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PubMed1995

Differential characteristics and regulation of arylamine and arylalkylamine N-acetyltransferases in the frog retina (Rana perezi).

Arylamine N-acetyltransferase activity (A-NAT: E.C.2.3.1.5) from Rana perezi retina was studied using p-phenetidine as specific substrate. Enzyme characteristics and regulation were compared with respect to the arylalkylamine N-acetyltransferase (AA-NAT: E.C.2.3.1.87) from the same tissue. A-NAT activity is distributed in both neural retina and choroid-pigmented epithelium complex, showing a 10-fold higher specific activity in neural retina. In contrast, AA-NAT activity is restricted to neural retina. Subcellular localization in neural retina indicated that both enzymatic activities are in the supernatant fraction (39,000 g, 20 min). p-Phenetidine acetylation was linear as a function of the neural retina amount in the assay (1/16 to 1 retina), and it is insensitive to phosphate buffer pH in the range 6.5-8.4. A-NAT kinetic showed a hyperbolic shape for both cosubstrates. Kinetic constants were KM = 11.2 microM, Vmax = 0.49 nmol/h/mg prot. for p-phenetidine (50 microM acetyl-CoA), and KM = 113.4 microM, Vmax = 3.1 nmol/h/mg prot. for acetyl-CoA (5 mM p-phenetidine). The additivity test for both enzymatic activities in retina homogenates demonstrated that both acceptor amines do not compete for the catalytic sites. Serotonin addition in the assay modifies differentially the kinetic characteristics of both enzymes. Serotonin acted as a strong mixed inhibitor, mainly competitive in nature (competitive Ki = 18.1 microM; non-competitive Ki = 1.9 mM) for AA-NAT. However, it acted as a weak inhibitor with respect to A-NAT, mainly non-competitive, (competitive Ki = 5.7 mM; non-competitive Ki = 8.7 mM).(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMedDate not supplied

Failure of dark adaptation to upregulate D-1 dopamine receptors in retina of senescent rats.

The effect of aging on the binding parameters of 3H-SCH 23390, the most selective ligand of D-1 DA receptors, was studied in membrane preparations from the rat retina. DA-stimulated adenylate cyclase activity was also measured in order to better characterize the changes in retinal D-1 DA receptors induced by aging. The binding studies revealed that the density of 3H-SCH 23390 was increased (34 and 73%) in the retina of 14- and 26-month-old rats, when compared to young adult animals, respectively. In contrast, aging failed to alter the sensitivity of the adenylate cyclase to the action of DA. In fact, DA (10(-6) M to 10(-4) M) elicited a similar enhancement in cyclic AMP formation in retinal homogenates of both adult and senescent rats. Since dark adaptation increases the density of D-1 DA receptors in the retina of adult rats we studied the effect of light deprivation on 3H-SCH 23390 binding and DA-sensitive adenylate cyclase activity in the retina of senescent rats. As previously shown (25) light deprivation increased 3H-SCH 23390 binding and enhanced DA-sensitive adenylate cyclase activity in the retina of young adult rats. On the contrary, dark adaptation failed to increase 3H-SCH 23390 binding and to enhance DA-sensitive adenylate cyclase activity in the retina of senescent rats. Taken together these results indicate that D-1 DA receptors in the retina of aged rats have biochemical and functional properties different from those found in the retina of adult animals; these changes may result in an altered response to the physiological stimuli elicited by environmental lighting.

Adenylyl Cyclases↗
PubMed1990

Analysis of the human, bovine and rat 33-kDa proteins and cDNA in retina and pineal gland.

A monoclonal antibody (mAb) was produced against a bovine retinal 33-kDa protein. Several clones of 33-kDa protein were isolated from each library of cDNA from human, bovine and rat retinas and rat pineal gland by mAb screening and by hybridization with cDNA probes. Each of the four cDNA sequences was determined and amino acid (aa) sequences were deduced from the nucleotide sequences. The latter were nearly identical in rat retina and rat pineal gland (99.6%) and were similar in human, bovine and rat retina (more than 87%). Each of these cDNAs had one long ORF and encoded 245 or 246 aa. The deduced aa sequences in rat retina and rat pineal gland were virtually identical and the sequences in human, bovine and rat retina were highly homologous (more than 88%). The predicted Mr for each of these proteins was 28,246 in the human, 28,176 in bovine, 28,143 in rat retina, and 28,129 in rat pineal gland. Each of the sequences has a putative site for phosphorylation by A kinase; we have confirmed that the putative site is Ser73. These results show that the 33-kDa proteins in the retina and pineal gland have the same sequences and the same phosphorylation site and suggest that the functional role of this protein is the same in the retina and pineal gland.

