Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Rod Cell Outer Segment”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 523 records · Page 29Linked to original sources
PubMed1989

Evidence that microtubules do not mediate opsin vesicle transport in photoreceptors.

The organization of the rod photoreceptor cytoskeleton suggests that microtubules (MTs) and F actin are important in outer segment (OS) membrane renewal. We studied the role of the cytoskeleton in this process by first quantifying OS membrane assembly in rods from explanted Xenopus eyecups with a video assay for disc morphogenesis and then determining if the rate of assembly was reduced after drug disassembly of either MTs or F actin. Membrane assembly was quantified by continuously labeling newly forming rod OS membranes with Lucifer Yellow VS (LY) and following the tagged membranes' distal displacement along the OS. LY band displacement displayed a linear increase over 16 h in culture. These cells possessed a longitudinally oriented network of ellipsoid MTs between the sites of OS protein synthesis and OS membrane assembly. Incubation of eyecups in nocodazole, colchicine, vinblastine, or podophyllotoxin disassembled the ellipsoid MTs. Despite their absence, photoreceptors maintained a normal rate of OS assembly. In contrast, photoreceptors displayed a reduced distal displacement of LY-labeled membranes in eyecups treated with cytochalasin D, showing that our technique can detect drug-induced changes in basal rod outer segment assembly. The reduction noted in the cytochalasin-treated cells was due to the abnormal lateral displacement of newly added OS disc membranes that occurs with this drug (Williams, D. S., K. A. Linberg, D. K. Vaughan, R. N. Fariss, and S. K. Fisher. 1988. J. Comp. Neurol. 272:161-176). Together, our results indicate that the vectorial transport of OS membrane constituents through the ellipsoid and their assembly into OS disc membranes are not dependent on elliposid MT integrity.

Animals↗
PubMed1990

Metabolic labeling of rod outer segment phospholipids in miniature poodles with progressive rod-cone degeneration (prcd).

The recessive genetic defect in miniature poodles which results in progressive rod-cone degeneration (prcd) has been investigated in an attempt to determine the biochemical abnormality involved. In the present study, the rod outer segments of young prcd affected miniature poodles and normal dogs have been compared with respect to the incorporation of intravitreally injected [3H]palmitic acid. [14C]linolenic acid, and [14C]docosahexaenoic acid into neutral lipids and phospholipids as well as [3H]palmitate and [14C]leucine into rhodopsin. In addition, 3 mm trephined punches of retinas were incubated with [3H]palmitic acid, [3H]arachidonic acid, [14C]linolenic acid, [3H]serine, [14C]glycerol and [14C]leucine. No difference in incorporation of labeled precursors into lipids or rhodospin was noted between prcd affected and normal retinas. Phosphatidyl choline appeared to function as a carrier of fatty acids to the rod outer segment where they were redistributed to other phospholipids. An interesting lack of conversion of the essential fatty acid linolenic acid to docosahexaenoic acid was noted in both normal and affected retinas. This conversion involves elongation and desaturation of linolenic acid and may take place primarily in extraretinal tissues such as the liver. This finding, in conjunction with a parallel study of plasma fatty acids which has shown significantly lower levels of docosahexaenoic acid in prcd affected poodles, points to a possible systemic defect in the metabolism or transport of docosahexaenoic acid, a fatty acid uniquely enriched in the photoreceptor outer segments.

Animals↗
PubMed1991

The NH2-terminal alpha subunit attenuator domain confers regulation of G protein activation by beta gamma complexes.

