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PubMed1982

Age-dependent effects of glutamate on photoreceptor cells of the isolated chick embryo retina.

Neurotoxic and gliotoxic effects of glutamate were studied in isolated chick embryo retinas of various ages. Employing a short-term incubation system, we found that initial cellular changes in retinas from 15- and 21-day chick embryos were localized to glial Müller cells as previously shown in retinas from 12-day embryos. In the older retinas, however, an additional selective lesion was consistently found in the presumptive photoreceptor cells. Similar photoreceptor damage was not seen in 12-day retinas, even after treatment with relatively high glutamate doses. Autoradiographic uptake of tritiated glutamate in retinas at 12 and 14 days was localized to the glial Müller cells; in the 14-day retinas, however, there was also uptake of the labeled amino acid into the developing inner segments. Uptake of tritiated glutamate in retinas at younger (8-day) and older (21-day) ages did not show any obvious localization of the label.

Age Factors↗

PubMed1985

Effect of glutamine on fructose 2,6-bisphosphate and on glucose metabolism in HeLa cells and in chick-embryo fibroblasts.

Glutamine caused a dose-dependent decrease in fructose 2,6-bisphosphate concentration in both HeLa cells and chick-embryo fibroblasts. The effect was complete within 15 min in HeLa cells, but required more than 9 h in the fibroblasts. Half-maximal effects were obtained with 0.1-0.3 mM-glutamine. In chick-embryo fibroblasts, but not in HeLa cells, glutamine induced a time-dependent decrease in the activity of phosphofructokinase-2, which correlated with the decrease in fructose 2,6-bisphosphate. Glutamine decreased the glycolytic flux by about 25% only in chick-embryo fibroblasts. The difference in glycolytic response between the two types of cells might correspond to a difference in the sensitivity of phosphofructokinase-1 for fructose 2,6-bisphosphate. In HeLa cells, glutamine caused a 2-3-fold stimulation of the synthesis of glycogen, a 50% decrease in the concentration of fructose 1,6-bisphosphate and a more than 80% decrease in the concentration of 5-phosphoribosyl pyrophosphate; the concentrations of hexose 6-phosphates and ATP were not affected.

Animals↗

PubMed1972

Studies on type C influenza virus in the chick embryo.

The effect of varying conditions of inoculation and incubation on the growth of type C influenza virus in the allantoic cavity of the developing chick embryo were investigated. It was found that the highest yields of both virus haemagglutinin and infectious virus were obtained following the inoculation of chick embryos at 8 days with subsequent incubation at 32 degrees C. Using the chick embryo allantoic cavity for titration of infectious virus, growth curves of allantoically propagated virus under varying inoculation and incubation conditions were determined.

Animals↗

PubMed1988

Magnetic resonance microscopy of chick embryos in ovo.

Magnetic resonance imaging (MRI) of the live 11-day chick embryo with special radiofrequency coils and 3-D imaging methods has produced contiguous 1.25-mm-thick slices with 200-microns pixel resolution, permitting definition of cardiac chambers, cerebral ventricles, spinal cord, liver, and lungs. It was the objective of this study to image younger chick embryos in ovo with higher spatial resolution through the application of implanted radiofrequency coils. Fertilized Arbor Acre eggs were windowed at 9, 6, and 4 days. Circular coils 18 mm in diameter tuned to 85.5 MHz were suspended around the developing embryo. The eggs were sealed with tape and maintained at 37 degrees C during the imaging procedure. MRI was performed in a 2.0-Tesla GE system utilizing a 3-D Fourier transform acquisition in sagittal and axial planes with a partial saturation sequence (TR = 400 ms, TE = 27 ms). Approximately 1 hour of imaging time was required to obtain 16 contiguous 600-microns-thick slices with 50-microns pixel resolution. Embryos remained viable through the imaging procedure. Embryos were photographed, fixed, and cleared for correlative anatomical study. Vitelline vessels, dorsal aorta, aortic arches, cardinal veins, and cardiac chambers were identified as areas of decreased signal intensity. Cerebral ventricles and the vitreous portion of the eye have signal intensities that are less than adjacent neural, scleral, and lens tissue. Further refinements in MR instrumentation and imaging sequences promise improvements in resolution and offer the potential for sequential observations of the intact embryo.

