Embryo Retina Research Articles

Alterations of the retina in chick embryos induced by systemic alpha-bungarotoxin application.

The application of alpha-bungarotoxin onto the chorio-allantoic membrane of chick embryos between the 11th and 18th day of incubation leads to alterations of retinal development. The most significant qualitative change is the appearance of retinal rosettes formed by receptor cells. These rosettes are infoldings of the receptor cell layer. Quantitatively, an enlargement in volume of the receptor and outer nuclear layer can be found together with a simultaneous decrease of the other retinal layers. The toxin seems to suspend the naturally occurring nerve cell death in the receptor cell population

Biosynthesis of gangliosides in cultured retina from chick embryos.

Retina tissue from 7-day chick embryos was maintained in culture for up to 10 days. After 5 days in culture the incorporation of [3H]leucine into proteins and of [3H]glucosamine into gangliosides was similar to that found in retinas from 12-day embryos. The incorporation of [3H]thymidine into DNA decreased steadily with time in culture; after 5 days it was about 20% of the initial value and approximately twice that determined in retinas from 12-day embryos. The radioactivity pattern of gangliosides labeled with [3H]glucosamine showed a predominance of the label in disialosyllactosylceramide (GD3); up to the 3rd day of culture. From then on, there was a progressive increase in the labeling of disialosylgangliotetraosylceramide (GD1a); by day 7 of culture, labeling of GD1a predominated and the labeling pattern was indistinguishable from that found in retinas from 12-day-old embryos. The specific activities of the CMP-NeuAc:GM3 sialosyl- and UDP-GalNAc:GM3 N-acetylgalactosaminyl-transferases decreased to 15% and increased to 400%, respectively, of the values determined in the retinas of 7-day embryos. The cultured retinas progressed in their organization into layers with culture time. The labeling transition from GD3 to GD1a was also detected after inhibition of the histotypic organization by addition of 5-bromo 2-deoxyuridine to the culture medium. Results suggest that high activity of GM3:sialosyl transferase and high labeling of GD3 are associated with the proliferative state of retina cells, while high activity of GM3:N-acetylgalactosaminyltransferase and high labeling of GD1a are associated with the non-proliferative, differentiated state of these cells.

Ultrastructure of Müller cells in the developing human retina.

The posterior retina of human embryos from 4 to 200 mm of crown-rump length was studied by electron microscopy. At 20 mm dense inner Müller-cell processes near ganglion cells contained rough endoplasmic reticulum, free ribosomes, small matrix particles, and some intermediate filaments. These processes soon had smooth endoplasmic reticulum. By 71 mm many of these inner processes were lucent and contained many intermediate filaments and glycogen particles. Müller-cell nuclei and outer processes were observed between differentiating cone cells at 66 mm, and these outer radial-cell processes soon contained many dense matrix particles and glycogen particles. As neurons in the inner nuclear layer differentiated by 100 mm, Müller-cell cytoplasm in the mid-retina was identified by its intermediate filaments and glycogen particles. Müller cells have composite glial features that appear in the horizontal retinal layers concomitant with neuronal differentiation and maturation in each layer.

The pattern of expression of chick delta-crystallin genes in lens differentiation and in trans-differentiating cultured tissues.

During development of the vertebrate lens, the lens epithelium undergoes a final stage of differentiation into lens fibre cells. Lens fibre cells can also be produced by trans-differentiation from certain extralenticular structures, all of which are of different developmental origin from lens, including embryonic neural retina and retinal pigmented epithelium. Delta-crystallin is the major lens protein in the chick and appears first in development; it is the major product in trans-differentiated retina of younger embryos. In both normal differentiation and trans-differentiation an increase of delta-crystallin coding RNA is detectable in the nucleus of cells prior to their terminal differentiation into lens fibres. The increase in transcription of delta-crystallin genes accompanying final differentiation of lens fibres, appears to take place slightly in advance of an increase in the capacity to process and transport this mRNA to the cytoplasm.

Formation of gap junctions by stem cells in the developing retina of the clawed frog (Xenopus laevis)

To ascertain whether gap junctions are preferentially formed by proliferated retinal stem cells, an analysis by electron microscopic autoradiography was made on the retina of the Xenopus embryo at stages 26, 29 and 31, after pulse labelling with 3H-thymidine. When the administration of the isotope was carried out for 0.5 or 3 h, retinal cells in S phase or S to M phase in the cell cycle were marked with the isotope, respectively. In these specimens, most gap junctions were found on the isotope-labelled cells, and few on the unlabelled cells. Several cells in mitosis also formed gap junctions. These results would suggest that gap junctions are formed by retinal stem cells in S to M phases of the cell cycle, but not by cells in G1 phase and post-mitotic cells.

A quantitative study of the position of the Golgi apparatus in the early developing chick eye.

A quantitative evaluation of the positional changes of the Golgi apparatus during the invagination of the lens placode and the presumptive neural retina of the chick embryo was carried out by silver impregnation. The Golgi apparatus is predominantly polarized in the apical process of the elongated interphasic cells; however, basal and lateral positions are also present. When the nuclei are located close to the luminal surface, basal and lateral positions increase significantly showing that a loss of Golgi apparatus polarization takes place, associated with the mitotic cycle. At the beginning of the invagination of lens and neural retina rudiments, a marked shift in orientation occurs between lateral and apical positions of the Golgi apparatus; this is associated with the nuclei located close to the luminal surface. The possible significance of these results is related to some processes concerning the invagination of these rudiments, such as interkinetic nuclear migration and secretion of extracellular materials.