Gs and Gi, respectively, activate and inhibit the enzyme adenylyl cyclase. Regulation of adenylyl cyclase by the heterotrimeric Gs and Gi proteins requires the dissociation of GDP and binding of GTP to the alpha s or alpha i subunit. The beta gamma subunit complex of Gs and Gi functions, in part, to inhibit GDP dissociation and alpha subunit activation by GTP. Multiple beta and gamma polypeptides are expressed in different cell types, but the functional significance for this heterogeneity is unclear. The beta gamma complex from retinal rod outer segments (beta gamma t) has been shown to discriminate between alpha i and alpha s subunits (Helman et al: Eur J Biochem 169:431-439, 1987). beta gamma t efficiently interacts with alpha i-like G protein subunits, but poorly recognizes the alpha s subunit. beta gamma t was, therefore, used to define regions of the alpha i subunit polypeptide that conferred selective regulation compared to the alpha s polypeptide. A series of alpha subunit chimeras having NH2-terminal alpha i and COOH-terminal alpha s sequences were characterized for their regulation by beta gamma t, measured by the kinetics of GTP gamma S activation of adenylyl cyclase. A 122 amino acid NH2-terminal region of the alpha i polypeptide encoded within an alpha i/alpha s chimera was sufficient for beta gamma t to discriminate the chimera from alpha s. A shorter 54 amino acid alpha i sequence substituted for the corresponding NH2-terminal region of alpha s was insufficient to support the alpha i-like interaction with beta gamma t. The findings are consistent with our previous observation (Osawa et al: Cell 63:697-706, 1990) that a region in the NH2-terminal moiety functions as an attenuator domain controlling GDP dissociation and GTP activation of the alpha subunit polypeptide and that the attenuator domain is involved in functional recognition and regulation by beta gamma complexes.

Adenylyl Cyclases↗
PubMed1985

Ultrastructural localization of S-antigen in retinal structures.

The localization of S-antigen, the soluble, uveitogenic, 51 kDa protein of the retina has been studied. Highly specific rabbit anti-bovine S-antigen antiserum reacted predominantly with the outer segment layer of the photoreceptor cells of rat retina in the indirect immunofluorescence technique, provided that the tissue was fixed by perfusion of the light adapted animal. More detailed information was obtained by immuno-electron microscopy using the same fixation technique and Protein A-coated gold particles for labeling. S-antigen was found to be predominantly bound to the rod outer segment plasma membranes and to the discs. The possible implication of this localization is discussed in view of the function of S-antigen.

Animals↗
PubMed1990

Retinal detachment prevents normal assembly of disk membranes in vitro.

The effects of retinal detachment upon disk membrane assembly in rod outer segments were assessed in Xenopus laevis retinas that had been maintained in eyecup cultures for up to 4 days. In these cultures, assembly of disk membranes occurred at a normal rate in regions of the retina that remained attached to the retinal pigment epithelium. In regions of the retina that were detached from the pigment epithelium, the assembly of new disk membranes either was abnormal or was inhibited. This result cannot be attributed to reduced access of cells in the detached retina to oxygen and metabolites. The experiments described here suggest that the apposition of the retina with the pigment epithelium is a necessary condition for normal disk membrane assembly in Xenopus retinas. This effect may be mediated by contact between the rod outer segments and the pigment epithelium, or by trophic factors in the subretinal space.

Animals↗
PubMed2002

Tamoxifen induces changes in the lipid composition of the retinal pigment epithelium cell line D407.

Tamoxifen, the antioestrogenic drug prescribed for long-term, low-dose therapy of breast cancer, induces retinopathy. This study evaluates the effects of tamoxifen on the human retinal pigment epithelial cell line D407, attempting to identify the underlying mechanisms on tamoxifen-induced retinopathy and the involvement of cellular membranes in the cytotoxic action mechanism. We demonstrate that the tamoxifen-induced decrease in the cell growth of the D407 cell line results from pyknosis and cell cycle arrest rather than from necrosis. Furthermore, D407 cells influence the lipid composition of both plasma membrane and intracellular membranes in response to tamoxifen. Tamoxifen increases the physical order of the lipid bilayer. We observed a compensatory decrease in the cholesterol content of the plasma membrane which results in an increase of the plasma membrane fluidity. In intracellular membranes the phosphatidylcholine content is reduced to 50% of the controls. This reduction may be related to the formation of a second messenger via phospholipase pathway and sustained activation of protein kinase C. Since increased plasma membrane fluidity as well as sustained activation of protein kinase C influence the rod outer segments binding and/or ingestion by retinal pigment epithelial cells, our results suggest that membrane-mediated pathways contribute to the tamoxifen-induced retinopathy.

Antineoplastic Agents, Hormonal↗
PubMed2006

Detection and localization of a putative cyclic-GMP-activated channel protein in the protozoan ciliate Stentor coeruleus.

