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PubMed1994

Nicotinic and muscarinic acetylcholine responses in the embryo chick ciliary ganglion cells.

Nicotinic and muscarinic acetylcholine (ACh) responses were investigated in acutely dissociated chick ciliary ganglion neurons using the nystatin perforated patch clamp technique. ACh-induced a rapid transient inward current in 100% of the neurons at a holding potential of -60 mV. This rapid inward current was mimicked by nicotine but not by muscarine. The reversal potential of the rapid inward current was +10.5 mV and the current was inhibited by d-tubocurarine and hexamethonium in a dose-dependent manner. In 57.6% of neurons, a slow inward current was also induced by ACh at a holding potential of -20 mV. This slow inward current was mimicked by muscarine but not by nicotine. The slow inward current became smaller at a hyperpolarized potential but not reversed, being consistent with the fact that this current was elicited by the inhibition of M-current. p-Fluorohexa-hydrosiladifenidol (P-F-HHSiD) strongly inhibited the slow inward current, suggesting that the current was elicited by the activation of M3 receptors.

Acetylcholine↗
PubMed1995

ATP responses in the embryo chick ciliary ganglion cells.

The ATP responses in ciliary neurons acutely dissociated from chick ciliary ganglia were investigated using the nystatin perforated patch recording mode. ATP induced a transient inward current at a holding potential of -50 mV in a concentration-dependent manner. The half-maximum effective concentration of ATP was 8.2 microM and the Hill coefficient was 0.9. The current-voltage relationship of the ATP response revealed a strong inward rectification at potentials more positive than -30 mV and the reversal potential was near 0 mV. The relative potencies of purinoceptor agonists were in the order of ATP > 2-methylthio-ATP > ADP. Neither adenosine, AMP nor alpha, beta-methylene-ATP induced any response. The ATP-induced inward current was blocked by suramin, a selective P2 purinoceptor antagonist, in a concentration-dependent manner. The half-maximum inhibitory concentration was 4.5 microM. The cytosolic Ca2+ concentration was increased by ATP and suramin inhibited the increase in a concentration-dependent manner. These results suggest that ATP operates non-selective cation channels by acting on P2y purinoceptors and has a role in the excitation of ciliary neurons.

Adenosine Diphosphate↗
PubMed1980

Effect of polyamines on ADP-ribosylation by chick-embryo-liver nuclei.

Effects of polyamines on poly(ADP-ribose) formation and DNA synthesis in the chick-embryo-liver nuclei were investigated. When 14-day chick-embryo-liver nuclei were incubated with [3H]NAD in the presence of 1 mM spermine, 2.5 mM spermidine, or 3.5 mM putrescine, a 9-fold increase in poly)ADP-ribose) formation was observed. Nuclei treated with nuclease showed high poly(ADP-ribose) synthetase activity as spermine-treated nuclei. However, no further increase in the polymer formation by polyamines was detected in the nuclease-treated nuclei. We found that an increase in the polymer formation by spermine was the result of an increase in both chain length and chain number of the polymer at 2.3- and 6-fold, respectively. The major ADP-ribosylated proteins were determined as two non-histone proteins of Mr 130 000 and 70 000. The experiment of DNA synthesis with nuclei ADP-ribosylated in the presence of spermine showed a 7-fold increase in [3H]dTMP incorporation into the acid-inaoluble fraction. A similar stimulation was also found with nuclei treated with other polysmines, spermidine and putrescine, in the presence of NAD. These results indicate that DNA synthesis in growing tissues containing polyamines at high levels, such as is the case with tumors and the fetus, is stimulated by polyamine-mediated ADP-ribosylation of the nuclear proteins.

Animals↗
PubMed1988

The antidysrhythmic effect of metoprolol in the epinephrine treated chick embryo.

