Action of ethanol on different skull and brain parameters in the chick embryo

Fifty microliters of ethanol diluted in 50 microl of distilled water were injected into the air chamber of chick eggs immediately before their incubation, and modifications in a series of parameters were recorded. The somatic weight of the ethanol-treated embryos was lower compared with control and vehicle-administered embryos during days 13, 15, 17, and 19 of incubation, but was the same on day 21. The brain weight was lower in the ethanol-treated embryos on all the days studied (days 13, 15, 17, 19, and 21 of incubation). Skull measurements showed that the transverse anteroposterior and sagittal diameters were significantly smaller in ethanol-treated embryos compared with control and vehicle-administered embryos on days 17, 19, and 21 of incubation.     

Effect of in ovo injection of vitamin C during incubation on hatchability of chickens and ducks

The aim of the investigation was to ascertain the influence of different doses of vitamin C injected at selected dates of incubation into the eggs of broiler breeders and Pekin ducks on hatchability. The injected vitamin C doses were administered into the air cell on the 13th, 15th and 17th days (3 and 6 mg--chickens) and on the 12th and 20th days (4 and 8 mg--ducks) of incubation. In the case of chickens, no significant differences were recorded between the control and experimental groups with regard to hatchability, although the highest value of hatchability from fertilized eggs was determined in the group injected with 6 mg of vitamin C on the 15th day of incubation. On the other hand, in ducks, significant differences were found between the control and experimental groups (4 mg of vitamin C administered on days 12 and 20 and 8 mg of vitamin C injected on day 20 of incubation) regarding hatchability. The value of this trait was higher in the group of eggs injected with ascorbic acid in comparison with the eggs which were not treated. On average, the difference amounted to 32.5 percentage points. Similarly, in the case of the number of dead embryos and unhatched chicks, better results were observed in the above-mentioned experimental groups. In summary, vitamin C injected into chicken eggs failed to influence hatchability. In the case of duck eggs, it was demonstrated that their injection on the 20th day of incubation with selected doses of vitamin C (4 and 8 mg/egg) improved hatchability by decreasing the proportions of dead and unhatched embryos.     

The development of the neurons of the glossopharyngeal (IX) and vagal (X) sensory ganglia in chick embryos

The timetable of cell generation, neuronal death and neuron numbers in the fused proximal glossopharyngeal (IX) and vagal (X) ganglion and distal IX and X ganglia were studied in normal and nerve growth factor (NGF) treated chick embryos. 3H-thymidine was injected between the 3rd and 7th days of incubation and embryos sacrificed on the 11th day. Neurons in the distal IX and X ganglia were generated between the 2nd and 5th days of incubation, the peak mitotic activity occurring on the 4th and 3rd days, respectively. Neurons of the proximal IX and X ganglion were generated between the 4th and 7th days, with maximum neuron generation on the 5th day of incubation. Counts of neurons in the 3 ganglia between the 5th and 18th days of incubation showed a maximum of 22,000 on the 8th day in the proximal IX and X ganglion and this decreased to 12,000 by the 13th day. In the distal IX ganglion, the neuron number decreased by 44% from 4,500 on the 6th day to 2,500 by the 11th day. A similar decrease of 43% was found in the distal X ganglion, the neuron number falling from 11,500 on the 7th day to 6,500 by the 11th day of incubation. Neuronal cell death in these ganglia extended from the 5th to the 12th day of incubation, maximum cell death occurring at or after the cessation of mitotic activity. NGF administration from the 5th to the 11th day of incubation did not have a measurable effect on the neurons of proximal IX and X and distal IX ganglia, but increased neuronal survival by 30% in the distal X ganglion.(ABSTRACT TRUNCATED AT 250 WORDS)     

Development of spontaneous motility in chick embryos. Normal development and the activating effect of strychnine and picrotoxin.

