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PubMed1994

Developmental changes in the expression and compartmentalization of the glucocorticoid receptor in embryonic retina.

Inducibility by glucocorticoids of the glutamine synthetase gene in chicken embryo retina and the transcriptional activity of the glucocorticoid receptor (GR) greatly increase between embryonic days 6 and 10 (E6, E10), although the level of GR does not markedly change during that time. This apparent discrepancy was investigated by examining the pattern of GR expression in undifferentiated E6 retina and in E10 retina, which consists mostly of differentiated cells. Two GR isoforms, 90 and 95 kDa, were found to be expressed at both of these ages at a similar total level but in different proportions: in E6 retina the level of the 90-kDa isoform was higher, whereas in E10 retina the 95-kDa receptor was higher. However, following treatment of the retinas with cortisol, the 95-kDa isoform became the predominant receptor at both ages. Immunohistochemical analysis revealed that the cellular localization of GR markedly changed in the course of development: in the undifferentiated E6 retina GR was expressed in virtually all cells, whereas in the more differentiated E10 and E12 retina, GR was detected only in Müller glia cells. The latter represent approximately 20% of the cells in this tissue and are the only cells in which glucocorticoid hormone induces the glutamine synthetase gene. We suggest that the compartmentalization of GR in Müller glia is a major aspect of the mechanism that modulates receptor activity during retina development and results in the temporal increase in the inducibility of glutamine synthetase and its specific localization in Müller glia cells.

Animals↗
PubMed2001

Extra-hepatic expression of serum albumin mRNA in mouse retina.

PURPOSE: In some mammals, serum albumin protein exists in the interphotoreceptor space (IPS), the space between photoreceptor cells and the retinal pigment epithelium. Serum albumin is synthesized largely in the liver, though low levels of extra-hepatic expression have been documented in several other tissues, including fetal rat kidney, pancreas, lung, and heart. The purpose of this study was to investigate whether serum albumin protein and mRNA are found in mouse retina. METHODS: Using albumin rabbit antibodies and HRP goat anti-(rabbit IgG), we performed immunoassays on mouse IPS washes to detect the presence of serum albumin protein. Protein extracts from IPS washes were subjected to Affigel Blue chromatography. This resin has an affinity for serum albumin. Reverse transcription-polymerase chain reaction (RT-PCR) of retina total RNA was performed to search for albumin mRNA. Also, real-time reverse transcription polymerase chain reaction (RT-RT-PCR) was employed to look at the levels of expression in different age groups. RESULTS: A constituent of the IPS washes specifically bound and eluted from Affigel Blue column, suggesting that the washes contained serum albumin. SDS PAGE revealed that the size of the constituent was 67 kDa, the size of serum albumin. This 67 kDa band reacted with mouse serum antibody. An RT-PCR amplified fragment of serum albumin mRNA from retina displayed the expected size. The sequence of this fragment is identical to authentic serum albumin cDNA sequence. RPE and choroid were negative for serum albumin mRNA. However, rd1(-)/rd1(-) retina was positive, suggesting that at least some serum albumin is synthesized in the inner layers of the retina. RT-RT-PCR showed that serum albumin mRNA levels in whole retina reached a maximum at about postnatal day 15 and gradually decreased to about one-sixth of maximum at 12 months age. CONCLUSIONS: Serum albumin protein and mRNA were found in mouse IPS and retina, suggesting that the protein is synthesized in the retina. The previously demonstrated ability of serum albumin to bind fatty acids and retinoids and its presence in the mouse IPS suggest a role for serum albumin in transporting retinoids in the retina or IPS, especially at young ages when concentrations appear greatest.

Animals↗
PubMed2001

Development and survival of tyrosine hydroxylase containing neurons in RCS rat retinae.