It was confirmed through electrocardiography that within two hours after epinephrine treatment, four day chick embryos either maintained normal rhythm or developed a severe cardiac dysrhythmia (22/93, 24% dysrhythmic). The ECG dysrhythmia in epinephrine treated embryos were characterized by periods of bradycardia, asystole, and various supraventricular or ventricular dysrhythmias. Within four hours after treatment, dysrhythmic embryos either reestablished normal rhythmicity or died. Electrocardiographic data also demonstrated that metoprolol pretreatment will block epinephrine induced dysrhythmias (0/46, 0% dysrhythmic). We conclude that metoprolol possesses antidysrhythmic properties in the epinephrine treated chick embryo.

Animals↗
PubMed2005

Perchloric acid-soluble protein regulates cell proliferation and differentiation in the spinal cord of chick embryos.

The role of perchloric acid-soluble protein (PSP) was investigated in chick embryos. Fluorescently labeled anti-chick liver (CL)-PSP IgG was injected into the yolk sac in ovo at embryonic day 3, and became localized in neuroepithelial cells. Within 12 h, morphological changes were observed in 37.5% of anti-CL-PSP IgG-injected embryos, and the neuroepithelial cells formed a wavy line. No significant changes were observed in embryos injected with non-immune IgG or PBS. Increased expression of PCNA and decreased expression of neuronal class III beta-tubulin were observed in the spinal cord after anti-CL-PSP IgG injection. These results suggest that PSP controls the proliferation and differentiation of neuroepithelial cells in chick embryos.

Animals↗
PubMed1990

Ontogeny of N-acetyltransferase activity rhythm in pineal gland of chick embryo.

1. N-acetyltransferase was present in pineal glands of 14-day-old chick embryos though no rhythm either in LL, DD or LD 12:12 was observed in this age. 2. Daily rhythm in pineal NAT activity was found in 18-day-old embryos incubated under LD 12:12 and LD 16:8 but no NAT rhythm was detected in DD or LL. 3. NAT rhythm persists for 2 days in constant darkness and it may be circadian in nature. 4. Presence of melatonin (85 +/- 8 pg/mg tissue) was detected in pineals of 18-day-old chick embryos.

Animals↗
PubMed2000

Epidermal growth factor regulates amino acid transport in chick embryo hepatocytes via protein kinase C.

System A-mediated amino acid transport, activation of different steps of signal transduction and involvement of different isoforms of protein kinase C (PKC) have been investigated in chick embryo hepatocytes after epidermal growth factor (EGF) stimulation. EGF rapidly (10 min) increased the rate of aminoisobutyric acid (AIB) uptake in chick embryo hepatocytes freshly isolated on the 19th day of embryonic life, while no change was detectable at other embryonal stages. The growth factor stimulation was abolished by PKC and tyrosine kinase inhibitors and was mimicked by 4-phorbol-12-myristate-13-acetate, dimethyl-2 (PMA). EGF treatment did not modify the phosphorylation of the isoform of phospholipase C (PLC-), and inositol trisphosphate (IP3) and intracellular calcium levels, but it induced an increase in PKC activity. Our data show that EGF regulates amino acid uptake, via PKC and without PLC- activation, only in the last period of chick embryo hepatocyte development. The effects of growth factor on PKC activity suggest the involvement of PKC- and - isoforms in EGF modulation of amino acid transport.

Amino Acids↗
PubMed1997

Glycoconjugate saccharidic moieties of the exocrine and endocrine pancreas in the chick embryo, newborn and adult.