Amino Acid Sequence↗
PubMedDate not supplied

Morphological recovery in the reattached retina of the toad Bufo marinus: a new experimental model of retinal detachment.

BACKGROUND: The retinal pigment epithelium (RPE) of the toad (Bufo marinus) has been used in many studies as a model for understanding its role and interaction with the neural retina. The toad's retina has been used to establish a new in vitro model of experimental retinal detachment (RD) and replacement . It has been shown that the electrophysiological measures of retinal function recovered following complete RD. The toad was chosen because its RPE is similar to the mammalian RPE . In this report, light microscopy was used to characterize the morphologic changes that occur in the RPE and neural retina following RD/replacement and to correlate these findings with recovery of electrophysiologic function. METHODS: Retinas from Bufo marinus were studied in vitro. The neural retina completely detached from the RPE and then replaced. At various times after replacement, neural retina-RPE tissues were processed for light microscopy. RESULTS: At 30 min after replacement, the subretinal space was greatly expanded, and the apical processes that normally ensheath the rod outer segments were short and no longer contacted the rod outer segments. The RPE was swollen, contained many vacuoles and the apical surface was rounded. By 2 h after replacement, the subretinal space was significantly resorbed and contained many shredded rod outer segments; RPE cells were still swollen, although less. During the next 5-10 h, the number of phagosomes in the RPE cytoplasm increased and the number of shredded rod outer segments in the subretinal space decreased. RPE cells regained their normal size and interdigitation of apical processes and rod outer segments were observed. CONCLUSIONS: These results demonstrate the re-establishment of morphological interactions between the RPE and neural retina within hours following RD/replacement. Morphological recovery coincides with recovery of electrophysiologic parameters. This is a good model to investigate the retinal pigment epithelium (RPE) and neural retina mechanisms involved in retinal adhesion and recovery from retinal detachment.

Animals↗
PubMed2004

Developmental changes of aggrecan, versican and neurocan in the retina and optic nerve.

We have used a monoclonal antibody to neurocan and specific polyclonal antibodies to the non-homologous glycosaminoglycan attachment regions of aggrecan and mRNA splice variants of versican to compare the localization and developmental changes of these structurally related hyaluronan-binding chondroitin sulfate proteoglycans in the rat retina and optic nerve. Staining for aggrecan and versican was first seen at embryonic day 16 in the optic nerve and retina, whereas neurocan was not detected in the embryonic eye. At postnatal day 0 (P0), beta-versican staining is largely confined to the inner plexiform layer whereas alpha-versican is also apparent in the neuroblastic layer. Both aggrecan and, much more weakly, neurocan immunoreactivity is present throughout the neonatal retina. At P9, aggrecan and versican immunoreactivity is most intense in the inner and outer plexiform and ganglion cell layers, accompanied by diffuse staining in the inner and outer nuclear layers. Aggrecan and alpha-versican are also present throughout the optic nerve and disk, whereas beta-versican and neurocan are confined to the laminar beams of the optic nerve. Between P0 and P9 there is a marked increase in beta-versican expression in the inner and outer nuclear layers and in the outer plexiform layer, whereas there is only weak staining of neurocan in the inner plexiform and ganglion cell layers of P9 retina. By 1 month postnatal the staining pattern of the fully differentiated retinal layers is essentially identical to that seen in the adult, where there is strong aggrecan and alpha-versican immunoreactivity in the retina and optic nerve, whereas beta-versican has essentially disappeared from the adult retina and, similarly to neurocan, is present only in the laminar beams of the optic nerve. The marked decrease of beta-versican in the retina is consistent with >90% decrease in its concentration in brain during postnatal development, suggesting that the developmental time-course for these proteoglycans in retina parallels that seen in other areas of the central nervous system.

