Impact of smoking, alcohol consumption, and NSAID use on risk for and phenotypes of eosinophilic esophagitis.

There are few data exploring modifiable risk factors for eosinophilic esophagitis (EoE). We aimed to determine if smoking, alcohol consumption, and nonsteroidal anti-inflammatory drug (NSAID) use were risk factors for EoE, and to assess their impact on EoE phenotypes and treatment outcomes. We performed a case-control study analyzing data collected from a prospective cohort of adults undergoing upper endoscopy for symptoms of esophageal dysfunction. Incident EoE cases were diagnosed via consensus guidelines. Exposure data were collected via standardized patient questionnaire. Follow-up assessments for cases were made after treatment, with histologic response defined as <15 eosinophils per high-power field (eos/hpf). Exposures were compared between EoE cases and controls, among EoE cases with and without fibrostenosis, and among EoE responders and nonresponders. A total of 115 cases and 225 controls were analyzed. Cases were less likely to have ever smoked cigarettes (23% vs. 47%, P < 0.001) or currently use NSAIDs (17% vs. 40%, P < 0.001) compared to controls. These relations persisted after multivariate analysis. Although alcohol use was more common among cases (75% vs. 51%, P < 0.001), the effect was abrogated after multivariate analysis. Smoking, alcohol, and NSAID use were not associated with the fibrostenotic phenotype. There was a trend toward improved histologic response among EoE patients concomitantly using NSAIDs (87% vs. 63%, P = 0.08; aOR 6.97 (95% CI: 0.81-60.3). In conclusion, NSAID and smoking were inversely associated with EoE compared to endoscopy-based controls. Alcohol use was more prevalent in the EoE cases, although not an independent risk factor. Concomitant NSAID use may improve treatment response and is worthy of future study.

Inositol incorporation into phosphoinositides in retinal horizontal cells of Xenopus laevis: enhancement by acetylcholine, inhibition by glycine.

The absorption of light by photoreceptor cells leads to an increased incorporation of [2-3H]inositol into phosphoinositides of horizontal cells in the retina of Xenopus laevis in vitro. We have identified several retinal neurotransmitters that are involved in regulating this response. Incubation with glycine, the neurotransmitter of an interplexiform cell that has direct synaptic input onto horizontal cells, abolishes the light effect. This inhibition is reversed by preincubation with strychnine. Acetylcholine added to the culture medium enhances the incorporation of [2-3H]inositol into phosphoinositides in horizontal cells when retinas are incubated in the dark. This effect is inhibited by preincubation with atropine. However, atropine alone does not inhibit the light-enhanced incorporation of [2-3H]inositol into phosphoinositides in the retina. gamma-Aminobutyric acid, the neurotransmitter of retinal horizontal cells in X. laevis, as well as dopamine and norepinephrine, have no effect on the incorporation of [2-3H]inositol into phosphoinositides. These studies demonstrate that the light-enhanced incorporation of [2-3H]inositol into phosphoinositides of retinal horizontal cells is regulated by specific neurotransmitters, and that there are probably several synaptic inputs into horizontal cells which control this process.

Membrane morphogenesis in retinal rod outer segments: inhibition by tunicamycin.

