Distribution of the small molecular weight GTP-binding proteins Rac1, Cdc42, RhoA and RhoB in the developing chick retina

Immunohistochemistry was used to determine the distribution of Rac1, Cdc42, RhoA and RhoB GTPases during development of the chick retina. All proteins appear as early as embryonic day 5 (E5) in cells of the vitreal margin, E7–8 in cells of the inner third of the inner nuclear layer and E9–10 in photoreceptors. From E10 until hatching, RhoA, Rac1 and Cdc42 were seen in perikarya and/or processes of amacrine, ganglion cells, and photoreceptors. Rho proteins were also observed in retinal Müller cells, with different distributions. RhoB showed a transient expression, being severely down regulated after E18. The distribution pattern of Rho proteins during the development of the chick retina suggests a concerted role in the differentiation of specific cell types, and probably during synaptogenesis.     

The morphology and coupling of Aplysia bag cells within the abdominal ganglion and in cell culture

The bag cells in the abdominal ganglion of Aplysia californica control egg-laying behavior by releasing a polypeptide (ELH) during an afterdischarge of synchronous action potentials. We have used intracellular injection of Lucifer Yellow to study the morphology and interconnections of the bag cells. These neurosecretory cells are typically multipolar and their processes extend in all directions out from the bag cell clusters into the surrounding connective tissue, where they branch in a complex manner. In some of the dye injection experiments, dye transfer from the injected cell to neighboring cells was observed. Freeze fracture of the bag cell clusters and their surrounding connective tissue revealed numerous gap junctions on bag cell processes within the clusters as well as on more distal processes. We have also examined the morphology and coupling between bag cells in primary culture. As in the intact ganglion, bag cells in culture were found to be multipolar. All pairs of bag cells whose somata or processes had formed contacts in culture were electrically coupled. The strongest coupling was observed between pairs of cells whose somata appeared closely apposed. In these cases transfer of Lucifer Yellow between cells could also be observed. It is therefore likely that the synchrony of bag cell action potentials during a bag cell afterdischarge is a result of coupling between individual cells in the bag cell cluster.     

The early development of retinal ganglion cells with uncrossed axons in the mouse: retinal position and axonal course

The carbocyanine dye, DiI, has been used to study the retinal origin of the uncrossed retinofugal component of the mouse and to show the course taken by these fibres through the optic nerve and chiasm during development. Optic axons first arrive at the chiasm at embryonic day 13 (E13) but do not cross the midline until E14. After this stage, fibres taking an uncrossed course can be selectively labelled by unilateral tract implants of DiI. The earliest ipsilaterally projecting ganglion cells are located in the dorsal central retina. The first sign of the adult pattern of distribution of ganglion cells with uncrossed axons located mainly in the ventrotemporal retina is seen on embryonic day 16.5, thus showing that the adult line of decussation forms early in development. A small number of labelled cells continue to be found in nasal and dorsal retina at all later stages. At early stages (E14-15), retrogradely labelled uncrossed fibres are found in virtually all fascicles of the developing nerve, intermingling with crossed axons throughout the length of the nerve. At later stages of development (E16-17), although uncrossed fibres pass predominantly within the temporal part of the stalk, they remain intermingled with crossed axons. A significant number of uncrossed axons also lie within the nasal part of the optic stalk. The position of uncrossed fibres throughout the nerve in the later developmental stages is comparable to that seen in the adult rodent (Baker and Jeffery, 1989). The distribution of uncrossed axons thus indicates that positional cues are not sufficient to account for the choice made by axons when they reach the optic chiasm.     

Multiple Neuronal Connexins in the Mammalian Retina

Gap junctions are abundant in the mammalian retina and many neuronal types form neural networks. Several different neuronal connexins have now been identified in the mammalian retina. Cx36 supports coupling in the AII amacrine cell network and is essential for processing rod signals. Cx36 is probably also responsible for photoreceptor coupling. Horizontal cells appear to be extensively coupled by either Cx50 or Cx57. These results indicate that multiple neuronal connexins are expressed in the mammalian retina and that different cell types express different connexins.     

Multiple neuronal connexins in the mammalian retina

Gap junctions are abundant in the mammalian retina and many neuronal types form neural networks. Several different neuronal connexins have now been identified in the mammalian retina. Cx36 supports coupling in the AII amacrine cell network and is essential for processing rod signals. Cx36 is probably also responsible for photoreceptor coupling. Horizontal cells appear to be extensively coupled by either Cx50 or Cx57. These results indicate that multiple neuronal connexins are expressed in the mammalian retina and that different cell types express different connexins.     

