Development Of Chick Retina Research Articles

The autophagic machinery is necessary for removal of cell corpses from the developing retinal neuroepithelium

Autophagy is a homoeostatic process necessary for the clearance of damaged or superfluous proteins and organelles. The recycling of intracellular constituents also provides energy during periods of metabolic stress, thereby contributing to cell viability. In addition, disruption of autophagic machinery interferes with embryonic development in several species, although the underlying cellular processes affected remain unclear. Here, we investigate the role of autophagy during the early stages of chick retina development, when the retinal neuroepithelium proliferates and starts to generate the first neurons, the retinal ganglion cells. These two developmental processes are accompanied by programmed cell death. Upon treatment with the autophagic inhibitor 3-methyladenine, retinas accumulated numerous TdT-mediated dUTP nick-end labelling-positive cells that correlated with a lack of the 'eat-me' signal phosphatidylserine (PS). In consequence, neighbouring cells did not engulf apoptotic bodies and they persisted as individual cell corpses, a phenotype that was also observed after blockade of phagocytosis with phospho-L-Serine. Supplying the retinas with methylpyruvate, a cell-permeable substrate for ATP production, restored ATP levels and the presentation of PS at the cell surface. Hence, engulfment and lysosomal degradation of apoptotic bodies were also re-established. Together, these data point to a novel role for the autophagic machinery during the development of the central nervous system.

Melatonin inhibitory effect on cAMP accumulation in the chick retina development

During vertebrate neurodevelopment, neuritogenesis and synaptogenesis are modulated by intracellular cAMP rises. Melatonin, which is implicated in neuronal differentiation, mainly inhibits this pathway. Here, an investigation about the profile of this effect during the vertebrate neurodevelopment is reported. In the embryonic chick retinas at days 8, 12, 14, 16 and at 2 days post-hatched (E8, E12, E14, E16 and PH), those control embryonic retinas incubated only with the phosphodiesterase inhibitor at days corresponding to commencement of neuronal differentiation (E8, E12) and PH, presented cAMP levels inhibited by melatonin. While the cAMP accumulation stimulated by forskolin was inhibited in the embryonic retinas at all testing days. Neither the unselective antagonist N-acetyl-2-benziltryptamine (luzindole) nor the selective Mel1b antagonist 4-phenyl-2-propionamidotetralin (4-P-PDOT) blocked the melatonin concentration-dependent inhibitory effect on cAMP accumulation in the retinas initiating differentiation (E7−E9), suggesting a tight binding between melatonin and their receptors. However, 4-P-PDOT competitively reverted the melatonin effect on cAMP stimulated by forskolin during synaptogenesis stages. Together, the melatonin effect on cAMP levels in chick retina, which is mainly through melatonin receptors, is depending on the developmental period observed, probably taking part in the mechanisms surrounding the melatonin action on neuronal differentiation.

Involvement of L-DNase II in Nuclear Degeneration during Chick Retina Development

During the development of the neural retina, 50% of the neurons die physiologically by apoptosis. In the chick embryo, the apoptotic wave starts at E8 and ends at E18, with a peak at E11. The onset of apoptosis is accompanied by the activation of several degradative enzymes. Among these, the activation of the endonucleases leads to the degradation of the genomic DNA of the cell which is thought to be the final event in apoptosis. Here, we have investigated the endonucleases activated during apoptosis associated with retinal development. We have found that Ca2+-Mg2+-dependent endonucleases, as well as acid endonucleases are activated. The results obtained in vitro using purified nuclei from chicken retina indicate that the endonuclease activity resulting from the activation of L-DNase II, an acid DNase is responsible for most of the DNA degradation observed in these cells.

Developmental regulation of group iii metabotropic glutamate receptors modulating adenylate cyclase activity in the avian retina

The regulation of adenylate cyclase by neurotransmitters is observed in early development of the chick retina. In the present work we show that L-2-amine-4-phosphonobutyric acid (L-AP4), the major agonist of group III metabotropic glutamate receptors (mGluRs), inhibits the accumulation of cyclic AMP induced by forskolin in the chick retina. This effect is observed after 8 days of development (E8), is maximal from E12–E17 and decreases at the post-hatching period (PH). The inhibition is also observed in cultures of retinal cells incubated for 2–8 days. We have also investigated the interaction between group III mGluRs and other receptors coupled to adenylate cyclase in the developing retina. The inhibition by L-AP4 is partially additive with that induced by the A1 adenosine agonist Cyclohexyladenosine and is not observed when cyclic AMP levels are increased with 2-chloroadenosine or dopamine. The group II mGluR agonist trans-(1S,3R)-1-amino-cyclopentanedicarboxylic acid has an inhibitory effect only on PH retinas, indicating that group II and group III mGluRs have a differential ontogenesis in this tissue. The results show that Group III mGluRs are expressed early during chick retina development and do not interact with other receptors known to be coupled to adenylate cyclase in the developing retina.