Isolated Xenopus laevis retinas were incubated with 3H-labeled mannose or leucine in the presence or absence of tunicamycin (TM), a selective inhibitor of dolichyl phosphate-dependent protein glycosylation. At a TM concentration of 20 micrograms/ml, the incorporation of [3H]mannose and [3H]leucine into retinal macromolecules was inhibited by approximately 66 and 12-16%, respectively, relative to controls. Cellular uptake of the radiolabeled substrates was not inhibited at this TM concentration. Polyacrylamide gel electrophoresis revealed that TM had little effect on the incorporation of [3H]leucine into the proteins of whole retinas and that labeling of proteins (especially opsin) in isolated rod outer segment (ROS) membranes was negligible. The incorporation of [3H]mannose into proteins of whole retinas and ROS membranes was nearly abolished in the presence of TM. Autoradiograms of control retinas incubated with either [3H]mannose or [3H]leucine exhibited a discrete concentration of silver grains over ROS basal disc membranes. In TM-treated retinas, the extracellular space between rod inner and outer segments was dilated and filled with numerous heterogeneously size vesicles, which were labeled with [3H]leucine but not with [3H]mannose. ROS disc membranes per se were not labeled in the TM-treated retinas. Quantitative light microscopic autoradiography of retinas pulse-labeled with [3H]leucine showed no differences in labeling of rod cellular compartments in the presence or absence of TM as a function of increasing chase time. These results demonstrate that TM can block retinal protein glycosylation and normal disc membrane assembly under conditions where synthesis and intracellular transport of rod cell proteins (e.g., opsin) are not inhibited.

Turnover of rod photoreceptor outer segments. I. Membrane addition and loss in relationship to temperature.

Membrane turnover in outer segments of Rana pipiens red rods (ROS) was studied in tadpoles maintained under cyclic lighting (12L:12D) at 23 degrees, 28 degrees, and 33 degrees C. Large fragments (greater than 2 microns in diameter or length) were shed from the ROS tips shortly after the onset of light. These were phagocytized by the pigment epithelium (PE) which caused an increase in the number of phagosomes greater than 2 microns in size (large phagosomes). Large phagosomes were present in highest numbers 2-4 h after light exposure and were degraded by 8-12 h. The proportion of ROS that shed each day after the onset of the light cycle increased with increment increases in temperatures (23 degrees C-18%, 28 degrees C-33%, 33 degrees C-42% per day), resulting, in a reduction in the average interval of time between repeated sheddings (23 degrees C-5.6 days, 28 degrees C-3 days, 33 degrees C-2.4 days) though the average numbers of disks shed from ROS at the various temperatures were not significantly different (23 degrees C-139.5 +/- 5.7, 28 degrees C-129.4 +/- 7.6, 33 degrees C-129.9 +/- 4.8 disks/shed packet). Phagosomes in the PE that were less than 2 microns in diameter (small phagosomes) were present in relatively constant numbers throughout the day, and their numbers increased at higher temperatures. The absence of a concomitant increase in small phagosomes as large phagosomes were degraded indicates that large phagosomes were not the major source of small phagosomes. When the PE was isolated to culture in the absence of the retina, these small phagosomes were degraded. The rate of disk addition to the ROS base was determined by autoradiography after [3H]leucine injection. The number of disks added per day increased with elevations of temperature (23 degrees C-32.4; 28 degrees C-55.9; 33 degrees C-65.5). The average number of disks added to the ROS between repeated sheddings (23 degrees C-181.4; 28 degrees C-167.7; 33 degrees C-157.2) was greater than the number of disks shed after light exposure. Inasmuch as the ROS show no net increase in length during the tadpole stages utilized, the remaining disks must be lost at some other time. Electron microscope analysis revealed the presence of small groups of disks in curled configurations at the tips of ROS, suggesting possible stages of detachment.(ABSTRACT TRUNCATED AT 400 WORDS)

Turnover of rod photoreceptor outer segments. II. Membrane addition and loss in relationship to light.