Expression of membrane-bound and soluble guanylyl cyclase mRNAs in embryonic and adult retina of the medaka fish Oryzias latipes

Localization of mRNAs for four membrane-bound guanylyl cyclases (membrane GCs; OlGC3, OlGC4, OlGC5, and OlGC-R2), three soluble guanylyl cyclase subunits (soluble GC; OlGCS-alpha(1), OlGCS-alpha(2), and OlGCS-beta(1)), neuronal nitric oxide synthase (nNOS), and cGMP-dependent protein kinase I (cGK I) was examined in the embryonic and adult retinas of the medaka fish Oryzias latipes by in situ hybridization. All of the membrane GC mRNAs were detected in the photoreceptor cells of the adult and embryonic retinas, but in different parts; the OlGC3 and OlGC5 mRNAs were expressed in the proximal part and the OlGC4 and OlGC-R2 mRNAs were expressed in the outer nuclear layer. The mRNA for nNOS was expressed in a scattered fashion on the inner side of the inner nuclear layer in the adult and embryonic retinas. The mRNAs (OlGCS-alpha(2) and OlGCS- beta(1)) of two soluble GC subunits (alpha(2) and beta(1)) were expressed mainly in the inner nuclear layer and the ganglion cell layer of the embryonic retina while the mRNAs of the soluble GC alpha(1) subunit and cGK I were not detected in either the adult or embryonic retina. These results suggest that NO itself and/or the cGMP generated by soluble GC (alpha(2)/beta(1) heterodimer) play a novel role in the neuronal signaling and neuronal development in the medaka fish embryonic retina in addition to the role played by phototransduction through membrane GCs in the adult and embryonic retinas.     

Connexin43 and connexin45 form gap junctions with different molecular permeabilities in osteoblastic cells.

We examined the expression and function of gap junctions in two rat osteoblastic cell lines, ROS 17/2.8 and UMR 106-01. The pattern of expression of gap junction proteins in these two cell lines was distinct: ROS cells expressed only connexin43 on their cell surface, while UMR expressed predominantly connexin45. Immunoprecipitation and RNA blot analysis confirmed the relative quantitation of these connexins. Microinjected ROS cells passed Lucifer yellow to many neighboring cells, but UMR cells were poorly coupled by this criterion. Nevertheless, both UMR and ROS cells were electrically coupled, as characterized by the double whole cell patch-clamp technique. These studies suggested that Cx43 in ROS cells mediated cell-cell coupling for both small ions and larger molecules, but Cx45 in UMR cells allowed passage only of small ions. To demonstrate that the expression of different connexins alone accounted for the lack of dye coupling in UMR cells, we assessed dye coupling in UMR cells transfected with either Cx43 or Cx45. The UMR/Cx43 transfectants were highly dye coupled compared with the untransfected UMR cells, but the UMR/Cx45 transfectants demonstrated no increase in dye transfer. These data demonstrate that different gap junction proteins create channels with different molecular permeabilities; they suggest that different connexins permit different types of signalling between cells.     

Differential, age-dependent MEK-ERK and PI3K-Akt activation by insulin acting as a survival factor during embryonic retinal development

Programmed cell death is a genuine developmental process of the nervous system, affecting not only projecting neurons but also proliferative neuroepithelial cells and young neuroblasts. The embryonic chick retina has been employed to correlate in vivo and in vitro studies on cell death regulation. We characterize here the role of two major signaling pathways, PI3K-Akt and MEK-ERK, in controlled retinal organotypic cultures from embryonic day 5 (E5) and E9, when cell death preferentially affects proliferating neuroepithelial cells and ganglion cell neurons, respectively. The relative density of programmed cell death in vivo was much higher in the proliferative and early neurogenic stages of retinal development (E3-E5) than during neuronal maturation and synaptogenesis (E8-E19). In organotypic cultures from E5 and E9 retinas, insulin, as the only growth factor added, was able to completely prevent cell death induced by growth factor deprivation. Insulin activated both the PI3K-Akt and the MEK-ERK pathways. Insulin survival effect, however, was differentially blocked at the two stages. At E5, the effect was blocked by MEK inhibitors, whereas at E9 it was blocked by PI3K inhibitors. The cells which were found to be dependent on insulin activation of the MEK-ERK pathway at E5 were mostly proliferative neuroepithelial cells. These observations support a remarkable specificity in the regulation of early neural cell death.     