Immunoblotting and immunocytochemical assays were employed to identify and localize a channel protein activated by cyclic GMP (cGMP) in the protozoan ciliate Stentor coeruleus. Analysis of whole-cell homogenate with antibodies raised against the alpha-subunit of the cGMP-activated channel protein from bovine rod outer segments and against cGMP revealed four major protein bands with molecular masses of 40 kDa, 63 kDa, and over 120 kDa, which bound cGMP. However, only a cGMP-binding protein of 63 kDa, corresponding to the alpha-subunit of the cGMP-activated ion channel protein from bovine rod outer segments, was found in the ciliate cortex fraction. The functional cGMP-activated channel protein was also shown to be present in the cortex fraction of S. coeruleus by patch-clamp measurements of artificial liposomes. Incorporation of the cortex fraction into liposomes resulted in the appearance of ion channel activity related to cGMP. The reconstituted protein channels were strongly inhibited by l-cis-diltiazem, a known potent blocker of many types of cyclic-nucleotide-activated channels. The results presented here are the first demonstration of the existence and localization of a putative cGMP-activated channel protein in the ciliate S. coeruleus. Cyclic-nucleotide-activated channel proteins are nonspecific cation channels which mediate the receptor potentials in photoreceptor cells and in cells of the olfactory epithelium. On the basis of these data, we suggest that the 63 kDa protein identified in Stentor coeruleus is also a cGMP-activated ion channel and that it may be involved as an effector in the photosensory transduction pathway leading to the motile photophobic response in this ciliate protist.

Animals↗
PubMed2000

Retinal vascular changes induced by the oxidative stress of alpha-tocopherol deficiency contrasted with diabetic microangiopathy.

It has been proposed that oxidative tissue damage is involved in the development of diabetic angiopathies. To evaluate this hypothesis, experiments were conducted to identify the retinal vessel changes induced by the oxidative stress related to alpha-tocopherol deficiency and examine possible similarities with the lesions characteristic of diabetic retinopathy. Twenty-one-day-old male Fisher 344 albino rats were divided randomly to receive a basal, chemically defined diet either with (adequate group) or without (deficient group) alpha-tocopherol. After 6 and 8 months, some rats (n = 3 per group) were killed and the eyes removed. In order to evaluate cell integrity and localization of lipofuscin-specific autofluorescence by light and fluorescence microscopy, some of the retinas were prepared for cryostat-sections while others were digested by elastase to isolate intact retinal vasculatures. After 8 and 14 months, the central retina of one eye per rat (n = 6 to 8 per group) was examined by electron microscopy for retinal capillary basement membrane (RCBM) thickening and other ultrastructural changes. At 6 and 8 months, the deficient rats exhibited extensive shortening and disarray of rod outer segments (ROS), marked loss of photoreceptor cells, and pronounced increases in the numbers of granules with lipofuscin-specific autofluorescence in the retinal pigment epithelium (RPE) and retinal vessels. At 14 months, the ultrastructure revealed that the damage to ROS involved disruption of membranes and that the capillary lipofuscin was contained mainly within the endothelial cells. Membrane remnants were found in the lipofuscin granules of both the RPE and retinal vessels. In addition, there was an increase in RCBM thickness (98.7 +/- 2.6 nm vs. 86.9 +/- 2.9 nm). RCBM thickening was the only finding common with diabetic retinopathy, and the thickening was 13.6%, significantly less than that reported in diabetic rat models with 8 and 14 months durations (34% and 53.1%, respectively). Capillary lipofuscin accumulation, which was prominent in the deficient rats, is not notable in diabetes. Both the moderate RCBM thickening and marked lipofuscin accumulations seen in alpha-tocopherol-deficient rats were similar to changes occurring in the aging process, though more pronounced. The spectrum of microangiopathies characteristic of diabetic retinopathy did not develop in alpha-tocopherol-deficient rats. These findings suggest that oxidative damage, though probably involved, is unlikely to play a predominant role in the development of diabetic retinal microangiopathies.

Animals↗
PubMed1994

Superoxide inhibits proliferation and phagocytic internalization of photoreceptor outer segments by bovine retinal pigment epithelium in vitro.