Glutamate decarboxylase activity in chick brain and retina

We have studied the effect of Triton-X-100 on glutamate decarboxylase (GAD) activity in brain and retina from chick embryos of 12 and 16 days' incubation and from chicks 4--6 weeks old. GAD activity was measured in five different homogenization media. Triton-X-100 inhibited the enzyme by about 60% in both brain and retina of 12-day embryos and by about 50% in 16-day embryos, independently of the homogenization medium. In chicks only about 20% inhibition by the detergent was observed in brain whereas no effect was found in retina. These results indicate that the evaluation of the experimental conditions of enzyme assays at different ages is essential for developmental studies of GAD activity in nervous tissue.

The environment of axonal migration in the developing chick retina: A scanning electron microscopic (SEM) study

We have made a SEM study of the basal intercellular spaces of the retina in chick embryos of different developmental stages. Since this is the environment where optic axons grow, the structural characteristics of this region might play some role in the orientation of axonal migration towards the choroid fissure. The basal region of undifferentiated retinas is formed by the vitreal expansions of neuroepithelial cells. In pre-axonal stages, the intercellular spaces between these expansions do not show any preferential orientation towards the fissure. The growth cones of ganglion cell axons appear in an apicobasal direction and turn towards the fissure immediately beneath the vitreal surface. Fasciculation is an early event during development and, in the more advanced stages, the vitreal expansions from retinal cells are placed in rows following the same orientation as the axon bundles. These observations are discussed in relationship to current hypotheses on axonal migration and orientation.

INDOLEAMINE ACCUMULATING NEURONS IN THE RETINA OF CHICKEN AND PIGEON: A Comparison with the Dopaminergic Neurons

Recently a special group of indoleamine accumulating neurons has been described in the retina of some mammals and goldfish. These neurons are characterized by their ability to accumulate indoleamines, whereby they become visible in the fluorescence microscope. They do not show any spontaneuos fluorescence. The indoleamine accumulating neurons are in this study shown to be present in the retina of chicken and pigeon. Their cell bodies differ from the earlier described cell bodies of the same type in other species in being larger and bottle shaped instead of round or oval, and in being situated further cutwards in the inner nuclear layer. Their terminals ramify in three sublayers in the inner plexiform layer. No indoleamine containing neurons could, however, be seen to fluoresce in normal retina of chick embryos, newborn chicken, older chicken or pigeons.

Effects of N2, O2′-dibutyril cyclic GMP on the nucleoside phosphotransferase activity of the retina of the chick embryos

In the retina of the chick embryo, 2 different forms of nucleoside phosphotransferase take part in the phosphorylation of thymidine. One is an unstable form with higher molecular weight. The other with lower m. wt is a stable form. This paper shows that N2,O2'-dibutyril cyclic GMP causes a marked decrement of the activity of the unstable nucleoside phosphotransferase.

Isolation of retina-specific cell-aggregating factor from membranes of embryonic neural retina tissue.

There is increasing evidence that developmental associations among embryonic cells are mediated by specific components of the cell surface. Earlier work has indicated that such components are extruded into the medium of primary monolayer cultures of embryonic cells, and that they represent the active constituents of the tissue-spedific cell-aggregating factors isolated fro- the supernatant medium of such cultures. We presently report that tissue-specific cell-aggregating factors can be obtained directly from embryonic tissues, and describe the isolation and partial purification of retina-spedific factor from a cell-membrane preparation derived from retina tissue of the chick embryo. Extraction of the purified membrane preparation with 1-butanol yielded an activity in the aqueous phase which resides in a protein probably a glycoprotein, with an estimated molecular weight of 50,000 in solution. This material could be obtained from retinas of embryos not older than 13 days, and only pre-13-day cells responded to its cell-aggregating activity. By these characteristics, this membrane-derived retina factor closely resembles the retina cell-aggregating glycoprotein previously purified from the supernatant medium of retina cell cultures. It is of special interest that the cell-aggregating protein is obtainable from cellular membranes during those stages of development when retina cells are most actively engaged in histological organization. Work in progress indicates that, by the procedures described herein, tissue-specific cell-aggregating factors can also be obtained from membrane preparations of other embryonic tissues.

Interaction of cortisol with the neural retina of the chick embryo in culture

The uptake of cortisol and the kinetics of hormone-receptor interaction in the cytosol and cell nucleus were investigated in the intact tissue in organ culture. Cortisol is concentrated by the neural retina. The accumulation of the free steroid is temperature dependent but the effect of temperature decreases with the increase of cortisol in medium. Cortisol binding to specific receptors in the cytosol shows a sigmoidal type of kinetics which correlates well with the kinetics of glutamine-synthetase induction by cortisol. The temperature dependent translocation of the receptor-hormone complexes to the nuclei and the effect of detergents on the binding to nuclei are presented.

Enzyme induction by corticosteroids in embryonic cells: steroid structure and inductive effect.

Glutamine synthetase in the developing retina of the chick embryo can be induced to increase by certain corticosteroids. The inductive effectiveness of various natural corticosteroids has been examined in organ cultures of embryonic retina and correlated with specific groupings on the steroid molecules.

CONTROL OF GLUTAMINE SYNTHETASE IN THE EMBRYONIC RETINA IN VITRO.

Glutamine synthetase activity in the neural retina of the chick embryo increases sharply during terminal differentiation of this tissue. This characteristic increase can be reproduced in cultures of retinal tissue fragments from late embryos. A similarly sharp increase can be elicited precociously in younger retina by culturing in medium with adult serum. Both the precociously elicited and the later increase in enzyme activity require continuous protein synthesis; both can be suppressed during the first 24 hours of culture by blocking RNA synthesis or by removing the adult serum. Subsequently, the increase in enzyme activity becomes progressively less dependent on RNA synthesis and on the continuous presence of adult serum. This transition is attained more rapidly in the older retina. The data suggest a progressive stabilization of the enzyme-forming system during differentiation.