The longitudinal development of spontaneous motility in chick embryos was studied by Kovach's method (Kovach 1970) from the 10th day of incubation up to hatching, in completely intact eggs. From the 10th to 12th day of incubation, very low amplitude movements of a burst character predominated in spontaneous motility. From the 13th day, both low and high amplitude movements could be distinguished. From the 18th day, high amplitude movements alternating with intervals of motor inactivity preponderated. This discontinuous motility, which was most pronounced on the 20th day of incubation, changed to periodic strong hatching movements. Reduction of spontaneous motility after the 17th day of incubation was not confirmed. Strychnine already activated spontaneous motility in 11-day embryos, but typical convulsions did not appear until the 15th incubation day. With picrotoxin, motility was likewise stimulated in 11-day embryos and paroxysmal activation did not occur until the 15th incubation day. In older embryos, convulsions were gradually succeeded by a continuous increase in spontaneous motility. The effect of picrotoxin had a much longer latent period than the effect of strychnine.     

Histochemistry of mucosubstances in the gallbladder epithelium of the chick embryo

Summary Mucosubstances of epithelial cells of the chick embryo gallbladder were investigated by histochemical methods from days 12–21 of incubation (stages 38–46). On incubation day 14, only neutral mucins were detected; on day 15, neutral and sulfated mucosubstances were observed in the epithelial cells that invaginate the underlying mesenchyme. In the same sites, at day 16 of incubation, neutral, carboxylated and sulfated mucins were seen. From the 17th day of incubation until hatching, neutral, carboxylated and sulfated mucosubstances were present in the surface cells and in the cells lining the epithelial invaginations. During this period, the chemical characteristics of the secretory material are similar to those observed by Yamada and Hoshino (1972) in the fowl.Dedicated to Professor Dr. T.H. Schiebler on the occasion of his 65th birthday     

Histochemistry of mucosubstances in the gallbladder epithelium of the chick embryo

Mucosubstances of epithelial cells of the chick embryo gallbladder were investigated by histochemical methods from days 12-21 of incubation (stages 38-46). On incubation day 14, only neutral mucins were detected; on day 15, neutral and sulfated mucosubstances were observed in the epithelial cells that invaginate the underlying mesenchyme. In the same sites, at day 16 of incubation, neutral, carboxylated and sulfated mucins were seen. From the 17th day of incubation until hatching, neutral, carboxylated and sulfated mucosubstances were present in the surface cells and in the cells lining the epithelial invaginations. During this period, the chemical characteristics of the secretory material are similar to those observed by Yamada and Hoshino (1972) in the fowl.     

Development of the cerebrospinal fluid in chick embryos. Physicochemical properties.

The development of the physicochemical properties of the cerebrospinal fluid (CSF) was studied in chick embryos from the 9th day of incubation up to hatching. Some of these properties were compared with the corresponding blood or blood plasma properties. During the second half of incubation the CSF pressure rose from 13.2 plus or minus 0.18 mm H2O in 9-day-old embryos to 80.7 plus or minus 0.48 mm H2O just prior to hatching. The critical stages of this development were the 13th to 15th and the 19th to 21st day of incubation. In 13- and 15-day-old embryos, CSF pressure fell sharply after the intracerebral injection of ouabain, but in 19-day embryos it was unaffected. Except for the 15th and 19th incubation day, the CSF pH was always lower than the plasma pH. From the 11th day of incubation up to hatching, the CSF pH fell from 7.36 plus or minus 0.002 to 7.2 plus or minus 0.005. On the 11th and 13th day, specific CSF resistance was higher than plasma resistance, whereas from the 17th incubation day it was significantly lower than the plasma value. During the second half of incubation, specific CSF resistance fell from 1.059 times 10(6) to 0.824 times 10(6) omega mm.m(-1). A difference between the D.C. potential of the venous blood and the CSF appeared for the first time in 15-day-old embryos, the CSF being negative in relation to the blood. By the end of the incubation period this potential difference rose to 10.82 times 0.07 mv.     