Differential expression of connexins during histogenesis of the chick retina

Gap junction (GJ) channels couple adjacent cells, allowing transfer of second messengers, ions, and molecules up to 1 kDa. These channels are composed by a multigene family of integral membrane proteins called connexins (Cx). In the retina, besides being essential circuit element in the visual processing, GJ channels also play important roles during its development. Herein, we analyzed Cx43, Cx45, Cx50, and Cx56 expression during chick retinal histogenesis. Cx exhibited distinct expression profiles during retinal development, except for Cx56, whose expression was not detected. Cx43 immunolabeling was observed at early development, in the transition of ventricular zone and pigmented epithelium. Later, Cx43 was seen in the outer plexiform and ganglion cell layers, and afterwards also in the inner plexiform layer. We observed remarkable changes in the phosphorylation status of this protein, which indicated modifications in functional properties of this Cx during retinal histogenesis. By contrast, Cx45 showed stable gene expression levels throughout development and ubiquitous immunoreactivity in progenitor cells. From later embryonic development, Cx45 was mainly observed in the inner retina, and it was expressed by glial cells and neurons. In turn, Cx50 was virtually absent in the chick retina at initial embryonic phases. Combination of PCR, immunohistochemistry and Western blot indicated that this Cx was present in differentiated cells, arising in parallel with the formation of the visual circuitry. Characterization of Cx expression in the developing chick retina indicated particular roles for these proteins and revealed similarities and differences when compared to other species.     

Gap junction communication and connexin 43 gene expression in a rat granulosa cell line: regulation by follicle-stimulating hormone

Follicle-stimulating hormone is the major regulator of growth and development of antral follicles in the ovary. Granulosa cells (GCs) in these follicles are coupled via gap junctions (GJs) consisting of connexin 43 (Cx 43). Because we and others have found that Cx 43 and GJs, respectively, are more abundant in large antral follicles compared with small antral and preantral follicles, we hypothesized that FSH may control Cx 43 gene expression, GJ formation, and intercellular communication. To directly address these points, we chose a rat GC line (GFSHR-17) expressing the FSH receptor and the Cx 43 gene. The functionality of FSH receptors was shown by the effects of porcine FSH, namely cell rounding, reduced cellular proliferation, and stimulation of progesterone production of GFSHR-17 cells, which are effects that were detectable within hours. Treatment with FSH also statistically significantly increased Cx 43 mRNA levels, as shown after 6 to 9 h in Northern blots. These effects were antedated by altered GJ communication, which was observed within seconds. Using a single-cell/whole-cell patch clamp technique, we showed that FSH rapidly and reversibly enhanced electrical cell coupling of GFSHR-17 cells. Increased GJ communication was associated with statistically significantly decreased phosphorylation of Cx 43, which was observed within 10 min after FSH addition, during immunoprecipitation experiments. Our results demonstrate, to our knowledge for the first time, that the gonadotropin FSH acutely and directly stimulates intercellular communication of GFSHR-17 cells through existing GJs. Moreover, FSH also increases levels of Cx 43 mRNA. These changes are associated with reduced proliferation and enhanced differentiation of GFSHR-17 cells. In vivo factors in addition to FSH may be involved in the regulation of GJ/GJ communication between GCs in the follicle, but our results suggest that improved cell-to-cell coupling, enhanced Cx 43 gene expression, and possibly, formation of new GJs are direct consequences of FSH receptor activation and may antedate and/or initiate the pivotal effects of FSH on GCs.     

Developmental expression of dynamin in the chick retinotectal system.

Dynamin I, a GTPase involved in the endocytic cycle of synaptic vesicle membranes, is believed to support axonal outgrowth and/or synaptogenesis. To explore the temporal and spatial patterns of dynamin I distribution in neuronal morphogenesis, we compared the developmental expression of dynamin with the expression of presynaptic membrane proteins such as SV2, synaptotagmin, and syntaxin in the chick primary visual pathway. Western blots of retina and tectum revealed a steady increase of synaptotagmin and syntaxin from embryonic Day 7 (E7) to E11, whereas for the same time frame no detectable increase of dynamin was found. Later stages showed increasing amounts of all tested proteins until the first postnatal week. Immunofluorescence revealed that SV2, synaptotagmin, and syntaxin are present in retinal ganglion cell axons from E4 on. In later stages, the staining pattern in the retina and along the visual pathway paralleled the formation and maturation of axons. In contrast, dynamin is not detectable by immunofluorescence in the developing retina and optic tectum before synapse formation. Our data indicate that, in contrast to the early expression of synaptotagmin, SV2, and syntaxin during axonal growth, dynamin is upregulated after synapse formation, suggesting its function predominantly during and after synaptogenesis but not in axonogenesis.(J Histochem Cytochem 47:1297-1306, 1999)     