AIM: Dopamine serves a variety of functions in the retina. Abnormalities of the retinal dopaminergic system have been described in the Royal College of Surgeons (RCS) rat as well as other models of retinal degeneration. Dopamine has been implicated in several retinal dysfunctions of retinitis pigmentosa. Dopaminergic amacrine cells respond to light by increasing their tyrosine hydroxylase (TH) activity and the rate of dopamine turnover. This study has, therefore, examined the ontogenesis of TH containing cells in the RCS rat retina to assess whether progressive photoreceptor degeneration affects the development or survival of TH containing cells in any way. METHODS: TH immunoreactivity in developing dystrophic RCS rat retinae (postnatal day (PN) 0, 3, 6, 14, 18, 26, 32, 56, 85, 91, 12 month and 15 month) and normal retina (PN day 0, 6, 14, 19, 26, 30, 33, 54 and adults) was compared. RESULTS: TH immunoreactivity in dystrophic retina closely resembled that in normal retina. In both groups, very faintly immunoreactive cells were detected in the proximal retina at PN 0. Immunoreactivity increased until PN 14, when faintly immunoreactive interplexiform (IP) fibers and fibers in the outer plexiform layer could be observed. In both groups, the IP connections reached their mature level of development at about PN 30. Thus the developmental expression of TH immunoreactive cells resembled that of non-dystrophic retina in both chronology as well as types of cells. These cells survived even in the advanced stages of degeneration. CONCLUSIONS: The results suggest that the abnormalities in the dopaminergic system of the RCS retinae are not associated with abnormal ontogeny or survival of TH synthesizing cells.

Aging↗
PubMed2002

Upregulation of glucose-dependent insulinotropic polypeptide and its receptor in the retina of streptozotocin-induced diabetic rats.

The pathology of diabetic retinopathy includes dilatation and beading of retinal vessels, and vascular sheathing. To gain a better understanding of the molecular events leading to diabetic retinopathy, we investigated disease-specific gene responses by screening differential expression using cDNA microarray. Male Sprague-Dawley rats were intraperitoneally injected with streptozotocin (STZ, 50 mg/kg) or the control buffer and were maintained for 6 weeks. Total RNA extracted from the retinas of both groups was used for cDNA microarray analysis. Signals from all the spots representing hybridized DNA were quantified and compared between the normal and diabetic rat retinas. Among 1176 genes analyzed, the retinal expression of glucose-dependent insulinotropic polypeptide (GIP) was found to increase in STZ-induced diabetic rats compared to controls. GIP is a secreted protein, known to be released from the small intestine, which potentiates glucose-induced insulin secretion from the pancreas. However, the expression of GIP and its receptor (GIPR) has not been previously noted in the rat retina. To further validate the expression of GIP in the rat retina and to determine its possible role in the development of early diabetic retinopathy, we investigated its expression by RT-PCR, Northern blotting, and immunohistochemistry in normal and diabetic rat retinas. GIP mRNA and protein are not only expressed in the rat retina, but their levels are greater in the diabetic rat as compared to controls. And GIPR expression was also upregulated in the retinas of STZ-induced diabetic rats. We here demonstrate for the first time the expression of GIP and GIPR in the rat retina. And we also revealed some genetic events in the early stage of diabetic retinopathy including the de novo increment of GIP and GIPR expression in the retina.

Animals↗
PubMed1971

Rhodopsin and porphyropsin fields in the adult bullfrog retina.

Though it had been supposed earlier that the bullfrog undergoes a virtually complete metamorphosis of visual systems from vitamin A(2) and porphyropsin in the tadpole to vitamin A(1) and rhodopsin in the adult, the present observations show that the retina of the adult frog may contain as much as 30-40% porphyropsin, all of it segregated in the dorsal zone. The most dorsal quarter of the adult retina may contain 81-89% porphyropsin mixed with a minor amount of rhodopsin; the ventral half contains only rhodopsin. Further, the dorsal zone contains a two to three times higher concentration of visual pigments than the ventral retina. The pigment epithelium underlying the retina contains a corresponding distribution of vitamins A(1) and A(2), predominantly vitamin A(2) in the dorsal pigment epithelium, exclusively vitamin A(1) in the ventral zone. The retina accepts whatever vitamin A the pigment epithelium provides it with, and turns it into the corresponding visual pigment. Thus, a piece of light-adapted dorsal retina laid back on ventral pigment epithelium regenerates rhodopsin, whereas a piece of light-adapted ventral retina laid back on dorsal pigment epithelium regenerates predominantly porphyropsin. Vitamin A(2) must be made from vitamin A(1), by dehydrogenation at the 3,4-bond in the ring. This conversion must occur in the pigment epithelium, presumably through the action of a vitamin A-3,4-dehydrogenase. The essential change at metamorphosis is to make much less of this dehydrogenase, and to sequester it in the dorsal pigment epithelium. Some adult bullfrogs, perhaps characteristically taken in the summer, contain very little porphyropsin-only perhaps 5%-still sequestered in the dorsal retina. The gradient of light over the retinal surface has little if any effect on this distribution. The greater density of visual pigments in the dorsal retina, and perhaps also-although this is less clear-the presence of porphyropsin in this zone, has some ecological importance in increasing the retinal sensitivity to the dimmer and, on occasion, redder light received from below.