Aggrecans↗
PubMed2005

Changes in rhodopsin kinase and transducin in the rat retina in early-stage diabetes.

To establish changes in phototransduction in diabetes, the effects of high glucose on rhodopsin kinase (RK) and transducin (G(t)), as well as recoverin, were examined in the retina of STZ-induced diabetic rats. Diabetes was induced by single intraperitoneal injection of STZ (50mg/kg) to Sprague-Dawley (SD) rats and the animals were sacrificed after 6 weeks. Immunohistochemistry (IHC) and Western blot analysis were carried out using antibodies against RK and G(talpha) (alpha subunit of G(t)) in the STZ-induced diabetic retina and the control retina. The expression level of recoverin protein was also analysed. In the diabetic retina, while the expression of RK protein increased, that of G(talpha) and recoverin proteins decreased. RK immunoreactivity (IR) appeared generally in the retina, and its signal increased in the outer limiting membrane (OLM), some rod cells in the outer segment layer (OSL) and at the tip of the outer plexiform layer (OPL) in the diabetic retina. G(talpha)-IR also appeared in the OPL and in photoreceptor layer. In the diabetic retina, G(talpha)-IR significantly decreased in the OPL, indicating RK-IR increase. This study illustrates the alterations in RK, G(talpha) and recoverin in the diabetic retina that may induce dysfunctions in phototransduction even in early-stage diabetes.

Animals↗
PubMed2005

The protective role of squalene in alcohol damage in the chick embryo retina.

The developing CNS, and in particular the visual system, is very sensitive to the effects of alcohol. Alcohol causes lipid peroxidation. Squalene, the major olive oil hydrocarbon, is a quencher of singlet oxygen and prevents the corresponding lipid peroxidation. We presumed that squalene can protect against the alcohol-induced damage already observed during the development of the chick retina. Alcohol+squalene was administered directly into the yolk sac of the egg of White Leghorn chicks at day 6 of incubation. The lipid composition of the retina was analyzed in embryos at E7, E11, E15 and E18. The proportions of phospholipids, free and esterified cholesterol, diacylglycerides and free fatty acids were estimated using the Iatroscan TLC/FID procedure. Gas chromatography and mass spectrometry were used to determine the fatty acid composition. The morphological study was carried out at E11 using semithin sections, and by means of immunohistochemical techniques at E19. Comparing the results obtained in control embryos, the administration of alcohol+squalene reduces the effects of alcohol on the total lipid composition of the retina during development. The effects were, in fact, of less magnitude than in embryos treated only with alcohol. The major phospholipid species of alcohol+squalene-treated embryos exhibited total recuperation at E15. As far as fatty acids are concerned, no significant changes were observed with regard to control embryos during development. From a morphological point of view, the retinas of alcohol+squalene-treated embryos show at E11 fewer cellular alterations than the retinas of alcohol-treated embryos. In this respect, the retinas of alcohol+squalene-treated embryos exhibited: a columnar cell arrangement similar to that observed in control retinas; few pycnotic cells and very few alterations in ganglion cell layers and in the optic nerve fibers layer. At E19 the recuperation of the expression of myelin oligodendrocyte specific protein (MOSP) in alcohol+squalene-treated embryos was recorded. Since squalene reduces the deleterious effects caused by alcohol on the lipid composition and the structure of the retina, squalene could act as a naturally occurring agent for the prevention of damage caused by abusive alcohol ingestion during pregnancy.

Animals↗
PubMed2006

Expression of nicotinamide adenine dinucleotide phosphate-diaphorase in the retina of postnatal golden hamsters deprived of light stimulation.

