A subgroup of bipolar cells in human retina is GABA-immunoreactive.

The distribution of GABA in the perifoveal and the near and far peripheral region of human retina was studied with peroxidase anti-peroxidase immunocytochemistry applied on semithin epoxy resin sections. Among the labeled amacrine cells in these regions, four types can be identified: putative diffuse A2 amacrines, stratified semilunar amacrines, interstitial amacrines and small displaced amacrines. GABA-immunoreactive interplexiform cells and ganglion cells also occur. Contrary to previous post-embedding studies, our preparations show that some bipolar cells in the near and far peripheral region are GABA-immunopositive. This indicates that a number of bipolar cells in human retina does have an enhanced GABA content.

New aspects of dopaminergic interplexiform cell organization in the goldfish retina.

Dopaminergic interplexiform cells (DA-IPCs) in the goldfish retina have been reexamined by light and electron microscopic immunocytochemistry with antisera against dopamine (DA) or tyrosine hydroxylase (TH). Successful immunostaining with a specific anti-DA antiserum offers further direct support for DA-IPCs. Anti-DA immunocytochemistry in combination with [3H]-DA autoradiography shows 92% colocalization of the two markers, indicating that [3H]-DA autoradiography is a reliable technique for identification of DA-IPCs. Incubations with anti-TH antiserum show that immunoreactive DA-IPCs have a homogeneous distribution, with an average frequency of 71 +/- 8 cells/mm2 in retinas of 14-15 cm long goldfish. Their arrangement is distinctly nonrandom. Electron microscopy of TH-immunoreactive cell processes confirms that horizontal cell axons synapse onto DA-IPCs and adds the following junctional arrangements to the circuit diagram of the DA-IPC: 1) adjacent serial synapses between DA-IPCs, external horizontal cells, and putative glycinergic interplexiform cells, 2) junctional appositions between DA-IPCs and photoreceptor cells, 3) junctional appositions between neighbouring DA-IPCs, and 4) the "gap junctional complex," typically consisting of a DA-IPC process juxtaposed with a gap junction between horizontal cell axons. The gap junction is flanked by clusters of small, round vesicles and groups of electron-dense structures resembling intermediate filaments. These morphological results support the functional involvement of DA-IPCs in adaptive retinomotor movements and in horizontal cell gap junction modulation and/or dynamics. They also suggest particular interaction between the dopaminergic and the glycinergic IPC system in the outer plexiform layer of goldfish retina.

Three types of GABA-immunoreactive cone horizontal cells in teleost retina.

Peroxidase-anti-peroxidase immunocytochemistry, applied on serial semithin epoxy resin sections, was used to examine the localization of endogenous GABA in horizontal cells in the retina of a marine teleost, the dragonet (Callionymus lyra L.). The immunostaining shows that not only the external H1 cone horizontal cells label with antibodies against GABA, but also the H2 and H3 cone horizontal cells in the inner nuclear layer. The distribution of the H1 cells corresponds to that of the single cones. They are square-patterned and in the dorsal retina their density equals 20,000 cells/mm2. The estimated density of the immunostained H2 and the H3 cells in the dorsal retina is 9500 and 1300 cells/mm2, respectively. The H2 and H3 cells are not geometrically arranged, but nearest-neighbor analysis shows that these horizontal cell types do have a very regular disposition. We suggest that GABA is the likely neurotransmitter substance used by all cone horizontal cell types in teleost retina.

Patterns of glutamate-like immunoreactive bipolar cell axons in the retina of the marine teleost, the dragonet.

Oblique 1 microns-sections through the dorsal inner plexiform layer of the light-adapted dragonet retina were processed for postembedding, silver-enhanced immunogold labeling after incubation with a glutamate-specific antiserum. Light microscopy showed strongly immunolabeled boutons grouped into distinct square patterns. These patterns were compared with the successive grids of bipolar axonal boutons revealed by electron microscope analysis of serial, oblique sections through the entire dorsal inner plexiform layer. With one exception, all types of patterned bipolar synaptic boutons could be clearly identified in the immunoreactive staining pattern. The elevated levels of endogenous glutamate in most bipolar synaptic boutons suggest that the large majority of bipolar cell types use glutamate as their neurotransmitter. However, some bipolar synaptic boutons displaying low levels of glutamate indicate that a small number of bipolar cells may contain another neuroactive substance.