Adaptation, Ocular↗
PubMed2005

Localization of endothelin-converting enzyme in bovine optic nerve and retina.

A significant loss and remodeling of the lamina cribrosa tissue leading to the excavation of the optic nerve is seen in glaucoma. Elevated endothelin-1 (ET-1) levels are detected in the aqueous humor of patients of open-angle glaucoma and in the plasma of patients with normal- tension glaucoma. Optic nerve damage, including axonal loss, can be mimicked by ET-1 injection near the optic nerve. ET-1 is produced from its precursor Big ET-1 (38 amino acids) by endothelin-converting enzyme (ECE). Although ET-1 and its receptors have been identified in the retina, little is known of the distribution of ECE at the optic nerve. Presently, ET-1 receptors and Big ET-1 converting activities were characterized in bovine optic nerve and the retina. The ET(B) receptor was detected in both the optic nerve and retina by immunoblotting and cross-linking, using 125I-ET-1. However, the ET(A) receptor was detected only in the retina. Big ET-1 conversion activities were detected in the plasma membrane (PM) of bovine retina, but not in the PM of the optic nerve. The retinal PM Big ET-1 converting activity was inhibited by phosphoramidon, thiorphan, and acidification. Furthermore, ECE cytosolic activities were detected in both the optic nerve and retina. Unlike the PM-ECE, cytosolic Big ET-1 converting activities were activated by acidification (pH 6.4), suggesting the involvement of ECE-2-like activity and/or cathepsin activity. Pepstatin, a potent inhibitor of cathepsins, inhibited the optic nerve (ON) cytosolic conversion of Big ET-1 peptide by 50%, and the combination of pepstatin and phosphoramidon, a potent inhibitor of ECE, inhibited the ON cytosolic activity by 86%. By contrast, the combination of both inhibitors weakly inhibited the cytosolic retinal Big ET-1 converting activity. Western blotting revealed the presence of ECE-1 at the PM of the retina not the ON. ECE-2 and cathpesins B, D, and L were detected only in the cytosol of both the retina and ON. In summary, it appears that ET-1 could be produced in the retina and optic nerve by at least two ECE subtypes and, perhaps, cathepsins. Big ET-1 converting activity may be an important target in preventing ET-1-induced optic nerve pathology.

Animals↗
PubMed1981

Comparison of the binding of [3H]spiperone and [3H]domperidone in homogenates of mammalian retina and caudate nucleus.

The specific binding of [3H]spiperone and [3H]domperidone, as defined by 1 microM-(+)butaclamol, was compared in homogenates of bovine retina and caudate nucleus. Scatchard analyses of saturation data for [3H]spiperone binding yielded dissociation constants (Kd) of 0.35 nM in the retina and 0.64 nM in the caudate nucleus. Comparison of the maximum number of binding sites (Bmax) present in each tissue indicated that the density of sites in bovine caudate nucleus (270 fmol/mg protein) was approximately three times higher than in bovine retina (92 fmol/mg protein). This difference was even more marked in guinea pig tissues, with a ratio of 7:1 between corpus striatum and retina. The pharmacological analysis of [3H]spiperone binding in both the bovine retina and caudate nucleus indicated an interaction with dopaminergic rather than serotonergic sites. However, inhibition curves obtained to dopaminergic agonists in the bovine retina were significantly steeper than those observed in the bovine caudate nucleus, as reflected in the greater Hill coefficients obtained for these agents in the retina. Furthermore, only a small amount of specific [3H]domperidone binding was observed in either the bovine caudate nucleus or the guinea pig striatum, whilst no specific [3H]domperidone binding was detectable in homogenates of either bovine or guinea pig retina. These data suggest that the retina possesses only a small population of dopaminergic D2 sites and that these binding sites may differ from those present in the caudate nucleus.