Animals↗

PubMed1953

Studies on the psittacosislymphogranuloma group. III. The effect of aureomycin on the propagation of virus in the chick embryo.

The findings presented indicate that aureomycin could become associated with tissue of the chick embryo by both hematogenous distribution and direct adsorption. Treatment of chick embryos infected with MP virus with 1 mg. of aureomycin by the allantoic route caused an inhibition of virus growth in the allantoic membrane. The drug had no effect on "inert" virus, and appeared to have little effect on adsorption of virus to host tissue. Complete inhibition of growth during the time interval corresponding to the first cycle of multiplication could be achieved only if the drug was administered within 6 to 8 hours after virus inoculation. Partial inhibition of virus multiplication could be achieved even if the administration was delayed as late as 24 hours after infection. In these experiments the chief role of the antibiotic appeared to be one of virustasis reflected in a prolongation of the latent period (non-infectious phase). The virus was able to resume its growth when a critical low level of the drug in the allantoic membrane was reached. When infectivity titrations were carried out using various tissues and organs of treated and untreated embryos, it was found that no virus was detectable in the brains of treated embryos as late as 192 hours after inoculation of virus. This was in contrast with the findings in allantoic membranes and livers of such embryos; these organs showed virus at 120 and 144 hours, respectively. In untreated controls, virus appeared in membranes at 24 hours, in the liver at 48 hours, and in the brain at 72 hours.

Animals↗

PubMed1985

Conversion of 5-aminolaevulinate into haem by homogenates of human liver. Comparison with rat and chick-embryo liver homogenates.

To assess whether the synthesis of haem can be studied in small amounts of human liver, we measured kinetics of the conversion of 5-aminolaevulinate into haem and haem precursors in homogenates of human livers. We used methods previously developed in our laboratory for studies of rat and chick-embryo livers [Healey, Bonkowsky, Sinclair & Sinclair (1981) Biochem. J. 198, 595-604]. The maximal rate at which homogenates of human livers converted 5-aminolaevulinate into protoporphyrin was only 26% of that for rat, and 58% of that for chick embryo. In the absence of added Fe2+, homogenates of fresh human liver resembled those of chick embryos in that protoporphyrin and haem accumulated in similar amounts, whereas fresh rat liver homogenate accumulated about twice as much haem as protoporphyrin. However, when Fe2+ (0.25 mM) was added to human liver homogenates, mainly haem accumulated, indicating that the supply of reduced iron limited the activity of haem synthase, the final enzyme in the haem-biosynthesis pathway. Addition of the potent iron chelator desferrioxamine after 30 min of incubation with 5-amino[14C]laevulinate stopped further haem synthesis without affecting synthesis of protoporphyrin. Thus the prelabelled haem was stable after addition of desferrioxamine. Since the conversion of 5-amino[14C]laevulinate into haem and protoporphyrin was carried out at pH 7.4, whereas the pH optimum for rat or bovine hepatic 5-aminolaevulinate dehydratase is about 6.3, we determined kinetic parameters of the human hepatic dehydrase at both pH values. The Vmax was the same at both pH values, whereas the Km was slightly higher at the lower pH. Our results indicate that the synthesis of porphyrins and haem from 5-aminolaevulinate can be studied with the small amounts of human liver obtainable by percutaneous needle biopsy. We discuss the implications of our results in relation to use of rat or chick-embryo livers as experimental models for the biochemical features of human acute porphyria.

Aminolevulinic Acid↗

PubMed2001

Developmental effects of lead acetate on the chick embryo metanephros.