Nicotinamide Adenine Dinucleotide Phosphate-Diaphorase (NADPH-d) expressing neurons in the retina of golden hamsters have been identified to be a subset of amacrine cells that provide a major source of Nitric Oxide (NO) in retina. This subset of amacrine cells in mouse retina was recently proved to contain the circadian clock gene Per1 (D.Q. Zhang, T. Zhou, G.X. Ruan, D.G. McMahon, Circadian rhythm of Period 1 clock gene expression in NOS amacrine cells of the mouse retina, Brain Res., 1050 (2005) 101-109). However, it remains unknown whether these clock-related NADPH-d amacrine cells can be regulated by light stimulation and thus synchronized to ambient day/night cycle. A previous study has reported that NADPH-d expressing amacrine cells in postnatal hamsters exhibited a surge after eye-opening (D. Tay, Y.C. Diao, Y.M. Xiao, K.F. So, Postnatal development of nicotinamide adenine dinucleotide phosphate-diaphorase-positive neurons in the retina of the golden hamster, J. Comp. Neurol., 446 (2002) 342-348) suggesting a possible effect of light on the NADPH-d amacrine cells. In order to further reveal the relationship between NADPH-d amacrine cells and light stimulation, the present study focuses on the changes of the expression of NADPH-d in the retina of postnatal hamsters reared in completely deprived light conditions. Prior to eye opening, P12 hamster pups were subjected to either bilateral eyelid suturing or dark rearing. On P28 a subgroup of light deprived hamsters was returned to lighting conditions and the expression of NADPH-d activities in the retina was assessed. In hamsters reared in the 12:12 light-dark cycle, the number of NADPH-d amacrine cells in the ganglion cell layer (GCL) increased right after eye-opening and reached the adult level gradually. However, hamsters subjected to both bilateral eyelid suturing and dark rearing, the number of NADPH-d amacrine cells in GCL was maintained at a low level but increased again upon returning to the 12:12 light-dark condition. In contrast, the number of NADPH-d expressing amacrine cells in the inner nuclear layer (INL) remained low and unaltered regardless of the lighting environment. This study demonstrates that there are two subpopulations of NADPH-d expressing amacrine cells with respect to different locations in the retina of hamsters. Different from those in INL, the NADPH-d amacrine cells in GCL of postnatal hamsters are dependent on the lighting environment implicating that these clock-related amacrine cells and the production of NO might be under a modulation of light stimulation.

Animals↗
PubMed2006

Ischemia-reperfusion alters the immunolocalization of glial aquaporins in rat retina.

Glial cells control the retinal osmohomeostasis, in part via mediation of water fluxes through aquaporin (AQP) water channels. By using immunohistochemical staining, we investigated whether ischemia-reperfusion of the rat retina causes alterations in the distribution of AQP1 and AQP4 proteins. Transient ischemia was induced in retinas of Long-Evans rats by elevation of the intraocular pressure for 60 min. In control retinas, immunoreactive AQP1 was expressed in the outer retina and by distinct amacrine cells, and AQP4 was expressed by glial cells (Müller cells and astrocytes) predominantly in the inner retina. After ischemia, retinal glial cells in the nerve fiber/ganglion cell layers strongly expressed AQP1. The perivascular staining around the superficial vessels altered from AQP4 in control retinas to AQP1 in postischemic retinas. The data suggest that the glial cell-mediated water transport in the retina is altered after ischemia especially at the superficial vessel plexus.

Animals↗
PubMed2005

Cytochrome-c oxidase is one of several genes elevated in marginal retina of the chick embryo.

The retinal ciliary margin is particularly relevant for the correct generation and regeneration of vertebrate retinae, since pluripotent stem cells are located there throughout development, and--at least in some species--even until adult stages. Our aim was to identify factors (genes) which are involved in processes of proliferation and differentiation in the developing chicken retina. Reverse transcription-polymerase chain reaction differential display was used to identify genes that were differentially expressed in chick central and peripheral embryonic retina. Candidate genes analyzed through sequencing and database searches were confirmed by Northern blot analysis and histochemistry. A series of differentially expressed genes were detected, including a neuronal cell adhesion molecule, an esterase, and homeobox gene products. One of the sequenced products was identified as subunit I of cytochrome-c oxidase (COX-1), an enzyme which is central to energy metabolism and particularly relevant for developing nervous systems. Northern blot analysis confirmed its up-regulation in the chick peripheral retina, being maximal at embryonic day 7. In the retinal pigmented epithelium its expression is lower than in the retinal periphery but higher than in central retina. COX histochemistry revealed distinct laminar patterns in central retina, but also an elevated level of activity in the peripheral retina throughout development. These data not only show that the developing ciliary margin of the chick retina has high energy requirements, but also indicate that COX-1 could play essential roles in developing cells and in stem cells of the eye periphery.

