John E. Dowling

Organization of the Primate Retina: Electron Microscopy

The retinae of monkey and man have been studied by electron microscopy to identify cell types, their processes and synaptic contacts. In the inner plexiform layer, the morphological characteristics of the three types of cells (bipolar, ganglion and amacrine) are described and seven synaptic relationships are identified. The bipolar terminals contain ribbons at points of synaptic contact, and, at these points, there are typically two postsynaptic processes, one a ganglion cell dendrite, the other an amacrine cell process. This synaptic arrangement is here termed a dyad. The amacrine cell processes themselves make synaptic contacts with ganglion cell dendrites and somata, other amacrine cell processes, and, most frequently, with the bipolar cell terminals. Often, the amacrine-bipolar contact is adjacent to a bipolar-amacrine junction, forming a reciprocal synaptic arrangement between the bipolar and the amacrine. In the more peripheral retina, large bipolar cell terminals (probably of rod bipolars) are occasionally observed adjacent to the perikarya of the ganglion cells. At these junctions, areas of fusion between the plasma membranes are seen, suggesting that such axosomatic junctions could be electrical. In the outer plexiform layer, synapses have been identified only in the receptor cell bases where receptor cells contact bipolar and horizontal cell processes. Synaptic contacts of the horizontal cells have not been clearly identified, but their strategic terminations in the receptor cell ending are described and interpreted as possibly synaptic. A model of the retina, based on the described anatomy, is presented and correlated with ganglion cell physiology.

The interplexiform cell system - I. Synapses of the dopaminergic neurons of the goldfish retina

Interplexiform cells are a class of retinal neuron that extends processes widely in both plexiform layers. In goldfish they contain dopamine and readily take up certain biogenic amines. Two of these amines, 6-hydroxyopamine (6-HDA) and 5, 6-dihydroxytryptamine (5,6-DHT), induce fine structural changes in the neurons that accumulate them, allowing the processes of the cells to be recognized by electron microscopy. Typically, the synaptic vesicles within the processes show electron-dense cores. The terminal cytoplasm may also show increased density, as may the cellular and cytoplasmic membranes, presumably an indication of degenerative changes induced by the drugs. 5, 6-DHT gives more readily observable changes than 6-HDA but labels both dopaminergic and indoleamine-accumulating neurons. The terminals of the indoleamine-accumulating terminals were therefore removed by intraocular injections of 5, 7-dihydroxytryptamine (5, 7-DHT) prior to the labelling with 5, 6-DHT. This procedure allowed an analysis of the dopaminergic terminals without interference by the terminals of the indoleamine-accumulating cells. The dopaminergic neurons were found to make synapses of the conventional type. In the outer plexiform layer they contacted both external horizontal cells and bipolar cell dendrites, but not hotoreceptor terminals or intermediate (rod) horizontal cells. No synapses onto the dopaminergic processes were found in the outer plexiform layer despite an extensive search. In the inner plexiform layer the dopaminergic processes were observed to be both pre- and postsynaptic to amacrine cells and their processes. No synaptic contacts between dopaminergic processes and bipolar cell terminals or ganglion cell dendrites were seen. We conclude that the dopaminergic interplexiform cells provide a centrifugal pathway for information flow in the retina from inner to outer plexiform layer.

Synaptic Organization of the Frog Retina: An Electron Microscopic Analysis Comparing the Retinas of Frogs and Primates