Induction of feather malformations in chick embryos by cadmium: protection by zinc

Various doses of cadmium chloride were injected to chick embryos between the seventh and 14th day of incubation. Doses over 15 micrograms/egg produced high mortality and, when injected between the tenth and 11th day, widespread curling of the feathers in the surviving embryos. A different type of malformation, consisting of hemorrhagic atrophy of the distal part of the feathers, was observed in the embryos injected with similar doses during the 12th day. No feather malformations were observed in embryos injected before the ninth or after the 12th day of incubation. The simultaneous injection of an equimolar amount of zinc sulfate prevented the feather malformations.     

Electrophoretic characterization of glycosaminoglycans during development of the chick embryo skin

Glycosaminoglycans extracted by CPC precipitation from chick embryo skin at 9, 12, 15, 18, 21 days of incubation were separated by three different electrophoretic methods on acetate cellulose strips. We observed the presence of Hyaluronic acid, Dermatan Sulfate and Chondroitin-4-Sulfate during the whole period considered and of Heparan Sulfate only after the 9th day. Dermatan Sulfate increases until the 15th day then decreases progressively; on the contrary Hyaluronic acid and Chondroitin-4-Sulfate decrease during days 9 to 15 then increase until hatching. Heparan Sulfate appear the 9th day then increases progressively until hatching.     

Further studies on the lipid metabolism of the normal and vitamin B12-deficient chick embryo

1. The triglyceride, cholesterol ester and total phospholipid fractions were isolated from the livers and yolk sacs of normal and vitamin B₁₂-deficient chick embryos after 13, 15, 17, 19 and 21 days of incubation, and the fatty acid compositions were determined. 2. At all stages of incubation, the concentration of cholesterol ester in the livers of the normal embryos were greater, and on days 15 and 17 the concentrations of triglyceride were considerably less, than the corresponding concentrations in the livers of the deficient embryos. 3. Between day 13 and day 21 of incubation the concentration of oleic acid in the liver triglycerides of the normal embryos increased, whereas the concentrations of palmitic acid and docosahexaenoic acid decreased. Vitamin B₁₂ deficiency resulted in higher concentrations of palmitic acid in the liver triglycerides on days 15, 17 and 19, higher concentrations of C₁₈ polyunsaturated acids on days 13 and 15 and lower concentrations of oleic acid on days 13, 15, 17 and 19. 4. At all stages of development, cholesterol oleate accounted for almost 80% of the total liver cholesterol esters in both normal and deficient embryos. 5. As development of the normal embryos progressed, the concentrations of palmitic acid and arachidonic acid in the liver phospholipid decreased, whereas the concentrations of stearic acid and docosahexaenoic acid increased. Vitamin B₁₂ deficiency resulted in markedly higher concentrations of stearic acid and palmitic acid and markedly lower concentrations of arachidonic acid and docosahexaenoic acid in the liver phospholipids. 6. Vitamin B₁₂ deficiency did not influence the fatty acid composition of the triglyceride, cholesterol ester and phospholipid fractions either in the yolks of fertile unincubated eggs or in the yolks obtained from eggs that had been incubated for 13, 15, 17, 19 and 21 days.     

An attempt to activate the spontaneous motility in chick embryos with tetanus toxin

The effect of tetanus toxin on spontaneous motor activity was studied in chick embryos between the 11th and 19th day of incubation. The toxin--dose 20 mg/kg egg weight ( = 2.86 X 10(3) mouse MLD) in 25 microliters isotonic NaCl solution--was injected into the tibial or the wing muscles. Tetanus toxin induced demonstrable activation of embryonic motility from the 15th day of incubation onwards. Activation attained 155-200% of resting activity. The activating effect was manifested for the first time by motor paroxysms in 17-day and particularly 19-day embryos. Tetanus toxin activation was effectively depressed by glycine (100 mg/kg e.w.) and GABA (100 mg/kg e.w.), the former having a stronger effect than the latter. The effect of tetanus toxin on spinal embryos was relatively more pronounced, while the depressant effect of the inhibitory amino acids (especially glycine) was weaker. The results are evaluated as further evidence that central inhibitory mechanisms are connected up in regulation of the spontaneous motor output activity of chick embryos on about the 15th day of incubation.     