High levels of extracellular glutamate are present in retina during neonatal development

The three major classes of neurons which comprise the primary visual pathway in retina are glutamatergic. These cells are generated in two separate developmental stages, with one subclass of photoreceptors (cones) and ganglion cells generated before birth; and the other subclass of photoreceptors (rods) and bipolar cells generated during the first week after birth. Gas chromatography/mass spectroscopy analysis coupled with a new method for collecting small samples of extracellular fluids from retina were used to determine the levels of endogenous glutamate present during differentiation and synaptogenesis of these different cell types. As expected the total retinal content of glutamate increased during the postnatal period in synchrony with the generation and maturation of glutamatergic cells. However, a significant proportion of the endogenous pool was found extracellularly at birth. Intracellular glutamate is localized within cell bodies and growing processes of cones and ganglion cells at this time but few glutamatergic synapses are present. The extracellular concentration of glutamate actually declined during the most active period of synaptogenesis, reaching very low levels in the adult. The high concentrations of extracellular glutamate in neonatal retina could play an important role in a variety of developmental events such as dendritic pruning, programmed cell death and neurite sprouting. Special issue dedicated to Dr. Kinya Kuriyama.     

Spatial Distribution of Excitatory Synapses on the Dendrites of Ganglion Cells in the Mouse Retina

Excitatory glutamatergic inputs from bipolar cells affect the physiological properties of ganglion cells in the mammalian retina. The spatial distribution of these excitatory synapses on the dendrites of retinal ganglion cells thus may shape their distinct functions. To visualize the spatial pattern of excitatory glutamatergic input into the ganglion cells in the mouse retina, particle-mediated gene transfer of plasmids expressing postsynaptic density 95-green fluorescent fusion protein (PSD95-GFP) was used to label the excitatory synapses. Despite wide variation in the size and morphology of the retinal ganglion cells, the expression of PSD95 puncta was found to follow two general rules. Firstly, the PSD95 puncta are regularly spaced, at 1–2 µm intervals, along the dendrites, whereby the presence of an excitatory synapse creates an exclusion zone that rules out the presence of other glutamatergic synaptic inputs. Secondly, the spatial distribution of PSD95 puncta on the dendrites of diverse retinal ganglion cells are similar in that the number of excitatory synapses appears to be less on primary dendrites and to increase to a plateau on higher branch order dendrites. These observations suggest that synaptogenesis is spatially regulated along the dendritic segments and that the number of synaptic contacts is relatively constant beyond the primary dendrites. Interestingly, we also found that the linear puncta density is slightly higher in large cells than in small cells. This may suggest that retinal ganglion cells with a large dendritic field tend to show an increased connectivity of excitatory synapses that makes up for their reduced dendrite density. Mapping the spatial distribution pattern of the excitatory synapses on retinal ganglion cells thus provides explicit structural information that is essential for our understanding of how excitatory glutamatergic inputs shape neuronal responses.     

Expression and function of the neurotransmitter serotonin during development of the Helisoma nervous system

Exogenous serotonin has been shown to evoke a neuron-selective inhibition of neurite outgrowth and synaptogenesis in identified Helisoma neurons in vitro. We demonstrate here that serotonin is present in the embryonic nervous system of Helisoma and can act as a regulator of neuronal development in vivo. Serotonin-like immunoreactivity was first observed in neurons at an early stage of nervous system development (E20). Throughout embryogenesis, the number of serotonin-immunoreactive neurons increased in a stereotypic pattern that was unique for each type of ganglion. Strikingly, the number of serotonin-immunoreactive neurons continued to increase throughout adult life. Transient perturbation of endogenous serotonin levels during embryogenesis had profound effects on the development of specific identified neurons. Embryos treated with 5,7-dihydroxytryptamine and raised to maturity showed aberrations in neuronal morphology, neuronal dye coupling, and strength of electrical synaptic connections. These effects were restricted to neurons known to be sensitive to the growth-inhibitory effects of serotonin in vitro. These results support the hypothesis that neurotransmitters are an important class of regulatory factors during normal development of the nervous system.     