Animals↗
PubMed1991

Protein kinase C (alpha and beta) immunoreactivity in rabbit and rat retina: effect of phorbol esters and transmitter agonists on immunoreactivity and the translocation of the enzyme from cytosolic to membrane compartments.

Using a monoclonal antibody against protein kinase C (PKC) that recognises the isoenzymes alpha, beta I, and beta II, positive immunoreactivity was observed throughout the cytoplasm of bipolar cells in both rat and rabbit retinas. PKC immunoreactivity was also associated with the outer segment of photoreceptors in the rabbit retina and presumed amacrine cells in the rat retina. The PKC immunoreactivity in the retina was unaffected in content or localisation in rats kept in continuous dark or light conditions over a period of 6 days. The localisation of PKC immunoreactivity in retinas was similar in 6-day-old, 16 day-old, or adult rabbits. However, the content of PKC was lowest at the youngest stage and highest in the adult rabbit retinas. Of the two active phorbol esters studied, only phorbol 12,13-dibutyrate (PDbut) at a concentration of 1 microM caused the PKC immunoreactivity in rabbit retina bipolar cells to be "transported" from the perikarya towards the axonal terminal processes. Biochemical analyses showed that most of the cytosolic PKC was translocated to the membrane compartment following such treatment. The other phorbol ester, phorbol 12-myristate 13-acetate, even at a concentration of 10 microM did not cause a similar transport of PKC immunoreactivity in the bipolar cells, although a partial translocation of the enzyme could be followed biochemically. Both the translocation and transport of PKC by PDbut could be reversed by simply incubating the retinas in physiological solution for 60 min. The "transport" and translocation processes were not obviously affected by the transport inhibitor colchicine or by known PKC inhibitor such as staurosporine, H-7, sphingosine, or polymyxin B. In addition, agonists known to stimulate inositol phosphates in the retina, viz., carbachol, noradrenaline, and quisqualate, or 4-aminopyridine did not cause a translocation or "transport" of PKC as observed for the phorbol esters.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗
PubMed1999

Quantitative estimation of RNA transcripts suggests persistence of Pax-6 expression in the postembryonic chick retina.

Pax-6 is expressed during early embryonic development of the eye. Very little is known about its expression in the functionally mature retina. We have detected Pax-6 transcripts in the ganglion cell- and amacrine cell layers at days 3, 10, 17 and 31 posthatching and in 2- to 3-month-old chick retina by in situ hybridization. These observations were confirmed by a quantitative analysis. Competitive RT-PCR with a homologous internal control revealed a significant reduction (p < 0.001) in the number of Pax-6 transcripts in day 17 retina [(0.39 +/- 0.13) x 10(10)/mg tissue] compared to day 3 retina [(1.65 +/- 0.48) x 10(10)/mg tissue]. Although significantly lower than at day 3, the day 31 retina [(0.7 +/- 0.16) x 10(10)/mg tissue] and retina from 2- to 3-month-old chicks [(0.9 +/- 0.28) x 10(10)/mg tissue] contained an increased number of Pax-6 transcripts in comparison to day 17. On the basis of the amount of RNA, the number of Pax-6 transcripts in the day 3 retina [(0.45 +/- 0.14) x 10(10)/microg RNA] relative to day 17 retina [(0.4 +/- 0.08) x 10(10)/microg RNA] did not change significantly (p = 0.29). However, at day 31 and at 2-3 months of age an increased number of Pax-6 transcripts [(0.65 +/- 0.14) x 10(10)/microg RNA and (0.65 +/- 0.2) x 10(10)/microg RNA, respectively)] were found. In view of the known association of Pax-6 expression with proliferation and emergence of different cell types, these data suggest that cell types in ganglion and inner nuclear cell layers may retain proliferative potential for an extended period in the young adult retina.