The developmental effects of lead acetate were studied in the chick embryo metanephros, the third renal rudiment that acquires morphological characteristics of functioning kidney already during the prenatal life. Lead exposure was obtained by applying a lead acetate solution on the chick embryo chorioallantoic membrane at the days 9, 10 and 11 of incubation. Quantitative evaluation of the lead concentration assessed by furnace atomic absorption spectrophotometry at the days 14 and 21 of incubation demonstrated metal presence both in the chorioallantoic membrane (CAM) and in metanephros (MN). The lead concentration was higher in CAM than in MN; the metal amount was similar in the CAM of 14 and 21 day embryos, but significantly higher in the 14day embryo MN than in the 21 day embryo MN. Morphological observations on metanephros tissue of control and lead-treated embryos were performed under light, electron transmission and electron scanning microscopes. Peculiar attention was devoted to the expression of the junctional protein connexin 43, the major component of the gap junctions in the renal cells. The results indicated that lead treatment does not intervene in the general differentiation of the metanephric nephrons. The lead is reabsorbed by the proximal tubule cells that are engulfed by endocytotic vacuoles and metal deposits and show long term degenerative changes. Expression of Cx43 protein and ultrastructure of gap junctions between proximal tubule cells appeared to be unchanged. The morphological aspects of the MN corpuscles and tubules agree with the suggestion of a lead cytotoxic effect but do not corroborate, at least in this experimental model, the view of primary damage exerted by lead on the gap junctions of the renal epithelial cells.

Abnormalities, Drug-Induced↗

PubMed1989

17 beta-estradiol secretion in normal and hypophysectomized chick embryos.

Ovaries from decapitated, sham-operated and intact 18-day-old chick embryos were cultured in Medium 199 for 6 h, and the amount of 17 beta-estradiol released into the culture media was determined by radioimmunoassay. Ovaries from decapitated embryos secreted significantly lower amounts of 17 beta-estradiol than those from intact embryos, but there was no difference when they were compared to ovaries from sham-operated embryos in this respect. On an ovarian weight basis, 17 beta-oestradiol production was significantly different between the 3 groups of embryos, the ratio being highest in the decapitates. 17 beta-Estradiol concentration was the same in serum from both decapitated and intact female embryos. Considering these results, it is concluded that the hypophysis does not control 17 beta-estradiol secretion by the chick embryo ovary even near hatching time.

Animals↗

PubMed1975

Coordinated motor output in the hindlimb of the 7-day chick embryo.

Electromyographic recordings from individual identified ankle muscles of the 7-day chick embryo (stage 31) were used to determine the organization of motor output at a developmental stage shortly after the onset of spontaneous motility in the leg. During spontaneous motility of the embryo, the electromyographic recordings from the gastrocnemius, peroneus, and tibialis muscles displayed bursts of motor unit activity which alternated with periods of little or no activity. Since the control of skeletal muscle in the chick embryo is neurogenic rather than myogenic, these findings imply that the motoneurons to a given muscle are driven by a common source. Since flexor and extensor muscles are attivated at different times, different central connections to flexor and extensor motoneurons must be present in the central nerbous system of the 7-day embryo. Moreover, since inhibition is known to play an important role in the selective activation of agonist and antagonist muscles, the present results suggest that functional inhibitory synapses may be present in the lumbosacral central nervous system at this stage of development. The basic pattern of muscle activation observed in the 7-day embryo is similar to that seen in older embryos. Since these patterns appear prior to the time at which motor responses to sensory stimulation of the leg can be demonstrated, it is likely that the neural pattern generating circuits for selective activation of muscles are established in the central nervous system without reliance on functional reflexes.

Animals↗

PubMed1997

Histological changes during development of the cerebellum in the chick embryo exposed to a static magnetic field.