Biochemical differentiation of nascent neurite junctions: unilateral localization of adheron components

Previous work showed that adhesive contacts made by growth cone filopodia involve extracellular filaments that can be labeled with an antiserum to adherons (adhesion-promoting complexes in conditioned medium). Here, adheron antigens were found to be differentially expressed by particular cell types and synaptic layers during chick retina development, and this differential expression at the cellular level was retained in culture. When applied to living cells, adheron antibodies characteristically patched at filopodial junctions. Monospecific antibodies to purpurin and a heparan sulphate proteoglycan (HSPG), two components of adherons, labeled a subpopulation of junctions. Most interestingly, anti-purpurin and anti-HSPG bound only to one end of adhesive filaments. Such localization to discrete microdomains provides support for a heterotypic adhesive mechanism in junction formation.

Effect of p-mercuribenzoate on the subestimation of angiotensin-converting enzyme measurement during chick retina development

The time course of dipeptidase activity and the effect of p-mercuribenzoate (PCMB) on the subestimation of the fluorometric determination of angiotensin-converting enzyme (ACE, EC 3.4.15.1) during development was studied. ACE and dipeptidase activities were measured fluorometrically in homogenates of the developing chick retina using Hip-His-Leu and His-Leu as substrates, respectively, both either in the presence or in the absence of 1 mM PCMB. ACE activity was inhibited by captopril (IC 50 1.7 nM), MK 422 (IC 50 4.8 nM), BPP 9a (IC 50 0.25 μM) and BPP 5a (IC 50 1.2 μM), thus suggesting that avian retinal ACE catalytically resembles the mammalian enzyme. Dipeptidase activity varied 3.4-fold throughout development, leading to a large and variable (28–83%) subestimation of ACE activity during chick retina ontogenesis. PCMB (1 mM) inhibited 67–94% dipeptidase activity during development, thus greatly reducing any subestimation of ACE activity determination during the development of the chick retina.

Binding of FITC-coupled peanut-agglutinin (FITC-PNA) to embryonic chicken retinas reveals developmental spatio-temporal patterns.

FITC-labeled peanut agglutinin (PNA), a lectin from peanut (Arachis hypogaea) which binds specifically to D-galactose, was used as a probe to trace the distribution of PNA-lectin receptors during early development of chick retina. We found that specific fluorescence within the inner plexiform layer (IPL) and the outer plexiform layer (OPL) appears at different developmental stages. The specific fluorescence of the IPL first appears near the central area of the ventro-posterior quadrant of the 7-day retina, then extends progressively towards the periphery, reaching the ora serrata earlier on the posterior than on the anterior side. The development of the specific fluorescence of the OPL follows the same scheme, but it appears later (starting at day 9) than that of the IPL and it is less pronounced. The spatio-temporal patterns of the expression of the lectin receptors show relationships to those of withdrawal of proliferating cells from mitosis, and of other events related to differentiation.

Membrane alterations during chick neural retina development: A freeze-fracture study

As part of a study of cell surface differentiation during chick retina development, a freeze-fracture study of neural retinas from 5 to 10 day embryonic chicks was undertaken. Three classes of changes have been detected. (1) As cells differentiate and become recognizable by their position within the tissue, they acquire characteristic numbers of intramembrane particles in the surfaces in each layer. (2) Small gap junctions appear between cells at the outer limiting membrane of the 5 day retina. At 6 days, they are larger, more numerous and are also found in deeper layers of the tissue. By the seventh day, the size and number of the junctions is greatly reduced; they are not visible after the tenth day. (3) The characteristic lack of particles in the outer limiting membrane of the mature retina appears at the ninth day of incubation, at the time that presumptive photoreceptors extend through the outer limiting membrane. Tight junctions between cells were not observed during this study.