Age Factors↗
PubMed1999

Choline acetyltransferase and acetylcholinesterase in the normal, developing and regenerating newt retinas.

The presence of the choline acetyltransferase (ChAT) and acetylcholinesterase (AChE) was demonstrated in the adult newt retina using immunocytochemical and histochemical techniques. Within the inner plexiform layer (IPL), two ChAT-positive bands were detected at relative depths of 0-15% and 45-60% of the total thickness (100%) of the IPL. AChE-positive band occupied approximately 0-60% of the IPL width with an intensive AChE-positive band at a depth of 20-40% within the IPL. Localizations of maximum ChAT and AChE activity were not exactly the same in the IPL of the mature retina. To elucidate whether retinal regeneration follows the same sequence of cellular differentiation steps that occur in retinal development, we examined the time course of appearance of the cholinergic neurons and AChE activity in both developing and regenerating retinas. The ChAT-positive cells were first detected in the retina just before or at the beginning of the morphological development of the IPL in both developing and regenerating retinas. AChE activity first became detectable in somata located at the most proximal layer of the retina before the ChAT-positive cells could be detected and well before the IPL developed in both developing and regenerating retinas. During subsequent development and regeneration, the outer plexiform layer, the IPL, and somata close to either side of the IPL became AChE-positive. The fact that the time course of the appearance of ChAT and AChE molecules during regeneration was similar to that observed during development suggests that common mechanisms may control both the development and the regeneration of the newt retina.

Acetylcholinesterase↗
PubMed2003

Circadian oscillator function in embryonic retina and retinal explant cultures.

Retinal circadian oscillators regulate many aspects of retinal function. Investigations of these oscillators and the biochemical cascades that entrain them would be greatly facilitated if experimental paradigms could be identified that permit long-term monitoring of retinal circadian oscillator function in vitro. The purpose of this study was to determine if chicken retinas maintained in explant culture conditions could serve in this capacity. Retinal circadian oscillator function was studied by monitoring iodopsin transcription under cyclic light, constant dark, and following reversal of the light cycle. Rhythms observed in the explant cultures were compared to those observed in retinas of embryos (in ovo) and post-hatch chickens. Robust iodopsin transcript rhythms were observed for up to 9 days in explant cultures maintained under cyclic light. These rhythms persisted for 48 h in constant darkness and the time course for re-entrainment of the rhythm to a reversed light/dark cycle was similar to that observed in post-hatch chicken retinas. These results show that circadian oscillators located within the retina play a key role in the regulation of iodopsin transcription in retinal explant cultures and in retinas of post-hatch chickens. Interestingly, our data show that iodopsin transcription in retinas of intact embryos is primarily, if not entirely, driven by light. These results show that the circadian oscillators driving iodopsin transcription in embryonic retinal explant cultures exhibit functional characteristics similar to those found in post-hatch chicken retina, supporting use of this paradigm in further studies of entrainment of these oscillators in retina.

Animals↗
PubMed2000

Difference in glutamate release between retina and cerebral cortex following ischemia.

The difference in ischemic tolerance between the retina and cerebral cortex may be attributable to a difference in glutamate release during ischemia. Glutamate release in the retina and the cerebral cortex was compared in rats. A dialysis electrode for real-time glutamate measurement was perfused with L-glutamate oxidase, and the current evoked between two voltage-clamped electrodes was detected. Two electrodes were implanted in the retina through the choroid and cerebral cortex in 12 anesthetized rats, each mounted on a stereotaxic frame. Global ischemia was induced by ligation on both carotid arteries and hypotension was induced by blood withdrawal. Under control conditions, the glutamate concentration in the retina was 164 +/- 231 (mean +/- standard deviation) microM, being significantly higher (P < 0.05) than that in the cerebral cortex (83 +/- 105 microM). In 10 of the 12 animals, the glutamate concentration in the retina decreased to a minimum of 134 +/- 149 microM (P < 0.01, compared with the value for the cerebral cortex), but that in the cortex increased to 410 +/- 305 microM (averaged highest value). Immediately after the start of reperfusion, the glutamate concentration in the cortex decreased rapidly to 101 +/- 27 microM, but that in the retina increased gradually to almost the control level (148 +/- 204 microM). In the other two animals, the glutamate concentration remained unchanged. In conclusion, glutamate release in the retina does not proceed as rapidly as that in the cerebral cortex during 20 min of ischemia, and in fact decreases. This opposite trend shown by the two organs may be due to the slow depletion rate of ATP in the retina. This may explain the differing neuronal tolerance to ischemia in these two organs.