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.     

Changes induces by haloperidol (antidepressant drug) on the developing retina of the chick embryo

Morphological and histological studies were done on the retina of chick embryos of 6th, 10th and 16th days of incubation by a single dose of haloperidol (0.25 mg/egg), injected on day zero and 5 of incubation. To get an idea about the extent of the teratogenic effect of this drug on the development retina. Sign of malformation in the chick embryo after administration of haloperidol were seen as absent of the ear vesicles, eyes or decreased size of them. Retardation of growth of the retina at 6th, 10th and 16th days treated chick embryo were observed as evidenced of reduction of the size of the retina, associated with sign of degeneration of the retina cells.

Conclusion

The injection of haloperidol drug give rise to several side effects as retardation of growth and degeneration of the cells. The decrease of the thickness of the layers and less density of the cells was related to direct effect of the drug on the cells of this organ. Also on the DNA formation and on the retardation of cellular mitotic activates, therefore the retina appeared decrease in thickness and less cell density with degeneration its cells.     

Development of spontaneous motility in chick embryos. Sensitivity to aminergic transmitters.

The consequences of systemic administration of aminergic transmitters (n-adrenaline 16 microgram/kg egg weight; serotonin 2.5 and 5 mg/kg e.w.; dopamine 2.5 and 5 mg/kg e.w.) for the spontaneous motility and heart rate of 11- to 19-day chick embryos were studied intack eggs. The following results were characteristic for all three transmitters: a) when administered to 11- and 13-day embryos their effect was non-significant; the first signs of activity did not appear until the 15th day of incubation. The effect on 17- and 19-day embryos was stronger. b) After the 15th day of incubation, all these transmitters had a predominantly inhibitory effect on spontaneous motility; in 17- and 19-day embryos this acquired a periodic character. c) The changes in spontaneous motility did not correlate significantly in any way with the relatively small heart rate changes. It is concluded from the results that aminergic mechanisms begin to participate in regulation of the spontaneous motility of chick embryos from the 15th day of incubation, and not before.     

Ontogenetical development of the chick and duck subcommissural organ. An immunocytochemical study

The ontogenetical development of the subcommissural organ (SCO) was investigated in chick embryos collected daily from the 1st to the 21st day in incubation. Some duck embryos, and adult chickens and ducks were also studied. Immunocytochemistry using an anti-Reissner's fiber (RF) serum as the primary antibody was the principal method used. In the chick embryos the events occurring at different days of incubation were: day 3 morphologically undifferentiated cells in the dorsal diencephalon displayed immunoreactive material (IRM); days 4 to 6 immunoreactive cells proliferated, formed a multilayered structure and developed processes which traversed the growing posterior commissure and ended at the brain surface; day 7 blood vessels penetrated the SCO, scarce hypendymal cells appeared, the first signs of ventricular release of IRM were noticed, appearance of IRM bound to cells of the floor of the Sylvius aqueduct; day 7 to 10 the number of apical granules and amount of extracellular IRM increased progressively; day 11 RF was observed along the Sylvian aqueduct, day 12 RF was present in the lumbar spinal cord; day 13 IRM on the aqueductal floor disappeared; days 10 to 21 hypendymal cells proliferated, developed processes and migrated dorsally, ependymal processes elongated and their endings covered the external limiting membrane. In adult specimens the ependymal cells lacked basal processes and the external membrane was contacted by hypendymal cells. the duck SCO appears to follow a similar pattern of development.     