A battery of horseradish peroxidase-conjugated lectins (PNA, SBA, DBA, WGA, ConA, LTA and UEAI) was used to study the distribution of glycoconjugate sugar residues in the exocrine and endocrine pancreas of chick embryos, 1-day-old chicks and adult animals. During the period of incubation considered here, the time of appearance and changes in the oligosaccharides were noted in the acinar cells, in the endocrine cells of the "light islets" and in the capillary endothelial cells. In the developing exocrine pancreas, the basolateral surface of the acinar cells reacted with PNA, WGA, LTA and ConA for the entire period of incubation while SBA, WGA, LTA and ConA reactivity was detected in the apical plasma membrane. Zymogen granule membrane reacted with PNA, WGA and LTA from the 12th day onward, while ConA reactivity was detectable from the 7th day onward. Owing to its early appearance, alpha-D-mannose, revealed by ConA lectin, might in some way contribute to the maturation of the zymogen granule. In the 1-day-old chick and in the adult the cellular surface reacted with WGA and ConA. The zymogen granule membrane reacted with PNA, WGA and ConA in the 1-day-old chick. In the adult, WGA and ConA reactivity was observed in the zymogen granule membrane, whereas at this site PNA reactivity was revealed only after neuraminidase digestion. The hypothesis formulated by others that granule membrane lectin reactivity is equal to the apical membrane reactivity, owing to exocytotic processes, is not consistent with our results in the chick embryos, the 1-day-old chick and the adults. The surface and the granules of the islet beta cells in different periods of incubation reacted with WGA, SBA and, after neuraminidase treatment, with PNA. The same reactivity was seen also in 1-day-old chick while in the adult the granules of the beta cells also reacted with ConA. These findings show the achievement of an almost adult-like state of glycosylation of the glycoconjugates in the beta cells at an early stages of development.

Aging↗
PubMed2000

Lipid-mediated gene transfection into chick embryo retinal cells in ovo and in vitro.

Several lipofection reagents were tested on chick embryo retinal cultures using green fluorescent protein (GFP) as a reporter gene; best results were obtained with the GenePORTER (GP) reagent, which yielded approximately 4.4% of the cells with intense GFP fluorescence. Cell survival and structural differentiation appeared normal, but one of the immunocytochemical markers studied (visinin) was less frequently observed in GP-treated cultures. When similar plasmid-GP mixtures were injected into chick embryo eyes in ovo, bright GFP-fluorescent cells were observed in different retinal layers, without detectable detrimental effects on retinal morphology. Particularly extensive reporter gene expression was obtained upon intraocular injection of GP plus naked DNA from a RCAS retrovirus, which resulted in the development of abundant radial columns of alkaline phosphatase-positive cells, separated by columns of negative cells. We conclude that lipid-based transfection offers a quick, simple and fairly innocuous means for gene delivery into proliferating and postmitotic retinal cells, in vitro as well as in the developing eye in ovo, and that transfection of naked retroviral DNA can lead to extensive expression of foreign genes by retinal cells, bypassing the time-consuming steps required for the generation of high-titer virion stocks.

Animals↗
PubMed2000

Thyroid hormone, glucagon, and medium-chain fatty acids regulate transcription initiated from promoter 1 and promoter 2 of the acetyl-CoA carboxylase-alpha gene in chick embryo hepatocytes.

High-carbohydrate feeding and triiodothyronine (T3) increase the abundance of acetyl-CoA carboxylase-alpha (ACC alpha) mRNA in avian hepatocytes, whereas starvation, glucagon, and medium-chain fatty acids decrease the abundance of ACC alpha mRNA. These changes in ACC alpha mRNA levels are mediated by alterations in the rate of transcription of the ACC alpha gene. In liver, ACC alpha transcription is initiated from two promoters, promoter 1 and promoter 2, resulting in transcripts that contain heterogeneity in their 5'-untranslated regions. Here, we investigated the role of promoter 1 and promoter 2 in mediating nutrient- and hormone-induced changes in ACC alpha mRNA abundance by measuring the level of transcripts expressed from promoter 1 and promoter 2 using a ribonuclease protection assay. The results indicated that both promoter 1 and promoter 2 were regulated by starvation/refeeding in livers of intact chicks and by T3, glucagon, and medium-chain fatty acids in chick embryo hepatocyte cultures and that alterations in the activity of promoter 2 accounted for a greater proportion of the changes in total ACC alpha mRNA abundance caused by nutrient and hormone treatment. Five DNase-hypersensitive sites were also identified between -500 and +1 bp relative to the transcription start site of promoter 2 in livers of intact chicks and in chick embryo hepatocyte cultures. In transient transfection analyses, this region of DNase hypersensitivity conferred regulation of transcription by T3, glucagon, and medium-chain fatty acids in chick embryo hepatocytes. Data from this study demonstrate that diet-induced changes in the activities of promoter 1 and promoter 2 in livers of intact chicks are mimicked in chick embryo hepatocyte cultures by manipulating the concentrations of T3, glucagon and medium-chain fatty acids in the culture medium and that cis-acting sequences mediating the effects of nutrients and hormones on promoter 2 activity are located immediately upstream of the transcription start site of this promoter.