Patterns of glutamate immunoreactivity in the goldfish retina.

Postembedding silver-intensified immunogold procedures reveal high levels of glutamate immunoreactivity in "vertical" elements of the goldfish retina: (1) Red-sensitive and green-sensitive cones display strong glutamate immunoreactivity, especially in their synaptic terminals, but blue-sensitive cones are poorly immunoreactive. (2) All type Mb (on-center) and Ma (off-center) mixed rod-cone bipolar cells and all identifiable cone bipolar cells are highly glutamate immunoreactive. We find no evidence for bipolar cells that lack glutamate immunoreactivity. (3) The majority of the somas in the ganglion cell layer and certain large cells of the amacrine cell layer resembling displaced ganglion cells are strongly glutamate immunoreactive. (4) Despite their high affinity symport of acidic amino acids, the endogenous levels of glutamate in Müller's cells are among the lowest in the retina. (5) GABAergic neurons possess intermediate levels of glutamate immunoreactivity. Quantitative immunocytochemistry coupled with digital image analysis allows estimates of intracellular glutamate levels. Photoreceptors and bipolar and ganglion cells contain from 1 to 10 mM glutamate. The bipolar and ganglion cell populations maintain high intracellular glutamate concentrations, averaging about 5 mM, whereas red-sensitive and green-sensitive cones apparently maintain lower levels. Importantly, photoreceptor glutamate levels are extremely volatile, and in vitro maintenance is required to preserve cone glutamate immunoreactivity in the goldfish. GABAergic horizontal and amacrine cells contain about 0.3-0.7 mM glutamate, which matches the values predicted from the Km of glutamic acid decarboxylase. Müller's cells and non-GABAergic amacrine cells contain less than 0.1 mM glutamate. Though Müller's cells are known to possess potent glutamate symport, they clearly possess equally potent mechanisms for maintaining low intracellular glutamate concentrations.

Glutamate-like immunoreactivity in the retina of a marine teleost, the dragonet.

The localisation of endogenous glutamate in the dragonet retina was investigated by light microscopic postembedding silver-enhanced immunogold labeling after incubation with an anti-glutamate antiserum. Rod and cone inner segments and synaptic terminals, as well as the inner plexiform layer, are moderately labeled. Bipolar cells and ganglion cell bodies show strong labeling. In the dorsal inner plexiform layer, the levels with square-patterned bipolar synaptic boutons can be identified by their prominent glutamate-immunoreactivity. These results support the idea that the majority of the neurons that constitute the direct, centripetal pathways through the retina use glutamate as their neurotransmitter.

Displaced small amacrine cells in the retina of the marine teleost Callionymus lyra L.

The displaced small amacrine cells (DSA cells) in the dorsal pure cone part of the retina of the marine teleost Callionymus lyra have been analysed in a combined light and electron microcopical study. These unistratified cells have their dendritic arborization at 70% of the depth of the inner plexiform layer (P5 level). The DSA cells constitute a dense population and have variable dendritic field sizes. The bipolar input occurs in the P5,1 and the P5,2 pattern layer. The short, central DSA dendrites make ribbon synapses with midget mixed di-cone bipolar cells and with two types of pure cone bipolar cells. The amacrine input and output occurs in the fibrous layer that separates both pattern layers. The dendritic arborization is most extensive and the dendrites of neighbouring DSA cells are interconnected. The thick, central DSA dendrites are presynaptic to adjacent DSA cells and possibly to large bistratified and diffuse ganglion cells. The fine, peripheral DSA dendrites receive input from neighbouring DSA cells and probably from large uni- and bistratified and diffuse amacrine cells. A matching population of regularly placed small amacrine cells (RSA cells) has been observed. Their unistratified dendritic arborization is situated at 20% of the depth of the inner plexiform layer. The synaptic relations of RSA cells have not yet been completely analysed in detail. However, results up till now indicate that they most probably receive input from two bipolar cell types, one of which may be a pure cone type. In addition, the large bistratified amacrine and ganglion cells may be synaptically connected to the RSA cells as well as to the DSA cells.

Synaptic contacts between bipolar and photoreceptor cells in the retina of Callionymus lyra L.