Nitric oxide specifically reduces the permeability of Cx37-containing gap junctions to small molecules

Gap junction intercellular communication (GJIC) plays a significant role in the vascular system. Regulation of GJIC is a dynamic process, with alterations in connexin (Cx) protein expression and post-translational modification as contributing mechanisms. We hypothesized that the endothelial autacoid nitric oxide (NO) would reduce dye coupling in human umbilical vein endothelial cells (HUVECs). In our subsequent experiments, we sought to isolate the specific Cx isoform(s) targeted by NO or NO-activated signaling pathways. Since HUVEC cells variably express three Cx (Cx37, Cx40, and Cx43), this latter aim required the use of transfected HeLa cells (HeLaCx37, HeLaCx43), which do not express Cx proteins in their wild type form. Dye coupling was measured by injecting fluorescent dye (e.g., Alexa Fluor 488) into a single cell and determining the number of stained adjacent cells. Application of the NO donor SNAP (2 microM, 20 min) reduced dye coupling in HUVEC by 30%. In HeLa cells, SNAP did not reduce dye transfer of cells expressing Cx43, but decreased the dye transfer from Cx37-expressing cells to Cx43-expressing cells by 76%. The effect of SNAP on dye coupling was not mediated via cGMP. In contrast to its effect on dye coupling, SNAP had no effect on electrical coupling, measured by a double patch clamp in whole cell mode. Our results demonstrate that NO inhibits the intercellular transfer of small molecules by a specific influence on Cx37, suggesting a potential role of NO in controlling certain aspects of vascular GJIC.     

Connexin 40 expression in bovine and rat retinas

Retinal neurons are extensively coupled through gap junction intercellular channels, but few connexin subtypes have been identified in mammalian retinal neurons. Based on previous findings that retinal gap junctional coupling is modulated by both dopamine and nitric oxide, presumably through connexin phosphorylation, we examined whether the connexin phosphoprotein subtype, connexin 40 (Cx40), was expressed in mammalian retinas. Immunostaining of rat and bovine retinas using Cx40-specific antibodies from two independent sources showed punctate staining between cells in the outer nuclear layer (ONL) and a sublayer of cells within the inner nuclear layer (INL). In addition, sparse punctate staining was detected in the ganglion cell/axon fiber layers (GCL/AFL). No punctate staining was observed in the outer (OS) or inner segment (IS) layers, and rarely in the outer plexiform layer (OPL) or inner plexiform layer (IPL). Double immunostaining of bovine retinas with antibodies to G(o), which stains bipolar cells, and to Cx40, showed little overlap, suggesting these bipolar cells do not express Cx40. Western blot analysis of alkaline-extracted bovine retinal membranes revealed Cx40 immunopositive bands of about 40 kD (monomer) and 80 kD (dimer). In both locations (monomer and dimer), the bands appeared as doublets, and their immunoreactivity was abolished when the antibody was pre-adsorbed with immunogenic Cx40 peptide. The doublet at 40 kD co-migrated with an immunopositive doublet present in heart membranes. Treatment with alkaline phosphatase altered the banding pattern of Cx40. The results suggest that the connexin phosphoprotein subtype, Cx40, is expressed within the neural layers of the mammalian retina.     

Extracellular recording in neuronal networks with substrate integrated microelectrode arrays

A photolithographically produced array of 60 substrate-integrated microelectrodes was used for extracellular recording. Neuronal electrical activity was recorded from chicken retinal ganglion cells with or without stimulation by diffuse light. The retina was removed from chicken embryos of embryonic day 14–18. Only cells recorded from day 18 retina would react to photostimulation, increasing their activity when stimulated, corresponding to the developmental time course of photoreceptor differentiation.     

Stage Specific Glycosylation Pattern for Lactoseries Carbohydrates in the Developing Chick Retina