The rate of disk addition to rod outer segments (ROS) varies widely in Xenopus laevis tadpoles kept in cyclic light (12L:12D). When measured as radioactive band (3H-band) displacement during the 2nd day after injection of [3H]leucine, 75% of the daily increment of displacement occurred during the first 8 h of light. During the same interval, the number of open disks at the ROS base increased more than threefold. During the last 8 h of darkness, 3H-band displacement was undetectable and the number of open disks was reduced. These observations suggest the possibility that disk addition may occur discontinuously. During the 3rd and 4th days after injection of [3H]leucine, maximal displacement of the 3H-band occurred later in the day than on the 2nd day, its movement no longer corresponding to the increase in open disks. This delay in 3H-band displacement may reflect a time delay as a result of propagation of compressive stress in an elastic ROS system. Maximal disk loss from ROS as reflected in counts of phagosomes in the pigment epithelium occurred within 1 h of light exposure, and phagosome counts remained high for 4 h before declining to a low level in darkness. Modified lighting regimes affected the daily rhythms of shedding and disk addition differently, suggesting that control mechanisms for the two processes are not directly coupled. During 3 days in darkness, disk addition was reduced 50% compared to controls (12L:12D), whereas shedding was reduced by about 40%. Although reduced in level, shedding occurred as a free-running circadian rhythm. There was no evidence of rhythmicity of disk addition in darkness. In constant light, the rate of disk addition was not different from controls, but shedding was reduced by about 80% after the 1st day. This resulted in a 21% increase in ROS length. Among animals kept on a 2.5L:21.5D cycle, the rate of disk addition was reduced by 40% while shedding was maintained near control levels, resulting in a slight decrease in ROS length. These observations indicate that normal shedding requires alternating light and darkness, and that the daily rhythm of disk addition is due primarily to daily stimulation by light.

Endocytosis and degradation of interstitial retinol-binding protein: differential capabilities of cells that border the interphotoreceptor matrix.

Between the pigment epithelium and the outer limiting membrane of the retina is an extracellular compartment filled with the interphotoreceptor matrix (IPM). A prominent component of the IPM is a glycoprotein known as interstitial retinol-binding protein (IRBP). Using in vitro techniques, we compared the ability of the cells that border this compartment to internalize colloidal gold (CG) coated with either IRBP or ovalbumin, a glycoprotein not found in the IPM. Neither IRBP-CG nor ovalbumin-CG was internalized by the Muller's cells. Both rod and cone photoreceptors take up IRBP-CG, which is observed in small vesicles and multivesicular bodies. Neither photoreceptor type takes up ovalbumin-CG. Acid phosphatase cytochemistry indicates that acid phosphatase reaction product in the multivesicular bodies co-localizes with IRBP-CG, which suggests that this molecule is degraded by rod and cone photoreceptors and is not recycled. The pigment epithelium internalizes IRBP-CG and ovalbumin-CG, both of which remain in small cytoplasmic vesicles near the apical plasma membrane. There is no indication that vesicles that contain either IRBP-CG or ovalbumin-CG fuse with the lysosomal system in the pigment epithelial cells during the incubation.

Photoreceptor outer segments: accelerated membrane renewal in rods after exposure to light.

The rate of rod outer segment renewal in Rana pipiens tadpoles under constant light and under diurnal conditions of 12 or 2 hours light per day is significantly increased compared to that in animals in darkness. Furthermore, during 24 hours in light after 6 days in darkness the rate of renewal is three to four times that in darkness. In Xenopus laevis tadpoles the rate of renewal is more than five times greater during the first 8 hours of a normal diurnal cycle than during the following 16 hours. These observations demonstrate that bursts of renewal activity occur as a response to light, and suggest that a normal pattern of light alternating with darness plays a fundamental role in the regulation of rod outer segment turnover.

Cyclic GMP accumulation causes degeneration of photoreceptor cells: simulation of an inherited disease.

Guanosine 3',5'-monophosphate (cyclic GMP) metabolism in developing eye rudiments of Xenopus laevis embryos in culture is disrupted by the phosphodiesterase inhibitor isobutylmethylxanthine. At low concentrations of inhibitor the rudiments develop normally, but at higher concentrations of the inhibitor, cyclic GMP accumulates in the rudiments and the retinal photoreceptor cells degenerate selectively. The isobutylmethylxanthine-induced photoreceptor degeneration is associated with an accumulation of cyclic GMP and, in this respect, it stimulates an early biochemical defect in the inherited degenerative disease of rd mice.