Few studies have been performed to evaluate the ultrastructural changes that exposure to static magnetic fields (SMF) can cause to the processes of cell migration and differentiation in the cerebellum during development. Thus, we have studied the development of the cerebellum in the chick embryo (n = 144) under a uniform SMF (20 mT). All of our observations were done on folium VIc of Larsell's classification. The cerebella of chick embryos, which were exposed solely on day 6 of incubation and sacrificed at day 13 of incubation [short exposure (S)1; n = 24], showed an external granular layer (EGL) that was less dense than the EGL in the control group (n = 24). The molecular layer (ML) exhibited a low number of migratory neuroblastic elements. Moreover, the internal granular layer (IGL) was immature, with the cellular elements less abundant and more dispersed than in controls. In chick embryos exposed on day 6 of incubation and sacrificed at day 17 (S2; n = 24), the outstanding feature was the regeneration of the different layers of the cerebellar cortex. The cerebellar cortex of chick embryos exposed continuously to an identical field from the beginning of the incubation up to day 13 [long exposure (L)1; n = 24] or day 17 (L2; n = 24) of incubation showed a higher number of alterations than that of group S1. Electron microscopy confirmed the findings from light microscopy and, at the same time, showed clear signs of cell degeneration and delay in the process of neuronal differentiation. This was more apparent in groups L1 (100%) and L2 (100%) than in groups S1 (95.4%) and S2 (65.2%). In conclusion, the present study showed that SMF can induce irreversible developmental effects on the processes of cell migration and differentiation of the chick cerebellar cortex.

Animals↗

PubMed1983

Diazepam inhibits the spreading of chick embryo fibroblasts.

Diazepam (Valium/Roche), a benzodiazepine derivative, reversibly inhibited the spreading of chick embryo fibroblasts in a dose-related manner. This inhibition was accompanied by: 1 Fewer and less organized microfilament bundles. 2 A reduction in myosin fluorescence along these structures. 3 A reduction in the total myosin content of cells compared to corresponding controls. Overall results suggest that myosin plays an active role in the spreading of chick embryo fibroblasts.

Animals↗

PubMed1996

Isolation of N-vinylprotoporphyrin IX after hepatic cytochrome P450 inactivation by 3-[(arylthio)ethyl]sydnone in chick embryos pretreated with phenobarbital, glutethimide, dexamethasone, and beta-naphthoflavone: differential inhibition of ferrochelatase by N-vinylprotoporphyrin regioisomers.

Several xenobiotics caused hepatic porphyrin accumulation through mechanism-based inactivation of cytochrome P450(P450) and heme alkylation. Loss of iron from the alkylated heme results in formation of an N-alkylporphyrin, which is a potent inhibitor of ferrochelatase. N-Vinylprotoporphyrin IX (N-vinylPP) was identified in chick embryo liver after in ovo administration of 3-[(arylthio)ethyl]sydnone (TTMS). Pretreatment of chick embryos with beta-naphthoflavone, which causes a 90-fold increase in P450 1A levels, did not increase the formation of N-vinylPP after TTMS administration, showing that the heme moiety of P450 1A does not contribute to the formation of N-vinylPP. Increased amounts of N-vinylPP were isolated from dexamethasone-, phenobarbital-, and glutethimide-pretreated chick embryos, and it is possible that P450 2H and/or a P450 3A-like isozyme contributes to the formation of N-vinylPP. The ring B-substituted (NB) regioisomer of N-vinylPP constituted the lowest percentage of the total regioisomers (9-13%) in untreated and drug-induced chick embryos, thus supporting the concept that ring B of heme is occluded by a protein residue in the P450 active site. Previously, the finding that the NB regioisomer of N-ethylprotoporphyrin IX had one fifth the potency of the ring A-substituted (NA) regioisomer as a ferrochelatase inhibitor led to a proposal that an A-C ring tilt was important in N-alkylprotoporphyrins for ferrochelatase inhibition. The finding in the present study that the NA and NB regioisomers of N-vinylPP are equipotent does not support the above proposal. The ring C-substituted (NC) and ring D-substituted (ND) regioisomers of N-vinylPP had low potency.

Animals↗

PubMed1993

Angiogenesis induced by head and neck squamous cell carcinoma xenografts in the chick embryo chorioallantoic membrane model.