Aging↗
PubMed2005

The organization of the inner retina in a pure-rod deep-sea fish.

The pure rod retina of a deep-sea eel species was used as a model system for the study of the differentiation of horizontal, bipolar, amacrine and ganglion cells. We wanted to test the hypothesis that the functional organization of the inner retina is less complex than in species with duplex, rod- and cone-containing retinae. We used immunocytochemistry, backfilling ganglion cells with fluorescent dextranes and microinjection of Lucifer Yellow, to visualize the micromorphology of the various cell types in a confocal microscope. The pure rod retina contains a single type of horizontal cell. The inner plexiform layer is 10-15 microm thick and shows three main sublayers. Bipolar terminals are found in all sublayers, but the majority are found in the inner sublamina b (PKC-immunoreactive cells, that in fish with duplex retinae receive a mixed rod-cone input). The neurochemical diversity of amacrine cells in terms of immunoreactivity does not differ from other teleosts; this similarity includes the pattern of dendritic stratification and ramification as revealed by microinjection. Ten different types of ganglion cells are distinguished based on the sizes of their perikaryon and dendritic field, and the stratification pattern in the inner plexiform layer. This is similar to the situation in catfish with retinae containing a single type of cone in addition to a majority of rods. In this respect, the differences between pure rod retinae and duplex retinae containing a single cone type were less obvious than hypothesized. In the deep-sea eel, the density of dendritic ramification in amacrine and ganglion cells was strongly reduced. This may be functionally related to the fact that vision in the deep sea environment relies exclusively on bioluminescence and is represented by burst-like emissions of point sources. This requires a mode of retinal signal processing that is less complex than in duplex retinae and involves a lower density of dendritic branching and synapses.

Animals↗
PubMed2005

rd1 mouse retina shows imbalance in cellular distribution and levels of TIMP-1/MMP-9, TIMP-2/MMP-2 and sulfated glycosaminoglycans.

BACKGROUND: The rd1 mouse retina displays fast degeneration of photoreceptors resulting in a depletion of almost all rod photoreceptors by postnatal day 21 (PN21). To evaluate the role of proteinases in the pathophysiology of this animal model of retinitis pigmentosa, C3H rd1 and congenic wild-type (wt) mice retinas were analyzed. MATERIAL AND METHODS: The cellular localization and levels of proteins, matrix metalloproteinases (MMPs), their endogenous inhibitors (TIMPs), total sulfated glycosaminoglycans (sGAG) and nature of saccharides in rd1 and wt retinal extracts were compared. RESULTS: MMP-2/TIMP-2 and MMP-9/TIMP-1 were predominantly localized in the interphotoreceptor matrix (IPM) of both genotypes, but MMP-2/TIMP-2 also appeared in the Muller cell fibers of rd1 retina. In rd1 retinal extracts the levels of total proteins were lower and those of active MMP-9, MMP-2, TIMP-1 and total sGAG were higher than those of wt extracts. Despite an increase in TIMP-1, active MMP-9/MMP-2 were disproportionately elevated in rd1 compared to wt retina. With increasing age, MMPs in wt retinas were decreased but were increased in rd1. The sialylation of proteoglycans in PN2 and PN7 rd1 retinas was lower, and galactosylation was higher than that in wt retinas. CONCLUSIONS: MMP-9/MMP-2 and TIMP-1/TIMP-2 are associated with IPM, possibly after secretion by retinal pigmented epithelial cells. In degenerating rd1 retina, MMP-2/TIMP-2 are associated with the Muller cell fibers, which apparently play a central role in modifying the balance between MMPs and TIMPs. Elevated sGAG and proteolysis due to an imbalance in the levels of TIMPs and active MMP-9/MMP-2 in rd1 retina possibly contribute to retinal degeneration in the rd1 mouse.

Animals↗
PubMed2006

A biphasic pattern of gene expression during mouse retina development.