Light stimulates the incorporation of inositol into phosphatidylinositol in the retina.

The incorporation of [3H]inositol into phosphatidylinositol in retinas of Xenopus laevis tadpoles or young adults in short-term organ culture is stimulated by light, compared to retinas maintained under identical conditions in darkness. Over 95% of the label incorporated into lipid was in phosphatidylinositol, and none was incorporated into retinal proteins. The stimulation of incorporation was localized by autoradiography to the outer plexiform layer, a neurophil composed primarily of horizontal cell processes.

Histopathology of the Retina from a Three Year-Old Suspected to Have Joubert Syndrome.

PURPOSE: To define the retinal pathology in a 3 year-old eye donor who died from complications of an undiagnosed genetic syndrome. METHODS: Eyes were fixed and analyzed using macroscopic fundus photography (MF), confocal scanning laser ophthalmoscopy (cSLO) and spectral-domain optical coherence tomography (SD-OCT). Small areas from the perifovea and periphery were processed for histology and indirect immunofluorescence, using antibodies specific to retinal proteins such as rhodopsin, cone arrestin, RPE65 and others. Available medical records were also reviewed. RESULTS: With all three imaging modalities, the affected donor's eyes lacked the distinct morphological detail typically observed with these techniques in postmortem control eyes. MF images showed a "photonegative effect" due to a hypopigmented macula relative to a hyperpigmented retinal background. cSLO imaging demonstrated a weak autofuorescence signal that was largely devoid of the usual retinal structures compared to the control. SD-OCT suggested disorganization of the affected retina, absence of a photoreceptor layer, and degeneration of the choroid in the macular area. Histologic findings indicated a highly disorganized photoreceptor layer in the macula and periphery. The RPE layer displayed thinning in some regions of the periphery and decreased pigmentation in most areas. Rods and cones were significantly reduced in the affected retina but a few cones were detected in the perifovea. Centrin-2 labeling was mostly absent from the connecting cilium of the photoreceptor cells. Medical record review pointed to a possible clinical diagnosis of Joubert syndrome. CONCLUSIONS: The retinal degenerative findings, and absence of centrin-2 labeling are compatible with the expected retinal phenotype in patients with Joubert syndrome.

Retinal ganglion cell morphology in the frog, Rana pipiens.

The morphology of retinal ganglion cells in the frog, Rana pipiens, has been examined in retinal flatmounts following backfilling of axons with horseradish peroxidase (HRP). Size and shape of the cell body and of the dendritic arbor, the dendritic branching pattern, and the depth of dendritic arborization within the inner plexiform layer (IPL) were all used to classify these cells. All of the ganglion cells so visualized can be grouped into one of 7 distinct cell classes. Class 1 contains the largest ganglion cells, with a soma size of 323 +/- 5.3 microns2 and dendritic fields of 86,819 +/- 11,817 microns2; the dendrites branch within strata 1 and 2 of the IPL. The second largest cells are class 2, with somas of 245 +/- 19.7 microns2 and dendritic fields of 55,983 +/- 7,392 microns2; the dendrites also branch within strata 1 and 2 of the IPL. Class 3 cells are the next largest class with somas of 211 +/- 11.8 microns2 and dendritic fields of 18,186 +/- 1,394 microns2; there are three varieties of class 3 cells based on the depth of branching of the dendrites: some cells are bistratified, others are tristratified, while still other cells arborize diffusely within the IPL. Class 4 cells are intermediate in size, with somas of 113 +/- 7.4 microns2 and dendrites of 4800 +/- 759 microns2; the dendrites arborize within strata 4 and 5 of the IPL. Class 5 cells have not been quantitatively analyzed because they are heterogeneous in soma and dendritic size. However, class 5 cells all have cell bodies displaced in location into the inner nuclear layer and all have a unique dendritic specialization: they send from 1 to 3 processes into the outer plexiform layer. Class 6 cells are the second smallest cell class with somas of 68.1 +/- 5.13 microns2 and dendritic fields of 888 +/- 182 microns2; the dendrites arborize within strata 3, 4, and 5 of the IP. Class 7 contains the smallest ganglion cells with somas of 62.1 +/- 2.86 microns2 and dendritic fields of 831 +/- 74.2 microns2; the dendrites arborize within strata 3, 4, and 5 of the IPL. The frequency of each cell class is inversely proportional to the size of the dendritic field. Thus, class 7 cells are the most frequent; class 1 cells are the least frequent. Furthermore, each of these 7 classes of ganglion cells has representative cells located in the inner nuclear layer.(ABSTRACT TRUNCATED AT 400 WORDS)