Experiments were performed to investigate the effect of free radical damage on two aspects of retinal pigmented epithelium (RPE) metabolism, namely, proliferation and phagocytosis. Bovine RPE cells were maintained in monolayer cultures, either as passaged (for proliferation and lysosomal activity assays) or primary cultures (for phagocytosis measurements). Free radicals (superoxide anions) were generated by a xanthine oxidase (XO)-hypoxanthine (HX) reaction. Total phagocytosis (binding plus ingestion of rod outer segments (ROS)) was quantitated by radioimmunoassay using a specific anti-opsin antibody and iodinated secondary antibody. In some cases, agents with known or possible protective influences against oxidative damage, i.e., superoxide dismutase (SOD), vitamin E, and basic fibroblast growth factor (bFGF), were tested for their activity in this model system. RPE cell proliferation was inhibited in a HX-XO dose-dependent manner, in the absence of cell toxicity. Modifications of cell morphology were also noticed. Either simultaneous exposure of RPE cells to ROS membranes and HX-XO or pretreatment of ROS membranes with HX-XO prior to their addition to RPE monolayers led to a statistically significant 20-30% decrease in phagocytosis relative to control values. This decrease was essentially observed in the binding phase of phagocytosis, indicating damage to ROS surface molecules as the primary event. Addition of SOD or vitamin E prevented this loss of phagocytic activity, whereas bFGF had no effect. Superoxide radicals did not, however, affect phagocytosis when RPE cells were exposed to them alone, prior to incubation with ROS; nor did they alter a later stage in the phagocytic process, acid phosphatase activity. This tissue culture model represents a convenient system for analyzing free radical damage in different aspects of RPE-photoreceptor behavior and may be useful in studying this phenomenon in several retinal disorders.

Acid Phosphatase↗
PubMed1994

The human photoreceptor membrane guanylyl cyclase, RetGC, is present in outer segments and is regulated by calcium and a soluble activator.

A human photoreceptor membrane guanylyl cyclase, RetGC, was recently cloned and expressed, but its localization and manner of regulation were not defined. We report here that RetGC is detected primarily in outer segments of human photoreceptor cells. Recombinant RetGC can be stimulated by a soluble retinal-specific factor. Ca2+ interferes with stimulation of RetGC by this factor with a cooperativity coefficient of 1.7 and EC50 near 200 nM. The Ca2+ sensitivities of recombinant RetGC and of guanylyl cyclase activity from rod outer segment membranes are very similar. Our findings indicate that RetGC is a photoreceptor-specific guanylyl cyclase which is stimulated by a retinal-specific activator and inhibited by physiologically relevant concentrations of free Ca2+. The Ca2+ sensitivity of RetGC may be responsible for some of the previously reported effects of Ca2+ on light adaptation and recovery of the dark state.

Amino Acid Sequence↗
PubMed1993

Structural and biochemical identity of retinal rod outer segment membrane guanylate cyclase.

Recent molecular cloning reports show that there are at least three membrane guanylate cyclases in vertebrate retina: (1) atrial natriuretic factor receptor guanylate cyclase (ANF-RGC), (2) C-type natriuretic peptide receptor guanylate cyclase (CNP-RGC), and (3) "retinal guanylate cyclase" (RetGC). The specific cellular localization of the first two cyclases is unknown, but RetGC is apparently localized in photoreceptor cells, suggesting that it participates in visual transduction. With the overall objective of identifying the guanylate cyclase that is linked to phototransduction, we compared the structural and regulatory properties of the biochemically characterized 112 kDa bovine rod outer segment membrane guanylate cyclase (ROS-GC) with those of RetGC, ANF-RGC and CNP-RGC. The N-terminal and two internal peptide sequences of purified ROS-GC had about 90% similarity with the corresponding sequences of the RetGC; the sequence identity with natriuretic peptide receptor cyclases was about 30%. A 19 amino acid long sequence from a tryptic peptide of ROS-GC had no corresponding sequence in the other three cyclases. ROS-GC was inhibited by ATP but ANF-RGC and CNP-RGC were activated by ATP in the presence of the respective peptide hormones. These results suggest that ROS-GC represents a new subtype of the membrane guanylate cyclase family that is structurally and biochemically distinct from the other retinal cyclases.

Adenosine Triphosphate↗
PubMed1986

G-proteins of fat-cells. Role in hormonal regulation of intracellular inositol 1,4,5-trisphosphate.

