin-O-deethylase during the development of chick embryos in ovo

Since the chick embryoin ovo is susceptible to the action of some agents needing metabolic activation we studied the development of the activity of cytochrome P450-dependent monooxygenases in embryo/fetal tissue. The activities of aldrinepoxidase (AE), 7-ethoxycoumarin-O-deethylase (ECOD) and 7-ethoxyresorufin-O-deethylase (EROD) were measured in whole embryos, liver and yolk sac tissue of the chick embryo during developmentin ovo from day 4 to day 15 of incubation (DI). In yolk sac tissue enzyme activities could be detected from DI 4 on. While EROD activity was only marginally developed, AE and ECOD activities were more pronounced in the earlier developmental period and showed a clear decrease by the time the liver activities rose. With the methods used AE activity could be measured in the homogenate of the whole embryo proper from DI 4 on while EROD and ECOD activity was not detectable before DI 6 or DI 7, respectively. In liver tissue enzyme activities of the three monooxygenases studied developed to a considerable degree from DI 9 on and tended to exhibit maximum values around DI 11–13. Studies on monooxygenase activities in extra-embryonically located tissues have not been published until now. The importance of the yolk sac as a metabolically relevant organ during embryogenesis is pointed out in this study.     

Effects of in ovo injection of carbamates on chick embryo hatchability, esterase enzyme activity and locomotion of chicks

Carbaryl and aldicarb, two carbamate pesticides used extensively throughout the United States, are known to act as acetylcholinesterase inhibitors. We have demonstrated previously that exposure to carbaryl and aldicarb in young chicks caused persistent locomotion alterations with no correlation to esterase inhibition. In this study, we investigated the effects of these carbamates when injected in ovo to chick embryos, at two time periods (days 5 and 15) during incubation. Carbaryl dosed at 45 mg kg-1 egg weight was extremely toxic to the embryos on day 5 of incubation. Hatchability was reduced to 0% as compared to 80% when carbaryl was injected on day 15 of incubation. Aldicarb at 1.5 mg kg-1 egg weight had no major effect on hatchability when injected either on day 5 or day 15 of incubation (hatchability = 90 and 100%, respectively). Plasma, liver and brain esterases were measured in the chick at different time points during incubation and after hatching. Brain acetylcholinesterase (AChE) and liver cholinesterase (ChE) were inhibited significantly during incubation in embryos dosed on day 15 with both carbaryl and aldicarb. Liver carboxylesterase was inhibited significantly during incubation with only the carbaryl treatment. All esterase enzyme activities returned to normal after hatching. Plasma ChE and carboxylesterase levels were not affected with either carbaryl or aldicarb treatment from 8 until 47 days after hatching. Neither carbamate had any effect on brain neuropathy target esterase (NTE) activity either during incubation or after hatching. The locomotion of chicks was affected in both treatment groups until 47 days after hatching. This study indicates that carbaryl and aldicarb may cause long-term delayed alterations in the chicks.