BACKGROUND: Between embryonic day 12 and postnatal day 21, six major neuronal and one glia cell type are generated from multipotential progenitors in a characteristic sequence during mouse retina development. We investigated expression patterns of retina transcripts during the major embryonic and postnatal developmental stages to provide a systematic view of normal mouse retina development, RESULTS: A tissue-specific cDNA microarray was generated using a set of sequence non-redundant EST clones collected from mouse retina. Eleven stages of mouse retina, from embryonic day 12.5 (El2.5) to postnatal day 21 (PN21), were collected for RNA isolation. Non-amplified RNAs were labeled for microarray experiments and three sets of data were analyzed for significance, hierarchical relationships, and functional clustering. Six individual gene expression clusters were identified based on expression patterns of transcripts through retina development. Two developmental phases were clearly divided with postnatal day 5 (PN5) as a separate cluster. Among 4,180 transcripts that changed significantly during development, approximately 2/3 of the genes were expressed at high levels up until PN5 and then declined whereas the other 1/3 of the genes increased expression from PN5 and remained at the higher levels until at least PN21. Less than 1% of the genes observed showed a peak of expression between the two phases. Among the later increased population, only about 40% genes are correlated with rod photoreceptors, indicating that multiple cell types contributed to gene expression in this phase. Within the same functional classes, however, different gene populations were expressed in distinct developmental phases. A correlation coefficient analysis of gene expression during retina development between previous SAGE studies and this study was also carried out. CONCLUSION: This study provides a complementary genome-wide view of common gene dynamics and a broad molecular classification of mouse retina development. Different genes in the same functional clusters are expressed in the different developmental stages, suggesting that cells might change gene expression profiles from differentiation to maturation stages. We propose that large-scale changes in gene regulation during development are necessary for the final maturation and function of the retina.

Animals↗
PubMed1990

Docosahexaenoic acid is the preferred dietary n-3 fatty acid for the development of the brain and retina.

The metabolism of individual dietary n-3 fatty acids was studied in n-3 fatty acid-deficient newly hatched chicks. Laying hens were fed the n-6 fatty acid, ethyl linoleate, as the only source of polyunsaturated fat. Chicks were then fed the n-3-deficient hens' diet, or one of three other diets supplemented with the ethyl ester of 18:3 n-3, 20:5 n-3 [eicosapentaenoic acid (EPA)], or 22:6 n-3 [docosahexaenoic acid (DHA)] at 0.44% of calories. At the end of 0, 1, 2, and 3 wk, the fatty acid composition of the brain, retina, liver, and serum was determined. Dietary EPA and DHA were equally effective at raising levels of DHA in the brain and retina. Dietary 18:3 was relatively ineffective in restoring brain and retina DHA. In the n-3-deficient chicks fed EPA or DHA, levels of DHA recovered to control values in both the brain and retina by 3 wk. Very little EPA accumulated in the brain or retina of chicks fed EPA. Hepatic synthesis of DHA from EPA appeared low, suggesting that the brain and retina synthesized the DHA that accumulated rapidly in these tissues after the feeding of EPA. The delta-4-desaturase enzyme was apparently very active, then, in the brain and retina. Retroconversion of dietary 22:6 to 22:5 and 20:5 was evident in the serum, liver, and retina but not in the brain. Thus, it was possible to study the relative metabolism and especially the interconversion of n-3 fatty acids in a environment uncomplicated by existing stores of these essential fatty acids. This study would suggest that 18:3 as the sole source of n-3 fatty acids in the diets of animals, including the human infant, may not be adequate for the biochemical development of the brain and retina and that dietary DHA is the preferred fatty acid of the n-3 series.

Animals↗
PubMed2004

Neural retina limits the nonviral gene transfer to retinal pigment epithelium in an in vitro bovine eye model.