The synaptic contacts in the inner and outer plexiform layers of the frog retina have been identified and studied by electron microscopy. In the inner plexiform layer, two types of synaptic contact were recognized. One type, believed to be the synaptic contact of the bipolar terminals, is characterized by a synaptic ribbon in the presynaptic cytoplasm. At such ribbon contacts, there are ordinarily two postsynaptic elements, both of which usually contain numerous synaptic vesicles and appear morphologically identical. The second type of synaptic contact in the inner plexiform layer has a more conventional morphology and is observed very much more frequently than are the ribbon contacts. It is characterized by a dense aggregation of synaptic vesicles clustered close to the presynaptic membrane and is thought to be the synaptic contact of the amacrine processes. The conventional synapses are presynaptic to ribbon-containing processes, ganglion cell dendrites, and other amacrine cell processes. Reciprocal contacts between processes making ribbon synapses, and processes making conventional synapses are often observed. Serial synapses between morphologically identical processes, presumably amacrine processes, are frequently seen; and up to four synapses in series between five adjacent processes have been observed. These findings suggest that in the inner plexiform layer of the frog: (1) bipolar terminals synapse primarily with amacrine processes; (2) amacrine processes synapse extensively with the processes of other amacrine cells; and (3) ganglion cells are driven primarily by the amacrine cells. In the outer plexiform layer, processes penetrate into invaginations in the bases of the receptor terminals and lie in close proximity to the synaptic ribbons of the terminals, where the processes presumably receive synaptic input from the receptors. Elsewhere in the outer plexiform layer, knob-like processes, probably from horizontal cells, make conventional synaptic contacts with other horizontal cell processes and probably with bipolar dendrites.

Early retinal development in the zebrafish, Danio rerio: Light and electron microscopic analyses

The morphological differentiation of the zebrafish retina was analyzed by using light (LM) and transmission electron (TEM) microscopy between the time of initial ganglion cell differentiation (≈32 hours postfertilization; hpf) and shortly after the point when the retina appears functional (≈74 hpf), i.e., when all major cell types and basic synaptic connections are in place. The results show that the inner retinal neurons, like the photoreceptor and ganglion cells, differentiate first within the ventronasal region, and differentiation subsequently spreads asymmetrically into the nasal and dorsal regions before reaching the ventrotemporal retina. In addition, we show that the attenuation of the optic stalk occurs in parallel with ganglion cell differentiation between 32 and 40 hpf. The first conventional synapses appear within the inner plexiform layer simultaneously with the first photoreceptor outer segment discs at 60 hpf; functional ribbon triads arise within photoreceptor synaptic terminals at 65 hpf; and synaptic ribbons occur within bipolar cell axon terminals at the time larvae exhibit their first visual responses (≈70 hpf). Although development is initially more advanced within the ventronasal region between 50 and 60 hpf, development across the retina rapidly equilibrates such that it is relatively comparable within all quadrants of the central retina by 70 hpf. An area within the temporal retina characterized by tightly packed and highly tiered cones emerges with subsequent development. Retinal differentiation in the zebrafish corresponds with that generally described in other vertebrates and can be correlated with the development of visual and electroretinographic responses in the animal. J. Comp. Neurol. 404:515–536, 1999.

Behavioral screening for cocaine sensitivity in mutagenized zebrafish

Understanding the molecular basis of addiction could be greatly aided by using forward genetic manipulation to lengthen the list of candidate genes involved in this complex process. Here, we report that zebrafish exhibit cocaine-induced conditioned place preference. In a pilot screen of 18 F2 generation families of mutagenized fish, we found three with abnormally low responses to cocaine. This behavior was inherited by the F3 generation in a manner that suggests the abnormalities were because of dominant mutations in single genes. Performance profiles in secondary behavioral screens measuring visual dark-adaptation and learning suggest that the defects were the result of mutations in distinct genes that affect dopaminergic signaling in the retina and brain.

The Site of Visual Adaptation

In response to background illumination, the adaptation properties of the b-wave are similar to those observed in the human eye with psychophysical methods. With increasing background luminance the b-wave sensitivity is diminished; except at the lowest background intensity the elevation of the log threshold is linearly related to the increase of background intensity, the relation having a slope of almost 1. The a-wave, however, behaves quite differently. At low background luminances it shows little adaptation. With higher background luminances the awave saturates, and no a-wave potential can be elicited with any stimulus intensity. The L-type S-potentials respond to background light in much the same way as the a-wave does. Thus, the b-wave is the first of the known responses in the visual system to show typical adaptation properties. This suggests that the site of visual adaptation may be in the bi-polarcell layer, the presumed locus of b-wave generation. Recent electron microscopic studies have demonstrated reciprocal synapses between the bipolar terminals and amacrine processes, and it is suggested that such a synaptic arrangement could account for visual adaptation by a mechanism of inhibitory feedback on the bipolar cells.