Animals↗
PubMed1998

The eyes of deep-sea fish. II. Functional morphology of the retina.

Three different aspects of the morphological organisation of deep-sea fish retinae are reviewed: First, questions of general cell biological relevance are addressed with respect to the development and proliferation patterns of photoreceptors, and problems associated with the growth of multibank retinae, and with outer segment renewal are discussed in situations where there is no direct contact between the retinal pigment epithelium and the tips of rod outer segments. The second part deals with the neural portion of the deep-sea fish retina. Cell densities are greatly reduced, yet neurohistochemistry demonstrates that all major neurotransmitters and neuropeptides found in other vertebrate retinae are also present in deep-sea fish. Quantitatively, convergence rates in unspecialised parts of the retina are similar to those in nocturnal mammals. The differentiation of horizontal cells makes it unlikely that species with more than a single visual pigment are capable of colour vision. In the third part, the diversity of deep-sea fish retinae is highlighted. Based on the topography of ganglion cells, species are identified with areae or foveae located in various parts of the retina, giving them a greatly improved spatial resolving power in specific parts of their visual fields. The highest degree of specialisation is found in tubular eyes. This is demonstrated in a case study of the scopelarchid retina, where as many as seven regions with different degrees of differentiation can be distinguished, ranging from an area giganto cellularis, regions with grouped rods to retinal diverticulum.

Animals↗
PubMed1998

Oscillatory potentials in the retina: what do they reveal.

This chapter is an overview of current knowledge on the oscillatory potentials (OPs) of the retina. The first section describes the characteristics of the OPs. The basic, adaptational, pharmacological and developmental characteristics of the OPs are different from the a- and b-waves, the major components of the electroretinogram (ERG). The OPs are most easily recorded in mesopic adaptational conditions and reflect rapid changes of adaptation. They represent photopic and scotopic processes, probably an interaction between cone and rod activity in the retina. The OPs are sensitive to disruption of inhibitory (dopamine, GABA-, and glycine-mediated) neuronal pathways and are not selectively affected by excitatory amino acids. The earlier OPs are associated with the on-components and the late OPs with the off-components in response to a brief stimulus of light. The postnatal appearance of the first oscillatory activity is preceded by the a- and b-waves. The earlier OPs appear postnatally prior to, and mature differently from, the later ones. The second section deals with present views on the origin of the OPs. These views are developed from experimental studies with the vertebrate retina including the primate retina and clinical studies. Findings favor the conclusion that the OPs reflect neuronal synaptic activity in inhibitory feedback pathways initiated by the amacrines in the inner retina. The bipolar (or the interplexiform) cells are the probable generators of the OPs. Dopaminergic neurons, probably amacrines (or interplexiform cells), are involved in the generation of the OPs. The earlier OPs are generated in neurons related to the on-pathway of the retina and the later ones to the off-channel system. Peptidergic neurons may be indirectly involved as modulators. The individual OPs seem to represent the activation of several retinal generators. The earlier OPs are more dependent on an intact rod function and the later ones on an intact cone system. Thus, the OPs are good indicators of neuronal adaptive mechanisms in the retina and are probably the only post-synaptic neuronal components that can be recorded in the ERG except when structured stimuli are used. The last section describes the usefulness of the oscillatory response as an instrument to study the postnatal development of neuronal adaptation of the retina. In this section clinical examples of of the sensitivity of the OPs for revealing early disturbance in neuronal function in different retinal diseases such as pediatric, vascular and degenerative retinopathies are also given.

Animals↗
PubMed1992

Light-sensitive melatonin synthesis by Xenopus photoreceptors after destruction of the inner retina.