Retina of the tadpole and frog: delayed dendritic development in a subpopulation of ganglion cells coincident with metamorphosis.

In this study, the morphology of tadpole retinal ganglion cells was compared to that of frogs to determine if changes in dendritic structure occur during metamorphosis. Ganglion cells were analyzed in the tadpole and frog after backfilling with horseradish peroxidase. Representative ganglion cells are present in the tadpole retina, which directly correspond to each of the 7 cell classes found in the frog. However, cells in 3 of these classes (1, 3, and 7) exist in morphologically immature states in retinas from tadpole stages St. XIV-XIX. New dendritic branches appear and the dendritic arbors of these ganglion cells expand during metamorphosis. We propose that the increased dendritic arborization may be followed by new synaptic contacts onto these cells, which contributes to the emergence of new physiological receptive field properties in the frog.

Localization of hydroxyindole-O-methyltransferase-like immunoreactivity in photoreceptors and cone bipolar cells in the human retina: a light and electron microscope study.

The localization of the melatonin-synthesizing enzyme hydroxyindole-O-methyltransferase (HIOMT) was examined by light and electron microscopic immunocytochemistry in the human retina. HIOMT-like immunoreactivity was observed in the photoreceptor layers and the inner nuclear layer (INL). The immunoreactive cells in the INL were more numerous in the central retina than in the peripheral retina and sent processes to both the outer plexiform and inner plexiform layers. The HIOMT immunoreactivity in the inner plexiform layer (IPL) appeared as punctate terminals in the proximal and distal one-thirds of that layer. At the ultrastructural level, HIOMT-like immunoreactivity was localized to the cytoplasm of rod and cone photoreceptors and to a population of cone bipolar cells. HIOMT-immunoreactive bipolar cell dendrites were observed to make both invaginating and flat synaptic contacts with cone pedicles. No immunoreactive invaginating contacts in rod spherules were observed. HIOMT immunoreactivity was observed in the bipolar cell cytoplasm in the INL, and in the bipolar synaptic terminals in the IPL. These terminals contained synaptic ribbons, which formed synaptic contacts with unlabeled cells in the IPL. HIOMT radioenzymatic assays confirmed the presence of HIOMT in the human retina. Average HIOMT activity of eight donors was determined to be 15.0 pmol/mg protein/hour +/- 7.2 S.D. The ultrastructural localization of HIOMT observed in this study, combined with reports from other laboratories, suggests that the cytoplasm of the photoreceptors and a population of cone bipolar cells may be the sites of melatonin synthesis in the human retina.

Serotoninergic neurons in the retina of Xenopus laevis: selective staining, identification, development, and content.