Acetyl-CoA Carboxylase↗
PubMed1997

Modulation of proliferative activity and amino acid transport in chick embryo hepatocytes by EGF and retinoic acid.

In this work, the proliferative activity of chick embryo hepatocytes and its regulation by factors affecting cell growth, such as epidermal growth factor (EGF) and retinoic acid, were studied. The transport of nutritional molecules, like amino acids, was also investigated; in particular, the uptake mediated by the Na+ -dependent system A, known to be under hormonal and growth factor control. Moreover, some steps in the transduction pathway of mitogenic signal were analyzed, both for EGF and retinoic acid treatment. Results obtained suggest that chick embryo hepatocytes growth in an exclusive serum dependent way shows an early sensibility to EGF stimulation as regards to the proliferative activity and amino acid transport, responds to retinoic acid treatment and lack of contact inhibition. Moreover, as far as the signal transduction is concerned, at early stages, they do not seem to be able to utilize the same signal molecules as observed in adult life.

Amino Acids↗
PubMed1980

[Effect of low temperature on the ultrastructure of hepatocyte nuclei in 11-day-old chick embryos].

A study was made of the ultrastructure of nuclei of 11-day old chick embryos exposed to low temperature (-15 degrees C) for various periods of time and returned to the normal conditions of incubations (+37 degrees). The exposure to the low temperature induced some drastic ultrastructural alterations in the nuclear pore complex that involved the loss of lateral components of annulus (annular granules) and of the central granule. Some pores displayed a diaphragm or its structures (peripheral granules), whereas in other pores the pore complex material was seen totally disorganized. The degree of chromatin condensation in such nuclei increased. After the normal condition of chick embryo incubation was resumed, the ultrastructural organization of the chromatin and the pore complex appeared to be normalized. The pore complex reorganization occurred one hour after heating. It is suggested that the abnormal pore structure may correspond to an inactive state of the pore.

Animals↗
PubMed1990

1 alpha,25-dihydroxyvitamin D3 stimulates colony formation of chick embryo chondrocytes in soft agar.

The effect of vitamin D metabolites on the growth of chick embryo chondrocytes in soft agar was examined. 1,25-Dihydroxyvitamin D3 [1,25(OH)2D3] at 10(-8)-10(-7) M induced colony formation by chick embryo chondrocytes in soft agar in the presence of 10% fetal bovine serum. Furthermore, 1,25(OH)2D3 increased the number of colonies in the presence of a maximal dose of basic fibroblast growth factor, a potent mitogen for chondrocytes in soft agar. However, 24R,25 (OH)2D3 and other metabolites had little effect on the soft agar growth of chondrocytes in the presence or absence of basic fibroblast growth factor. These results suggest that 1,25(OH)2D3 is an active metabolite which may be involved in supporting cartilage growth.

Agar↗
PubMed1992

Cardiac mechanics in the stage-16 chick embryo.

A theoretical model is presented for the tubular heart of the stage-16 chick embryo (2.3 days of a 21-day incubation period). The model is a thick-walled, pseudoelastic cylindrical shell composed of three isotropic layers: the endocardium, the cardiac jelly, and the myocardium. The analysis is based on a shell theory that accounts for large deformation, material nonlinearity, residual strain, and muscle activation, with material properties inferred from available experimental data. We also measured epicardial strains from recorded motions of microspheres on the primitive right ventricles of stage-16 white Leghorn chick embryos. Relative to end diastole, peak axial and circumferential Lagrange strains occurred near end systole and had similar values. The magnitudes of these strains varied along the longitudinal axis of the heart (-0.16 +/- 0.08), being larger near the ends of the primitive right ventricle and smaller near midventricle. The in-plane shear strain was less than 0.05. Comparison of theoretical and experimental strains during the cardiac cycle shows generally good agreement. In addition, the model gives strong stress concentrations in the myocardial layer at end systole.

Animals↗