The angiogenic potential of head and neck tumors compared to nonneoplastic control tissues was investigated by using the chick embryo chorioallantoic membrane (CAM) as a bioassay for angiogenesis. Eighty tumor specimens from 10 patients with squamous cell carcinoma of the head and neck were grafted onto the CAM of 7-day chick embryos. The presence of tumor in the original explant was confirmed histologically. Forty-four embryos (55%) survived and were evaluated histologically at day 17. Tumors were growing on or invading the CAM mesoderm in 30 of 44 embryos (68%). Before harvesting of the membranes, the tumors and surrounding blood vessels were photographed, and the angiogenic responses were graded by a panel of blinded observers. Tumor explants elicited a significantly greater angiogenic response than nontumor controls (p = .01). We conclude that head and neck squamous cell carcinomas can induce an angiogenic response in vivo, presumably secondary to the production of an unidentified angiogenic factor, and that the chick embryo CAM is an effective model for quantifying angiogenesis induced by head and neck tumors.

Allantois↗

PubMed2004

Drug-drug interactions between antiarrhythmic drugs in chick embryos.

The drug-drug interactions between antiarrhythmic drugs were studied in chick embryos. Fertilized eggs of White Leghorns were incubated and investigated. Procainamide or flecainide with and without propranolol was injected into the air sac of a fertilized egg. Electrocardiograms (ECGs) were recorded 0 to 60 min after the injection. After each drug injection alone, the heart rate was not different compared with the control. However, the heart rate was significantly decreased by combinations of procainamide and propranolol or flecainide and propranolol. In addition, arrhythmia was produced in combination with propranolol. These findings indicate that the drug-drug interactions between antiarrhythmic drugs have a marked influence on the heart rate in chick embryos.

Animals↗

PubMed1988

The ontogeny and induction by zinc of hepatic chick embryo metallothionein.

To avoid the maternal influences inherent in mammalian models, the chick embryo was examined for the effects of development on the ontogeny of hepatic metallothionein (MT) induction. Livers from embryos were examined for total and cytosolic zinc (Zn) and copper (Cu), and cytosolic MT on 12, 14, 16, and 18-21 days of incubation (DI) as well as 1, 7, and 14 days posthatch (ph). Cytosolic Zn levels fell, from a high on 12 DI (45.04 +/- 1.05 ng/micrograms DNA), to 25.79 +/- 1.05 ng/micrograms DNA on 19 DI and then increased to 44.73 +/- 3.47 ng/micrograms DNA by 1 day ph. Cytosolic Cu followed a reverse pattern, with a high of 24.10 +/- 0.74 ng/microgram DNA on 16 DI. Both 109Cd radioassay analysis and gel filtration chromatography of hepatic cytosols showed a developmental pattern for MT similar to that of cytosolic Zn. These results are in contrast to the developmental pattern of the rat. To establish if yolk Zn levels limit hepatic MT in the 1-day neonate, the yolks of unincubated fertile eggs were supplemented with 26, 52, or 78% of the endogenous Zn (768 micrograms/yolk). As a result, hepatic MT of 1-day neonates increased by 3.9-, 4.7-, and 7.1-fold, respectively. Thus the initial level of Zn in the yolk has a significant impact on the final concentration of MT in neonatal liver. A study of MT induction in the 18-DI embryo revealed that yolk sac administration of Zn (550, 2750, 4950 micrograms/yolk) increased hepatic MT by 5.8-, 23-, and 39-fold, respectively. This demonstrates the inducibility of MT even at the point of minimum endogenous MT (18 DI). In summary, our results show that (1) a marked difference exists between the developmental patterns of MT in avian and mammalian species, and (2) chick embryo hepatic MT is highly responsive to exogenous Zn introduced into the yolk.

Animals↗

PubMed1990

Effect of changes in circulating blood volume on cardiac output and arterial and ventricular blood pressure in the stage 18, 24, and 29 chick embryo.