We investigated the permeation of liposomal and polymeric gene delivery systems through neural retina into retinal pigment epithelium (RPE) and determined the roles of various factors in permeation and subsequent uptake of the delivery systems by RPE. Anterior parts and vitreous of fresh bovine eyes were removed. Retina was left intact or peeled away. Complexes of ethidium monoazide (EMA)-labeled plasmid DNA and cationic carriers (polyethyleneimine, poly-L-lysine, DOTAP liposomes) were pipetted on the retina or RPE. Two hours later the neural retina was removed, if present, and the RPE cells were detached. Contaminants were removed by sucrose centrifugation, and the RPE cells were analyzed for DNA uptake by flow cytometry. Cellular uptake of FITC-dextrans (molecular weight [mw] 20,000, 500,000 and 2,000,000), FITC-poly-L-lysine (mw 20,000), FITC-labeled oligonucleotide (15-mer), and naked EMA-labeled plasmid DNA was determined after pipetting the solutions on the RPE or neural retina. Location of the fluorescent materials in the retina was visualized with fluorescence microscopy. Neural retina decreased the cellular uptake of DNA complexes by an order of magnitude, the uptake of FITC-dextrans slightly, whereas delivery of polycationic FITC-poly-L-lysine to RPE was almost completely inhibited. Neural retina decreased the cellular uptake of FITC-oligonucleotides, while the uptake of uncomplexed plasmid was always negligible. Conclusions from FACS and fluorescence microscopy were similar: delivery of polymeric and liposomal DNA complexes into RPE are limited by the neural retina. This is due to the size and positive charge of the complexes.

Animals↗
PubMed2005

Retinal neurons regulate proliferation of postnatal progenitors and Müller glia in the rat retina via TGF beta signaling.

The number of proliferating cells in the rodent retina declines dramatically after birth. To determine if extrinsic factors in the retinal micro-environment are responsible for this decline in proliferation, we established cultures of retinal progenitors or Muller glia, and added dissociated retinal neurons from older retinas. The older cells inhibited proliferation of progenitor cells and Muller glia. When these experiments were performed in the presence of TGF(beta)RII-Fc fusion protein, an inhibitor of TGF(beta) signaling, proliferation was restored. This suggests a retina-derived TGF(beta) signal is responsible for the developmental decline in retinal proliferation. TGFbeta receptors I and II are expressed in the retina and are located in nestin-positive progenitors early in development and glast-positive Muller glia later in development. RT-PCR and immunofluorescence data show TGF(beta)2 is the most highly expressed TGF(beta)ligand in the postnatal retina, and it is expressed by inner retinal neurons. Addition of either TGF(beta)1 or TGF(beta)2 to postnatal day 4 retinas significantly inhibited progenitor proliferation, while treatment of explanted postnatal day 6 retinas with TGF(beta) signaling inhibitors resulted in increased proliferation. Last, we tested the effects of TGF(beta) in vivo by injections of TGF(beta) signaling inhibitors: when TGF(beta) signaling is inhibited at postnatal day 5.5, proliferation is increased in the central retina; and when co-injected with EGF at postnatal day 10, TGF(beta)inhibitors stimulate Muller glial proliferation. In sum, these results show that retinal neurons produce a cytostatic TGF(beta) signal that maintains mitotic quiescence in the postnatal rat retina.

Animals↗
PubMed1993

Developmental changes in carbachol-stimulated inositolphosphate release in pigmented rat retina.

Carbachol-stimulated release of inositolphosphates (IP) was studied in the whole retina from Long-Evans rats of different ages (day 5, 10, 15, 20, adult) following in vitro incorporation of [3H]myo-inositol. Unlike the albino rat retina, the pigmented retina was highly light-sensitive, making it necessary to dark adapt the animals and perform retinal dissections under low illumination to prevent light-induced IP release. Retinae from postnatal day 10 rats showed the highest amount of carbachol-stimulated IP released. This response to carbachol decreased with age until postnatal day 20 when it reached adult levels. The pigmented rat retina showed a sharp fall in the degree of carbachol (1 mM)-stimulated IP released at the time of eye-opening (450% above basal in retinae from 10 day old animals, as compared to 230% above basal in 15 day old retinae). Basal release of IP was not altered in the retina during development. Muscarinic cholinergic receptor density was, however, found to increase 5 fold with age, reaching adult levels by PND 20. Retinal weight and protein per retina also increased (four fold) from day 5 to adult; however, the in vitro incorporation of [3H]myo-inositol into phosphoinositides (calculated as per mg protein) did not change during development. Thus, in animals prior to eye opening, a much higher proportion of phosphoinositides appears to be hydrolyzed upon muscarinic receptor stimulation. During retinal development a change in sensitivity to the agonist-sensitive pool(s) of phosphoinositides may occur and/or there may be alterations in the efficacy of receptor coupling to the second messenger system resulting in the disassociation observed between the drastic increase in receptor number and the apparent decrease in receptor-stimulated release of IP.

