In in-vitro preparations of both macaque and human retina, intracellular injections of Neurobiotin and horseradish peroxidase were used to characterize the morphology, depth of stratification, and mosaic organization of a type of bistratified ganglion cell. This cell type, here called the small bistratified cell, has been shown to project to the parvocellular layers of the dorsal lateral geniculate nucleus (Rodieck, 1991) and is therefore likely to show color-opponent response properties.

In both human and macaque, the two dendritic tiers of the bistratified cell are narrowly stratified close to the inner and outer borders of the inner plexiform layer. The inner tier is larger in diameter and more densely branched than the outer tier and gives rise to distinct spine-like branchlets bearing large, often lobulated heads. By contrast the smaller, outer tier is sparsely branched and relatively spine-free.

In human retina, the small bistratified cells range in dendritic field diameter from ∼50 µm in central retina to ∼400 µm in the far periphery. The human small bistratified cells are about 20% larger in dendritic-field diameter than their counterparts in the macaque. However, when the difference in retinal magnification between human and macaque is taken into account, the small bistratified cells are similar in size in both species. In macaque, the small bistratified cell has a dendritic-field size that is ~10% larger than that of the magnocellular-projecting parasol ganglion cell. Human small bistratified ganglion cells tend to have smaller dendritic-field diameters than parasol cells. This is because parasol ganglion cells are larger in human than in macaque retina (Dacey & Petersen, 1992).

In macaque retina, intracellular injections of Neurobiotin revealed heterotypic tracer coupling to a distinct mosaic of amacrine cells and probable homotypic coupling to an array of neighboring ganglion cells around the perimeter of the injected cell's dendritic tree. The amacrine cell mosaic had a density of 1700 cells/mm2 in peripheral retina. Individual amacrines had small, densely branched and bistratified dendritic fields. From the homotypic coupling, it was possible to estimate for the small bistratified cell a coverage factor of ~1.8, and a density of ~1% of the total ganglion cells in central retina, increasing to ~6–10% in the retinal periphery.

The estimated density, dendritic-field size, and depth of stratification all suggest that the small bistratified ganglion cell type is the morphological counterpart of the common short-wavelength sensitive or ‘blue-ON’ physiological type.

Monoclonal antibodies to human leucocyte antigens, including anti-CD45 and anti-CD68, have been used to describe microglia in flatmounts of normal adult human retina for the first time. Anti-CD45 (the leucocyte common antigen) intensely labeled large numbers of cells in a regular distribution across the retina; anti-CD68 and anti-macrophage antibodies labeled fewer cells with distinctive morphologies, suggesting the presence of subpopulations of microglia in the human retina expressing leucocyte antigens.

The expression of GABA in the human fetal (12–25 weeks of gestation), postnatal (five-month-old), and adult (35-year-old) retinas was investigated by immunohistochemistry. GABA expression was seen as early as 12 weeks in the undifferentiated cells of the inner neuroblast zone; a few optic nerve fiber layer axons were clearly labeled, suggesting that some of the stained cell bodies were prospective ganglion cells, others could be displaced amacrine cells. From 16–17 to 24–25 weeks, intense labeling was found in the amacrine, displaced amacrine, and some ganglion cells. During this time period, horizontal cells (identified by calbindin immunohistochemistry), undergoing migration (periphery) and differentiation (center), expressed GABA prominently. In the postnatal retina, some horizontal cells were moderately labeled, but very weakly in a few cells, in the adult. The Müller cells developed immunoreactivity first weakly at 12 weeks and then moderately from 16–17 weeks onward. The staining was also evident in the postnatal and adult retinas, showing labeled processes of these glial cells. Virtually no axons in the adult optic nerve and nerve fiber layer were stained; the staining was restricted to a few, large ganglion cells and displaced amacrine cells. Some amacrines were also labeled. The possibility that GABA might play a role in horizontal cell differentiation and maturation is highlighted. Other evidences suggest that GABA might play a role in metabolism during retinal development.

Video-enhanced imaging of retinal wholemounts reveals an abrupt change in the composition of the photoreceptor mosaic at the edge of the human retina. Cone densities rise threefold and rod densities fall tenfold in a 1-mm-wide peripheral band. Antibodies directed against cones confirm the identification of the major subtypes of photoreceptors within this peripheral band. The cone-enriched rim is most highly developed along the nasal retinal margin, an area where the extreme lateral periphery of the visual field is imaged. This rim of cones may function as part of a rapid-acting alert mechanism under conditions of moderate and bright illumination.

The major neurons and neural circuits that are involved in the transmission of color signals through the human retina to produce the color and spatially opponent P cell or midget ganglion cell responses are described. The older findings of single cone to midget bipolar connectivity is reviewed, and the single midget bipolar cell to midget ganglion cell connectivity as revealed by a recent serial section electron microscope study is described in detail. Our present knowledge concerning the discrimination of the blue-cone subtype from the other longer wavelength cones in the human at the outer plexiform layer is summarized, and our most recent findings concerning horizontal cell connectivity to the different spectral types of cones are discussed. Finally, a hypothetical pathway is proposed for color-opponent surrounds of midget ganglion cells using both horizontal cells at the outer plexiform layer and amacrine cell pathways at the inner plexiform layer.