Penicillin activates spontaneous motility in chick embryos

The development of the motor reaction to i.v. injection of the sodium salt of penicillin G in a dose of 0.9 x 10(6) I.U./kg egg weight was studied in chick embryos (normal and spinal) from the 11th to the 19th day of incubation. Penicillin first caused standard activation of embryonal motility from the 15th day of incubation, in both normal and spinal embryos. Activation was at first continuous in character (a twofold increase in the frequency of spontaneous movements). In 17- and particularly in 19-day embryos a typical paroxysmal reaction developed, with pronounced intervals of motor rest. The proportion of the spinal component in the penicillin reaction was abut 40% and of the supraspinal component about 60% of total motor activity. In older embryos (after the 15th day of incubation), the motor reaction to penicillin could be effectively modified by the systemic administration of glycine and GABA. It is concluded from the results that penicillin does not activate embryonal motor activity until a given stage of development of the CNS has been attained. It is a developmental phenomenon with a spinal and a supraspinal component, in which central inhibitory mechanisms participate.     

Development of the differential effect of oxazepam on spontaneous and activated motility in chick embryos

Development of the acute depressant effect of oxazepam (in a dose of 10 mg/kg egg weight) on spontaneous and activated motor activity was studied in chick embryos (incubation age 11-19 days). The depressant effect of oxazepam was demonstrated for the first time in 13-day-old embryos. From the 15th day of incubation it led to prolonged depression of embryonic motility. From the 15th day the depressant effect of oxazepam depended on the connections of the central motor output with the prosencephalon. In mesencephalic, rhombencephalic and spinal preparations the effect of oxazepam was insignificant. In 13- and 15-day-old embryos oxazepam blocked the activating effect of strychnine and pentylenetetrazol, but not the picrotoxin-induced activation. The picrotoxin activation of embryonic motility was incompletely blocked for the first time in 17-day-old and completely only in 19-day-old embryos. The results are discussed with reference to heterochronia of development of the various components in the complex GABA receptor.     

Effects of thyroxine on mitochondrial morphology and uptake of 35S in embryonic chicken livers

Ultrastructural analyses of reactions of mitochondria in hepatocytes of chicken embryos to low levels of exogenous thyroxine (T4) reveal that such reactions (overall swelling accompanied by disruption of crest geometry) first take place at about 10 days of incubation, T4 having been administered on the 6th day. Physically altered mitochondria may be seen after 11-12 days of incubation but are no longer evident by 13 days. Correlated with the initial evidence of T4 effects on mitochondria at 10 days of incubation is a spurt in hepatocyte proliferation. The time correlation observed between T4 induced mitochondrial changes in morphology and abrupt increases in rates of cell proliferation, suggests that liver nuclear receptors for thyroxine are unavailable prior to 9-10 days of incubation. Golgi complexes within the hepatocytes appear to be especially active in the production of electron-opaque vesicles from at least the 8th day of incubation to 11-12 days. Uptake of 35S (probably into chondroitin sulphates) was found to be fifteen times greater on the 8th day of incubation than at 15 days. This correlates with the period of heightened activity of the Golgi complex. In livers exposed to T4 on the 6th day, uptake of 35S was higher on the 9th and 10th days of incubation as compared to controls.     

Action of acetazolamide on the chick embryo during late development.

Acetazolamide was injected into chick embryos on the 14th or 15th day of incubation. Doses ranging between 5 and 10 mg per egg produced a retardation in the growth of long bones. The affected bones contained a normal proportion of mineral as determined by ashing and presented a normal histological picture. On the basis of these findings, it is suggested that the alterations were not due to a specific direct effect of the drug on bones. The incorporation of 131-I by the thyroid glands of acetazolamide-injected embryos was analyzed radioautographically and quantitated on the same 6 mu-paraffin sections, with a thin window Geiger counter. The incorporation appeared notably reduced 3 h after the injection of acetazolamide and the reduction persisted for a least 24 h.the electron microscopical observation of thyroid follicular cells from similarly treated embryos showed that the cytological characteristics indicating an active protein synthesis were unmodified with respect to those found in control embryos. These results may indicate that acetazolamide inhibits the iodination of the throid hormone without interfering with the synthesis of the globulin. It is suggested that the growth retardation observed in the embryos treated with acetazolamide may be secondary to the action of the drug on the thyroid gland, although this action appears to be a transitory one.