Uptake of 3H-serotonin followed by autoradiography, and uptake of the serotonin analog 5,7-dihydroxytryptamine (5,7-DHT), with subsequent staining, were each used to define a unique set of neurons in the retina of the African clawed frog, Xenopus laevis. Both techniques demonstrated the same population of neurons, on the basis of perikaryal size, shape, and position within the retina. Two classes of amacrine cells accumulated 5,7-DHT at the proximal (vitread) margin of the inner nuclear layer; the two classes were distinguished by the size of their perikarya. Two similar populations of cells, observed in the ganglion cell layer with lower frequency, may represent "displaced" counterparts of these two amacrine cell types. A class of bipolar cells whose perikarya were located in middle-to-distal regions of the inner nuclear layer also accumulated 5,7-DHT and 3H-serotonin. Processes of these cells contributed to a dense plexus of fine fibers that appeared evenly distributed throughout the inner plexiform layer. 3H-Serotonin-accumulating cells first appeared in the developing retina at stage 35/36, a time immediately after retinal stratification but before elaboration of either plexiform layer. Electron microscopic analysis permitted an identification of 3H-serotonin-accumulating terminals in the inner plexiform layer. Serotonin-labeled terminals containing conventional contacts, suggestive of amacrine cells, were presynaptic to unidentified processes and postsynaptic to bipolar cells. Labeled terminals containing ribbon contacts, indicative of bipolar cells, were postsynaptic to amacrine cells. The amount of serotonin contained in isolated retinas was 15 pmol/mg protein as measured by HPLC with electrochemical detection. We attempted to stimulate the release of accumulated 3H-serotonin from mature retinas by increasing the K+-concentration in the bathing medium. Although preloaded glycine is readily released from 14C-glycine-accumulating neurons, from the same retinas there was no calcium-dependent, K+-stimulated release of 3H-serotonin. This finding suggests that serotonin and glycine are processed differently by retinal neurons, the consequence of which results in differing responses to 40 mM K+.

Interphotoreceptor matrix in the human retina: cone-like domains surround a small population of rod photoreceptors.

The interphotoreceptor matrix (IPM) in mammalian retinas is subdivided into rod and cone specific compartments: peanut agglutinin (PNA) binding glycoconjugates are associated with cones, whereas wheat germ agglutinin (WGA) binding glycoconjugates are associated with rods. To establish the identity of a photoreceptor cell type in the human retina with rod dimensions but with a matrix domain which stains with PNA, double label studies, using PNA-ferritin to decorate the extracellular domains and immunocytochemical techniques using a rod specific anti-opsin antibody were conducted. The PNA-binding domains were observed in the cone-associated IPM as well as in the IPM surrounding a small population of rod-shaped photoreceptors. The outer segments of these rod-shaped photoreceptors showed intense labeling with a rod specific anti-opsin antibody as did all other rods which were free of PNA-labeling. A quantitative analysis of all retinal quadrants indicates that this novel rod represents approximately 0.3% of the total rod population in the human retina.

Glycinergic neurons in the human retina.

Neurotransmitter-specific properties of glycinergic neurons in the human retina were studied using 11 pairs of eyes from donors ranging from 2 1/2 to 54 years in age. A mean endogenous level of 10.3 nmoles glycine per mg protein was measured by amino acid analysis in retinas isolated within 1 hour postmortem. When retinas were incubated with 3H-glycine (2 microM) and processed for autoradiography, label was found associated with neurons whose somata reside within the inner nuclear layer. Some heavily labeled neurons located at the vitread border of the inner nuclear layer were identified as amacrine cells based on ultrastructural verification of the conventional synaptic contacts made by their processes in distal regions of the inner plexiform layer. In proximal regions of the inner plexiform layer, dendrites of glycine-accumulating amacrine cells were postsynaptic to both ribbon and conventional synaptic contacts, suggesting input from bipolar and other, nonglycinergic amacrine cells. Their density (30 +/- 11 S.D. cells/mm linear retinal expanse) tended to be greater toward the central fundus. A second population of lightly labeled, probable bipolar cells was present in the middle of the inner nuclear layer; the density of this second set of glycine-accumulating cells approximated that of the heavily labeled population from the fovea, centrally, to the ora serrata, peripherally. Release of either accumulated or endogenous glycine was elicited by K+-depolarization in a Ca2+-dependent manner. Tissue fragments exposed for 6 minutes to normal medium, 40 mM K+-substituted medium, or K+-substituted medium with Co2+, release endogenous glycine into each bathing solution in average amounts of 0.6, 2.6, and 0.7 nmoles per mg protein, respectively. Together these data strongly implicate glycine as a neurotransmitter in the human retina.