Pertussis toxin abolishes hormonal inhibition of adenylate cyclase, hormonal stimulation of inositol 1,4,5-trisphosphate accumulation in rat fat-cells, and catalyses the ADP-ribosylation of two peptides, of Mr 39,000 and 41,000 [Malbon, Rapiejko & Mangano (1985) J. Biol. Chem. 260, 2558-2564]. The 41,000-Mr peptide is the alpha-subunit of the G-protein, referred to as Gi, that is believed to mediate inhibitory control of adenylate cyclase by hormones. The nature of the 39,000-Mr substrate for pertussis toxin was investigated. The fat-cell 39,000-Mr peptide was compared structurally and immunologically with the alpha-subunits of two other G-proteins, Gt isolated from the rod outer segments of bovine retina and Go isolated from bovine brain. After radiolabelling in the presence of pertussis toxin and [32P]NAD+, the electrophoretic mobilities of the fat-cell 39,000-Mr peptide and the alpha-subunits of Go and Gt were nearly identical. Partial proteolysis of these ADP-ribosylated proteins generates peptide patterns that suggest the existence of a high degree of homology between the fat-cell 39,000-Mr peptide and the alpha-subunit of Go. Antisera raised against purified G-proteins and their subunits were used to probe immunoblots of purified Gt, Gi, Go, and fat-cell membrane proteins. Although recognizing the 36,000-Mr beta-subunit band of Gt, Gi, Go and a 36,000-Mr fat-cell peptide, antisera raised against Gt failed to recognize either the 39,000- or the 41,000-Mr peptides of fat-cells or the alpha-subunits of Go and Gi. Antisera raised against the alpha-subunit of Go, in contrast, recognized the 39,000-Mr peptide of rat fat-cells, but not the alpha-subunit of either Gi or Gt. These data establish the identity of Go, in addition to Gi, in fat-cell membranes and suggest the possibility that either Go or Gi alone, or both, may mediate hormonal regulation of adenylate cyclase and phospholipase C.

Adenylate Cyclase Toxin↗
PubMed1989

Light exposure stimulates arachidonic acid metabolism in intact rat retina and isolated rod outer segments.

This paper presents evidence that short-term exposure to light increases synthesis of hydroxyeicosatetraenoic acid (HETE) and prostaglandin D2 (PGD2) and stimulates the uptake and metabolism of 20:4 in phospholipids and triacylglycerols in rat retina. There was a time-dependent increase in incorporation of 1-14C-20:4 into glycerolipids in both dark-adapted and light-exposed groups. Exposure to light for 15 or 30 min enhanced the acylation of 20:4 into phosphatidylcholine, phosphatidylinositol, phosphatidylethanolamine and triacylglycerols. In the light-exposed groups there was a large increase in the conversion of 20:4 to leukotriene B4, two diHETEs, 5-HETE, 15-HETE, and PGD2. The stimulation of HETE synthesis by light could be due to early stages of light-induced lipid peroxidation in visual cells. To examine this, we studied peroxidation of 20:4 in isolated rod outer segments (ROS). There was more oxidation of 20:4 in light-exposed ROS, as compared to ROS incubated in the dark. Vitamin E and nordihydroguaiaretic acid inhibited the light-induced formation of some of these products. The data indicate that photo-oxidation of 20:4 in ROS is accompanied by enzymatic oxygenation that is stimulated by light. Increased production of HETEs an PGD2 may be a consequence of the light-induced stimulation of the metabolism of 20:4 in membrane phospholipids in the retina.

Animals↗
PubMed1997

Retinopathy induced in mice by targeted disruption of the rhodopsin gene.