We studied the hemodynamic effects of changing volume loading in the chick embryo, before autonomic innervation, to test the hypothesis that the Frank-Starling mechanism functions in the embryonic myocardium. Dorsal aortic blood velocity was measured by pulsed Doppler. Heart rate and aortic diameter were also measured to calculate cardiac output and stroke volume index. Vitelline arterial and ventricular pressures were measured with a servo-null micropressure system in stage 24 embryos. Infusing isotonic solution intravenously resulted in linear increases in stroke volume index for stages 18 (y = 388x + 6.89), 24 (y = 466x + 7.86), and 29 (y = 549x + 4.96). The slopes and intercepts were statistically the same for all three stages. Similar volume loading in stage 24 embryos initially increased mean arterial pressure linearly, but at higher loading conditions, the rate of rise lessens. Thus, volume loading resulted in a decrease in vascular resistance. Withdrawing blood from stage 24 embryos resulted in a decrease in ventricular peak systolic and end-diastolic pressures. With reinfusion of the blood, systolic and end-diastolic pressures initially rose above baseline levels and later returned to normal. We conclude that a length-tension relation is present in the preinnervated embryonic heart and that vascular resistance changes inversely with loading conditions. We speculate that these mechanisms are the primary hemodynamic control mechanism in the early chick embryo.

Animals↗

PubMed2001

Does an equivalent of the "ventral node" exist in chick embryos? A scanning electron microscopic study.

The internal organs of vertebrates show species-specific left-right (L-R) asymmetries. Questions on the embryonic origin of these asymmetries have been fascinating embryologists since the 19th century. During the past years, remarkable progress has been made in answering these questions. Evolutionary highly conserved molecular signaling cascades have been identified that start from Hensen's node and transfer side-specific identity to the embryonic left and right halves. However, the question of what initiates these signaling cascades has remained unanswered. Studies on mouse embryos have shown that the ventral surface of Hensen's node consists of a ciliated epithelium called the ventral node. Recent findings suggest that the monocilia of ventral nodel cells generate a leftward flow of extracellular fluid possibly leading to the accumulation of an unknown morphogen at the left of the node, which then might start the signaling cascades. This hypothesis might explain the fact that gene defects causing ciliary dyskinesia are frequently associated with situs anomalies. Studies on chick embryos led to the discovery of the L-R signaling cascades. However, whether an equivalent of the ventral node exists in avian embryos remained unknown. Therefore, I examined the endoderm and epiblast of early chick embryos for the presence of monociliated cells. In the endoderm, a population of monociliated cells indeed was present. These cells, however, were neither confined to the area of Hensen's node nor did they form the predominant cell population at this location. In the epiblast, monociliated cells formed the predominant cell population at the periphery of the blastodisc but only a relatively small subpopulation of epiblast cells at Hensen's node. These findings suggest that, in the early chick embryo, an equivalent of the ventral node of mouse embryos neither exists on the ventral nor the dorsal surface of Hensen's node. It is unlikely that nodal cilia are required for initiating the L-R patterning in chick embryos.

Animals↗

PubMed1983

The contribution of the inferior endocardial cushion of the atrioventricular canal to cardiac septation and to the development of the atrioventricular valves: study in the chick embryo.

The contribution of the inferior endocardial cushion of the atrioventricular canal to cardiac septation and to the development of the atrioventricular valves was studied in the chick embryo by in vivo labelling techniques. The study was performed in White Leghorn chick embryos in which the dorsal cushion was labelled at stage 20-21 (Hamburger and Hamilton, 1951), when the endocardial cushions were not yet fused. The embryos were sacrificed at stage 35 (mature heart). These experiments allow us to conclude that the inferior atrioventricular cushion gives origin to: a) that part of the cardiac septum between the septal insertion of the antero-septal leaflet of the mitral valve and the fibrous ridge which is the equivalent to the human septal leaflet of the tricuspid valve (atrioventricular septum); b) the region of the interatrial and the interventricular septa adjacent to the atrioventricular septum; c) the portion of the antero-septal leaflet of the mitral valve which inserts into the septum; d) the fibrous ridge corresponding to the septal leaflet of the tricuspid valve. Microdissection shows that, when they appear at stage 18, the superior and inferior endocardial cushions of the atrioventricular canal are in continuity, without boundaries, with both the interatrial and interventricular septa. Therefore, each atrioventricular orifice opens into its corresponding ventricle, there being no stage in the development of the chick embryo heart in which the atrioventricular orifices are connected to the left ventricle at the same time. The development of the atrioventricular canal is similar in the chick and human.

Animals↗