Aging↗
PubMed1999

Expression of proteoglycan decorin in neural retina.

PURPOSE: To identify the expression of chondroitin/dermatan sulfate proteoglycan decorin in retina and to elucidate its changes during development and ischemia-reperfusion. METHODS: Expression of decorin in rat retina was investigated by reverse transcription-polymerase chain reaction (RT-PCR) and immunohistochemistry. Distributional changes during development and transient ischemia in model eyes also were investigated by immunohistochemical experiments. RESULTS: Gene expression of decorin core protein was identified in rat retina by RT-PCR. Decorin immunoreactivities were shown throughout the retina, especially in the ganglion cell layer. In developing rat retinas, at embryonic stages (embryonic day 16), decorin was distributed uniformly throughout the retina. As retina matured, the intensity of decorin immunostaining in retinal inner layers and retinal pigment epithelium increased. Furthermore, in experimental transient retinal ischemia, after transient downregulation of the decorin core protein gene between 24 and 48 hours after the ischemia, recovered (or increased) expression was shown by semiquantitative RT-PCR experiments. Immunohistochemical studies revealed strong decorin immunoreactivities in the damaged inner layers 1 week later. CONCLUSIONS: The expression of decorin was identified in adult and developing rat retina. The distributional changes of decorin during the retinal development suggest that this proteoglycan may play a role in the differentiation of retinal ganglion cells. Moreover, in rat ischemia-reperfusion models, the alterations in gene expression and immunohistochemical localization showed the contribution of this proteoglycan to the damage and repair processes in diseased retina.

Animals↗
PubMed1999

Immunocytochemical study of the distribution of a 16-kDa galectin in the chicken retina.

PURPOSE: To compare the distribution of a developmentally regulated 16-kDa galectin in the chicken retina at two different developmental stages: embryonic day 13 (ED13) and postnatal day 10 (PD10) retinas, by immunocytochemical analysis using light and transmission electron microscopy. METHODS: Semi-thin and thin sections from ED13 and PD10 retinas were incubated with the IgG fraction purified from a rabbit antiserum raised against the 16-kDa chicken galectin. After incubation with colloidal gold particle-labeled goat anti-rabbit IgGs, tissue sections were analyzed by light and transmission electron microscopy. To improve the observation by light microscopy, semi-thin immunostained sections were intensified by silver enhancement. RESULTS: In ED13 retinas a specific galectin labeling was detected in the region corresponding to the outer limiting membrane by light microscopy. This labeling seemed to be associated with the apical villi of Muller glial cells and their specialized junctions, as judged by transmission electron microscopy. In PD10 retinas, the more relevant finding revealed by light microscopy was the detection of a widespread immunostaining at the level of all retinal layers. The ultrastructural analysis indicated that the galectin labeling was detected at the cytoplasmic and nuclear compartments of Muller cells throughout the different retinal layers. Moreover, the labeling was detected in the inner limiting membrane in structures that resemble the end feet of Muller cells. The apical villi, and the specialized junctions of these glial cells, appeared more strongly stained in PD10 retinas than in ED13 retinas. Finally, highly intense labeling in a group of mitochondria localized in the inner segments of cone cells was observed. CONCLUSIONS: The present study clearly supports the idea that the subcellular distribution of the 16-kDa galectin changes during the development of the chicken retina. Morphologic changes associated with developmentally regulated expression and subcellular compartmentalization of the retinal galectin suggest that this lectin may be involved in the modulation of several processes in the visual system. Its presence in the apical villi of Miller cells may be related by modulatory functions between retina and pigment epithelium, but its presence in the cytoplasm and nucleus of these glial cells suggests a potential immunomodulatory role and its involvement in different metabolic processes between Muller and the other retinal cells. Finally, although the presence of galectins inside mitochondria has not been described before, this localization gives rise to the idea that this lectin may be involved in the modulation of mitochondrial processes.

Animals↗