Retinitis pigmentosa (RP) represents the most common mendelian degenerative retinopathy of man, involving death of rod photoreceptors, cone cell degeneration, retinal vessel attenuation and pigmentary deposits. The patient experiences night blindness, usually followed by progressive loss of visual field. Genetic linkage between an autosomal dominant RP locus and rhodopsin, the photoreactive pigment of the rod cells, led to the identification of mutations within the rhodopsin gene in both dominant and recessive forms of RP. To better understand the functional and structural role of rhodopsin in the normal retina and in the pathogenesis of retinal disease, we generated mice carrying a targeted disruption of the rhodopsin gene. Rho-/- mice do not elaborate rod outer segments, losing their photoreceptors over 3 months. There is no rod ERG response in 8-week-old animals. Rho+/- animals retain the majority of their photoreceptors although the inner and outer segments of these cells display some structural disorganization, the outer segments becoming shorter in older mice. These animals should provide a useful genetic background on which to express other mutant opsin transgenes, as well as a model to assess the therapeutic potential of re-introducing functional rhodopsin genes into degenerating retinal tissues.

Age Factors↗
PubMed1993

A method for the isolation of retinal pigment epithelial cells from adult rats.

PURPOSE: Isolating retinal pigment epithelium (RPE) cells from adult rat was the goal of this study. METHOD: A modification of the procedure of Mayerson et al was the method employed. After the cornea and lens were removed, the eyecup was treated with hyaluronidase plus collagenase. The neural retina was carefully removed with little RPE attached. The eyecup, free of retina, was treated with the same enzyme cocktail that loosens any attached rod outer segments as well as the attachment of the RPE to Bruch's membrane. RPE cells were isolated in calcium-free medium as sheets of cells. RESULTS: Light and electron microscopy revealed good cell morphology. Purity of the RPE preparation was established by microscopy, polyacrylamide gel electrophoresis, and lipid analysis. The viability of the isolated RPE was about 74% by trypan blue exclusion test. CONCLUSIONS: This technique enables RPE to be isolated from adult rat in quantities sufficient for biochemical analysis.

Animals↗
PubMed1989

Cholesterol heterogeneity in bovine rod outer segment disk membranes.

Rod outer segment disk membranes have been used to study visual transduction events. Numerous studies have also focused on protein-lipid interactions in these membranes. The possible heterogeneity of the disk membrane composition has not been addressed in such studies. Freeze fracture studies (Andrews, L. D., and Cohn, A. I. (1979) J. Cell Biol. 81, 215-220; Caldwell, R., and McLaughlin, B. (1985) J. Comp. Neurol. 236, 523-537) suggest a difference in cholesterol content between newly formed and old disks. This potential heterogeneity in disk membrane composition was investigated using digitonin. Osmotically intact bovine rod outer segment disk membranes prepared by Ficoll flotation were separated based on the cholesterol content of the disks. The addition of digitonin to disk membrane suspensions in a one-to-one molar ratio with respect to cholesterol produced an increase in the density of the membranes in proportion to the amount of cholesterol present. The digitonin-treated disks were separated into subpopulations using a sucrose density gradient. Disks were shown to vary in cholesterol to phospholipid ratio from 0.30 to 0.05. The ratio of phospholipid to protein remained constant in all disk subpopulations at approximately 65 phospholipids per protein. No significant change in the fatty acid composition of the disks was observed as a function of change in cholesterol content. This work demonstrates compositional heterogeneity in disk membranes which may ultimately affect function.

Animals↗
PubMed1992

Ascorbate treatment prevents accumulation of phagosomes in RPE in light damage.

In dark-reared albino rats, exposure to 2 or 3 hr of intense light interrupted by 2 hr dark periods resulted in extensive degeneration of photoreceptor cells and degeneration of the retinal pigment epithelium (RPE). Ascorbate (ie, vitamin C) administration prior to light exposure protected photoreceptors and the RPE from light damage. In the present study, ascorbate-treated and untreated rats were exposed to various cycles of intermittent light. Immediately after this light exposure, phagosome frequency in the RPE was morphologically evaluated in comparable 50 microns sections. In untreated rats, exposure to 2 or 3 hr of intermittent light resulted in a five- to sixfold increase in phagosome density compared to unexposed controls. In contrast, no increase in phagosome density was observed in ascorbate-treated rats. In these animals, under all lighting regimens, phagosome levels remained essentially identical to those in rats not exposed to light. After a single nondamaging light exposure, phagosome density remained at the level of dark controls in ascorbate-treated and untreated rats. These results indicate that phagosome frequency may serve as an index for light damage and that the protective effect of ascorbate may be linked to its capacity to prevent rod outer segment shedding and phagocytosis under intense light conditions.

Animals↗