A combined light and electron microscopic study of Golgi-impregnated retinas of the marine teleost Callionymus lyra L. revealed mixed bipolar cells (M types) contacting rods and cones and pure cone bipolar cells (C types). Five types of mixed bipolar cells can be differentiated on the basis of their synaptic contacts. Two out of the five mixed bipolar cell types contact double cones, single cones, and rods (mixed, dark, pale, single [Mdps and midget-Mdps]). Their endbuds make narrow cleft junctions, with each type of photoreceptor, and in addition, two endbuds end centrally in the synaptic ribbon complexes of the dark and pale double-cone pedicles. Three types of mixed bipolar cells contact only double cones and rods. The endbuds of one type (mixed, dark, pale, ribbon [Mdpr]) end centrally in the synaptic ribbon complexes of the dark and pale double-cone pedicles as well as of the rod spherules. The endbuds of two types (Mdp and midget-Mdp) make wide cleft junctions in dark and pale double-cone pedicles and in rod spherules. All pure cone bipolar cell types contact cones exclusively with narrow cleft junctions. Four types are seen: a type that contacts predominantly pale double-cone pedicles but also a few dark double-cone pedicles (Cp), a type that is connected with dark and pale double-cone pedicles in about equal numbers (Cdp), a type that makes synaptic contacts with pale double-cone pedicles and single-cone pedicles (Cps), and a type that is connected with both types of double cones and to single-cone pedicles (Cdps). A resemblance between the ultrastructural features of mixed bipolar cell synapses in Callionymus and in Carassius auratus is noted.

Stratification and square pattern arrangements in the dorsal inner plexiform layer in the retina of Callionymus lyra L.

The dorsal inner plexiform layer in the retina of Callionymus lyra consists of successive fibrous F layers and geometrically organized P layers. In the F layers no pattern arrangement of the bipolar synaptic buttons was observed. In the P layers, on the contrary, the bipolar synaptic buttons are grouped by three distinct square patterns. The alpha pattern measures 5 micron and corresponds to the square pattern of the photoreceptor cells. The beta and the gamma pattern measure 7 micron. One of both patterns corresponds to the single cone arrangement, while the other coincides with the crossings of the rows of double cone pairs. The alternation of F and P layers results in the stratified appearance of the inner plexiform layer.

Interbipolar contacts in the dorsal inner plexiform layer in the retina of Callionymus lyra L.

Several types of direct contacts have been observed between the square pattern-arranged bipolar synaptic buttons in the P layers in the dorsal inner plexiform layer of the Callionymus retina. These contacts are not randomly occurring interconnections, but they are systematically formed throughout the whole of the dorsal retinal zone and they may be the site of direct synaptic interactions between similar as well as dissimilar bipolar axons. Reciprocal ribbon synapses have been found in the P2,2 layer. Morphologically mixed synapses occur in the P5,1 layer. These contact zones show morphologic characteristics of both chemical and electronic synapses. Morphologically mixed synaptic complexes, representing a triple coupling between two adjacent similar bipolar buttons and an amacrine cell process, have been observed in the P1,1, the P2,1 and the P5,2 layer. The morphology of the interbipolar contacts in the P3,2 layer does not clearly point to a synaptic function. Our results suggest a high degree of functional stratification in the inner plexiform layer.

Synaptic contacts of the horizontal cells in the retina of the marine teleost, Callionymus lyra L.

The horizontal cell system in the retina of the fish Callionymus lyra L. was investigated light microscopically and electron microscopically. One type of rod horizontal cells, which are only found in the mixed ventral part, exclusively contacts spherules with lateral endknobs in the triads. Three types of cone horizontal cells occur. The first type, c-H1, contacts pedicles of pale and dark double cones and single cones. The processes always have a lateral position in the triads. This type has a constant contact pattern with the photoreceptor cells. The second type, c-H2, selectively contacts pedicles of pale double cones and the endbuds occupy a central position in the triads. In the single cone pedicles in the dendritic field of the c-H2 horizontal cell, the endbuds never seem to reach the triads. The third type, c-H3, only sends processes to pedicles of single cones where the endbuds occupy a central position in the triads. The synaptic connections of the horizontal cells of Callionymus differ from those observed in Nannacara anomala and Carassius auratus. The difference from the results obtained from Carassius is such that the information transfer model proposed for Carassius cannot apply to Callionymus.