Several lines of evidence indicate that retinal photoreceptors produce melatonin. However, there are other potential melatonin sources in the retina, and melatonin synthesis can be regulated by feedback from the inner retina. To analyze cellular mechanisms of melatonin regulation in retinal photoreceptors, we have developed an in vitro method for destruction of the inner retina that preserves functional photoreceptors in contact with the pigment epithelium. Eyecups, which include the neural retina, retinal pigment epithelium, choriod, and sclera were prepared. The vitreal surface of the retina in each eyecup was washed sequentially with 1% Triton X-100, water, and culture medium. This lysed the ganglion cells and neurons and glia of the inner nuclear layer, causing the retina to split apart within the inner nuclear layer. The damaged inner retina was peeled away, leaving photoreceptors attached to the pigment epithelium. The cell density of the inner nuclear layer was reduced 94% by this method, but there was little apparent damage to the photoreceptors. Lesioned eyecups produced normal melatonin levels in darkness at night, and melatonin production was inhibited by light. These results indicate that the inner retina is not necessary for melatonin production nor for regulation of photoreceptor melatonin synthesis by light. The lesion method used in this study may be useful for other physiological and biochemical studies of photoreceptors.

Animals↗
PubMedDate not supplied

Characterization and localization of an aldehyde dehydrogenase to amacrine cells of bovine retina.

An enzyme of bovine retina that catalyzes oxidation of retinaldehyde to retinoic acid was purified to homogeneity and a monoclonal antibody (mAb H-4) was generated. MAb H-4 recognized a single component (Mr = 55,000) in extracts of bovine retina and other bovine tissues. The antibody showed no cross-reactivity with extracts of rat, monkey, or human retinas. A 2067 bp cDNA was selected from a retina cDNA expression library using mAb H-4. The cDNA hybridized with a similarly sized, moderately abundant mRNA prepared from bovine retina. Nucleotide sequence analysis indicated that the cDNA contained a single open reading frame encoding 501 amino acids that have 88% sequence identity with the amino-acid sequence of human hepatic Class 1 aldehyde dehydrogenase. Amino-acid sequence analysis of purified enzyme demonstrated that the cDNA encodes the isolated enzyme. MAb H-4 specifically labeled the somata and processes of a subset of amacrine cells in bovine retinal sections. Labeled amacrine somata were located on both sides of the inner plexiform layer, and their processes ramified into two laminae within the inner plexiform layer. The inner radial processes of Müller (glial) cells were weakly reactive with mAb H-4. Weak immunostaining of amacrine cells was found in monkey retina with mAb H-4, but no signal was detected in rat or human retina. The results provide further evidence for metabolism and function of retinoids within cells of the inner retina and define a novel class of retinal amacrine cells.

Aldehyde Dehydrogenase↗
PubMedDate not supplied

Angiotensin II in the rabbit retina.

We investigated a putative local angiotensin II (AngII) system in the rabbit retina by examining AngII contents in the retina, vitreous humor, and choroid by radioimmunoassays and AngII synthesis in the retina and choroid by detection of angiotensin converting enzyme (ACE) mRNA. An antibody directed against AngII was used to localize possible cellular sources of AngII in the retina. To enhance immunoreactivity and to further examine AngII metabolism, tissues were preincubated in medium containing either protease inhibitors (PI), PI together with the AngII-precursor AngI, or PI and AngII. In some experiments the conversion of AngI to AngII was blocked by an ACE inhibitor. AngII concentration in the vitreous humor was only about 10% of the plasma concentration; in the retina and the choroid, however, AngII concentrations were 10 and 86 times higher, respectively, than in the plasma. ACE mRNA was present in both retina and choroid. Immunohistochemistry for AngII revealed faintly labeled amacrine cells at the inner border of the inner nuclear layer of the retina. Preincubation with PI resulted in an enhanced immunoreaction and in the labeling of fibers in the inner and outer plexiform layer; Müller cells and their processes as well as ganglion cells were now stained as well but the specificity of ganglion cell staining remains questionable. The immunoreaction was further enhanced when AngI or AngII was added to the incubation medium, whereas labeling totally disappeared when the conversion of AngI to AngII was blocked. No immunoreactive cells were detected in the choroid. In conclusion, the synthesizing enzyme for AngII is expressed in the retina and a specific AngII concentration is maintained there; AngII is localized in distinct cell types and can be metabolized within these cells. These data point to a local retinal AngII system that is protected and independent of blood-borne AngII.

Angiotensin II↗