Dopaminergic neurons in the human retina.

The utilization of dopamine in the adult human retina was examined by using high-affinity uptake, localization, synthesis, and release as neurotransmitter-specific physiological probes. Autoradiographic and histochemical studies have shown that dopamine-accumulating and dopamine-containing cells of the human retina belong to a population of neurons whose somata are located in the proximal regional of the inner nuclear layer. Some of these are amacrine cells which are pre- and postsynaptic to other amacrine cells exclusively in the inner plexiform layer. However, evidence is presented which indicates the existence of interplexiform dopaminergic neurons which send processes to both plexiform layers of the retina. These neurons contain a high concentration of dopamine, take up 3H-dopamine by a hig-affinity mechanism, and release endogenous or accumulated dopamine by a Ca2+-dependent mechanism upon depolarization with high extracellular K+. An endogeneous level of about 20 pmoles dopamine per mg protein was measured in freshly isolated retina using high-pressure liquid chromatography with electrochemical detection. These results demonstrate that mechanisms for dopaminergic neurotransmission are present in the human retina.

The emergence, localization, and maturation of neurotransmitter systems during development of the retina in Xenopus laevis: II. Glycine.

The high-affinity uptake and release of glycine was studied in retinas of Xenopus laevis. In the toad and tadpole retina, 3H-glycine was accumulated by a population of cells located predominantly in the inner nuclear layer. When retinas preloaded with 3H-glycine were subjected to high K+-concentrations, these retinas released large amounts of 3H-glycine by a Ca++-dependent mechanism. The appearance and maturation of these putative glycinergic properties was followed during retinal development. Our results indicate that the high-affinity uptake of glycine first appears around stage 33/34 whereas K+-stimulated glycine release cannot be detected until stage 42.

The emergence, localization, and maturation of neurotransmitter systems during development of the retina in Xenopus laevis. III. Dopamine.

The uptake, synthesis, and release of dopamine was studied in retinas of Xenopus laevis. In the tadpole and adult retina, 3H-dopamine is accumulated by cells located in the inner nuclear layer. Retinas preloaded with 3H-dopamine release this compound in response to high K+ concentrations in the medium. This release is probably Ca++-dependent as it is inhibited by Co++ in the medium. Adult retinas are also capable of synthesizing 3H-dopamine from 3H-tyrosine. The appearance and maturation of these dopaminergic properties were followed during retinal development. Our data indicate that synthesis of dopamine can first be detected as early as stage 35/36 whereas uptake of dopamine first occurs at stage 43. K+-stimulated release of preloaded 3H-dopamine from putative dopaminergic neurons is, however, not evident until stage 46. These results show that similar to the development of GABA-ergic and glycinergic properties, the uptake, synthesis, and release mechanisms for dopamine emerge at different stages during retinal differentiation in Xenopus Laevis.

The emergence, localization and maturation of neurotransmitter systems during development of the retina in Xenopus laevis. I. Gamma aminobutyric acid.

The high-affinity uptake, biosynthesis and release of GABA have been studied in the retina of Xenopus laevis. In the mature retina, [3H]-GABA is accumulated predominantly by horizontal cells. A second population of cells located in the inner nuclear layer (possibly a type of amacrine cell) also showed a specific GABA uptake. In addition, this retina contains significant activities of L-glutamic acid decarboxylase and also releases [3H]-GABA in response to increasing K+ concentrations in the medium. We have followed the appearance and maturation of these GABA-ergic properties during embryonic development of this retina. Our results indicate that these properties emerge in a precise temporal pattern during retinal differentiation: the specific neuronal uptake of GABA precedes GABA synthesis which is followed by K+-stimulated GABA release.