Based on the idea of differentiation-related changes in the glycosylation pattern of neurons, the expression of two cell surface oligosaccharide epitopes, N-acetyl-lactosamine (NALA), and its sulpho-glucuronyl derivative (HNK-1), was studied, by immunohistochemistry and Western blot experiments, in the developing chick retina beginning on day 2 of incubation (E2) until day 18 post-hatching. NALA was detectable on neuroepithelial cells as soon as the primary optic vesicles formed, and this pattern continued until E3. During subsequent retinal development NALA expression became progressively restricted in concert with the appearance of postmitotic neurons as revealed by neurite outgrowth, and with the formation of synaptic contacts until it disappeared at the end of the incubation period. The pattern of NALA expression was the inverse of HNK-1 which was detected for the first time at E3 on postmitotic ganglion cells accumulating at the vitreal surface. The number of HNK-1+ cells steadily increased until around E10, when the entire neural epithelium was labelled. Synchronously to synaptogenesis, most neurons lost their HNK-1 immunoreactivity. At the time of hatching the adult-like pattern was found, characterised by subpopulations of labelled horizontal, bipolar, amacrine, and ganglion cells. Immunoblot experiments demonstrated transient NALA glycosylation of protein bands, partially identical in their apparent molecular weight to those proteins with HNK-1 glycosylation. The observed temporospatial changes in the glycosylation patterns of distinct proteins during retinal development suggest NALA as a suitable marker for neuronal proliferation, and HNK-1 for differentiation and establishment of final synaptic configuration.     

Gap-junctional coupling measured by flow cytometry

A method is described which reliably quantifies the degree of intercellular communication via gap junctions by combining a dye-loading technique with fluorescence-activated flow cytometry. Our experiments expand former measurements of other groups by analyzing the time- and density-dependent onset of coupling with a fixed ratio of donor to recipient cells. The high sensitivity of this technique provides a better resolution than the microelectrode technique and allows the detection of small changes in gap-junctional coupling by examining a large number of cells in a single experiment. Suspended cells were loaded with the membrane-permeable dye calcein AM, which is intracellularly hydrolyzed by nonspecific esterases, and the resulting polyanionic calcein is thus trapped inside these donor cells. Gap junctions, however, are permeable for this fluorescent dye, as can be observed when suspended donor cells are added to recipient cells (i.e., monolayer cultures) in which case cell-cell contact is established within less than 60 min. In addition, one of these two cell populations can also be stained with a membrane-resident dye (e.g., DiI), which facilitates the identification of different cell populations (donors, recipients, and noncoupled cells) not only by epifluorescence microscopy but also by flow cytometry. Our analyses reveal that junctional coupling depends not only on the connexin type (homo- or heterotypic junction) but also on the origin (species) of the contacting cells (homo- or heterospecific contact). We confirm earlier reports in which homotypic-homospecific coupling was demonstrated with different techniques in connexin-transfected HeLa and RIN cells as well as in BICR/M1R(k) and 3T3/SV40 cells. In contrast to other publications, we show that a significant heterotypic-homospecific coupling between Cx40- and Cx43-HeLa transfectants can be resolved, whereas no coupling was detected for heterotypic-heterospecific contacts between Cx40-HeLa transfectants and the Cx43-expressing cell lines BICR/M1R(k), 3T3/SV40, and RIN.     

Study on RNA synthesis in the retina and retinal pigment epithelium of mice by light microscopic radioautography.

With the aim of determining the distribution of the incorporation of 3H-uridine in both retina and retinal pigment epithelium (RPE), the mouse eyes at embryonic day 9.5 (E 9.5), E 12.5, E 14.5, E 16.5, E 18.5 of gestational ages, and postnatal day 1 (P 1), P 3, P 7, P 14 were analyzed by light microscopic radioautography. Small pieces of the ocular tissues were labelled with 3H-uridine in vitro and light microscopic radioautographs were prepared. The average grain numbers per cell of the respective regions of tissues were calculated. In the retina, the grain numbers increased gradually from E 9.5 to P 1 and reached the maximal value at P 1, and then decreased until P 14. However, the grain numbers were more in the vitreal portion than those in the scleral portion at E 16.5 and then became more in the scleral portion from E 18.5 to P 14. It is considered that the ganglion and bipolar cells finish the RNA synthesis earlier, while the photoreceptor cells do it later during the fetal and postnatal development. In the RPE, the grain numbers gradually increased from E 12.5 to P 7 and then decreased until P 14. Considering the same ages, the grain numbers increased in the following order, anterior, equatorial and posterior regions during embryonic stages, but decreased in the same order after birth. Therefore, it is suggested that the activity of RNA synthesis in PE cells is higher in the posterior region than in the anterior region during embryonic stages. But the activity ascends generally and becomes relatively higher in the anterior region, after birth. Comparing the retina and RPE, it was noted that the grain numbers in the RPE were more important than in the retina and that the maximal value was at P 1 in the retina, while it was at P 7 in the RPE. From these results, it can be concluded that the RNA synthesis ceases earlier in the retina than in the RPE.