Synaptic relationships in the plexiform layers of carp retina

SummaryThe synaptic contacts made by carp retinal neurons were studied with electron microscopic techniques. Three kinds of contacts are described: (1) a conventional synapse in which an accumulation of agranular vesicles is found on the presynaptic side along with membrane densification of both pre- and postsynaptic elements; (2) a ribbon synapse in which a presynaptic ribbon surrounded by a halo of agranular vesicles faces two postsynaptic elements; and (3) close apposition of plasma membranes without any vesicle accumulation or membrane densification.In the external plexiform layer, conventional synapses between horizontal cells are described. Horizontal cells possess dense-core vesicles about 1,000 Å in diameter. Membranes of adjacent horizontal cells of the same type (external, intermediate or internal) are found closely apposed over broad regions.In the inner plexiform layer ribbon synapses occur only in bipolar cell terminals. The postsynaptic elements opposite the ribbon may be two amacrine processes or one amacrine process and one ganglion cell dendrite. Amacrine processes make conventional synaptic contacts onto bipolar terminals, other amacrine processes, amacrine cell bodies, ganglion cell dendrites and bodies. Amacrine cells possess dense-core vesicles. Ganglion cells are never presynaptic elements. Serial synapses between amacrine processes and reciprocal synapses between amacrine processes and bipolar terminals are described. The inner plexiform layer contains a large number of myelinated fibers which terminate near the layer of amacrine cells.

The Interplexiform Cell System. II. Effects of Dopamine on Goldfish Retinal Neurones

The effects of atomized solutions of dopamine and certain related compounds have been tested on the intracellularly recorded activity of receptor, horizontal, bipolar and amacrine cells in the goldfish retina. Dopamine depolarizes the cone L-type horizontal cells and reduces the amplitude of light-evoked responses. These effects on L-type horizontal cells are completely abolished by the α-adrenergie blocker, phentolamine, but only partially depressed by the β-blocker, propanolol. L-Dopa, noradrenalin, and serotonin do not have effects on L-type horizontal cells when applied at concentrations similar to those that cause maximal dopamine effects. The results suggest that the effects of dopamine on L-type horizontal cells are specific, and we propose that they mimic the effects of interplexiform cell activity. Dopamine has no effects on rod horizontal cells in goldfish and variable effects on C-type horizontal cells. On bipolar cells, dopamine alters the dark membrane potential, enhances the central response to light, and depresses the surround response. Dopamine also decreases the horizontal cell feedback evident in cone responses. Finally, dopamine strongly depolarizes the transient type of amacrine cells, but it has no significant effect on the sustained type of amacrine cells. Assuming that dopamine is the transmitter of interplexiform cells, we suggest that these neurons regulate lateral inhibitory effects mediated by L-type horizontal cells in the outer plexiform layer and transient amacrine cells in the inner plexiform layer. In addition, it appears as if interplexiform cells have specific effects on bipolar cells and are capable of regulating centre-surround antagonism in these cells. The net effect of interplexiform cell activity is to isolate the bipolars from the influence of the surround.

Synapses of Horizontal Cells in Rabbit and Cat Retinas

Horizontal cells in the retinas of cats and rabbits are morphologically similar; in both species, two types can be distinguished in Golgistained material. Horizontal cells and their processes are readily recognized in electron micrographs, and many of the horizontal cell processes appear to make synaptic contacts with dendrites and somata of bipolar cells, and probably with other horizontal cells. The synapses of the horizontal cell appear similar to chemical synaptic contacts described throughout the nervous system. With the finding of synaptic contacts, it seems clear that retinal horizontal cells should be classified as neurons.

Zebrafish: a model system for the study of eye genetics.

Over the last decade, the use of the zebrafish as a genetic model has moved beyond the proof-of-concept for the analysis of vertebrate embryonic development to demonstrated utility as a mainstream model organism for the understanding of human disease. The initial identification of a variety of zebrafish mutations affecting the eye and retina, and the subsequent cloning of mutated genes have revealed cellular, molecular and physiological processes fundamental to visual system development. With the increasing development of genetic manipulations, sophisticated techniques for phenotypic characterization, behavioral approaches and screening strategies, the identification of novel genes or novel gene functions will have important implications for our understanding of human eye diseases, pathogenesis, and treatment.