Retinitis pigmentosa (RP) is an inherited disease that causes primary degeneration of rod photoreceptors in the retina. Although the causal gene (e.g. rhodopsin) is thought to be expressed in all rods across the retina, the degeneration is typically nonuniform, with rods in the far periphery surviving significantly longer than those in the midperiphery and macula. Basic fibroblast growth factor (bFGF) is a putative survival factor for photoreceptors, and the characteristic regional pattern of rod cell survival in RP suggested that bFGF might be distributed nonuniformly in the human retina. We performed double-label immunocytochemistry on 15 normal human retinas, using anti-bFGF and other antibody markers for retinal neurons and glia. Immunoreactivity for bFGF was consistently absent from cones but was present in rods, populations of cone bipolar and amacrine cells, Müller glial cells, and astrocytes. In the macula, the percentage of bFGF-reactive rods was very low (~0.5%) but it increased in a central to peripheral gradient, accounting for up to ~88% of the rods in the far periphery. These findings suggest that a central to peripheral gradient of rod bFGF is present in normal human retina and may influence the pattern of photoreceptor degeneration in RP. The absence of bFGF in cones and the low number of bFGF-positive rods in the macula may correlate with the vulnerability of these cells in RP, age-related macular degeneration, and other retinal diseases.

To study the detailed morphology of human retinal ganglion cells, we used intracellular injection of horseradish peroxidase and Neurobiotin to label over 1000 cells in an in vitro, wholemount preparation of the human retina. This study reports on the morphology of 119 wide-field bistratified and 42 diffuse ganglion cells. Cells were analyzed quantitatively on the basis of dendritic-field size, soma size, and the extent of dendritic branching. Bistratified cells were similar in dendritic-field diameter (mean ± s.d. = 682 ± 130 μm) and soma diameter (mean ± s.d. = 18 ± 3.3 μm) but showed a broad distribution in the extent of dendritic branching (mean ± s.d. branch point number = 67 ± 32; range = 15–167). Differences in the extent of branching and in dendritic morphology and the pattern of branching suggest that the human retina may contain at least three types of wide-field bistratified cells. Diffuse ganglion cells comprised a largely homogeneous group whose dendrites ramified throughout the inner plexiform layer. The diffuse cells had similar dendritic-field diameters (mean ± s.d. = 486 ± 113 μm), soma diameters (mean ± s.d. = 16 ± 2.3 μm), and branch points numbers (mean ± s.d. = 92 ± 32). The majority had densely branched dendritic trees and thin, very spiny dendrites with many short, fine, twig-like thorny processes. Five of the diffuse cells had much more sparsely branched dendritic trees (<50 branch points) and less spiny dendrites, suggesting that there are possibly two types of diffuse ganglion cells in human retina. Although the presence of a diversity of large bistratified and diffuse ganglion cells has been observed in a variety of mammalian retinas, little is known about the number of cell types, their physiological properties, or their central projections. Some of the human wide-field bistratified cells in the present study, however, show morphological similarities to monkey large bistratified cells that are known to project to the superior colliculus.

To determine the number of wide-field, monostratified ganglion cell classes present in the human retina, we analyzed a large sample of ganglion cells by intracellular staining in an in vitro, whole-mount preparation of the retina. Over 1000 cells were labeled by horseradish peroxidase or Neurobiotin; some 200 cells had wide dendritic trees narrowly or broadly stratified within either the inner (ON) or outer (OFF) portion of the inner plexiform layer. Based on dendritic-field size and the pattern and extent of dendritic branching, we have distinguished six wide-field cell groups. The giant very sparse ganglion cells included both inner and outer stratifying cells and were unique both for their extremely large dendritic field (mean diameter = 1077 μm) and extremely sparsely branched dendrites. Four of the cell groups had similarly large dendritic fields, ranging in mean diameter from 737 to 791 μm, but differed in the pattern and extent of dendritic branching, with the number of dendritic branch points ranging from a mean of 33 to 129. Of these four groups, the large very sparse group and the large dense group included both inner and outer stratifying cells, while the large sparse and large moderate groups consisted of inner stratifying cells only. The thorny monostratified ganglion cells were distinct from the other cells in having medium size dendritic fields (mean diameter = 517 μm) and moderately branched, inner stratifying dendritic trees with many thin, spiny, twig-like branchlets. All six groups had medium-size cell bodies, with mean soma diameters ranging from 17 to 21 μm. Though the physiological properties and central projections of human wide-field, monostratified ganglion cells are not known, some of the cells resemble macaque ganglion cells known to project to the lateral geniculate nucleus, the pretectum, or the superior colliculus.

Ganglion cells with intraretinal axon collaterals have been described in monkey (Usai et al., 1991), cat (Dacey, 1985), and turtle (Gardiner & Dacey, 1988) retina. Using intracellular injection of horseradish peroxidase and Neurobiotin in in vitro whole-mount preparations of human retina, we filled over 1000 ganglion cells, 19 of which had intraretinal axon collaterals and wide-field, spiny dendritic trees stratifying in the inner half of the inner plexiform layer. The axons were smooth and thin (∼2 μm) and gave off thin (<1 μm), bouton-studded terminal collaterals that extended vertically to terminate in the outer half of the inner plexiform layer. Terminal collaterals were typically 3–300 μm in length, though sometimes as long as 700 μm, and were present in clusters, or as single branched or unbranched varicose processes with round or somewhat flattened lobular terminal boutons 1–2 μm in diameter. Some cells had a single axon whereas other cells had a primary axon that gave rise to 2–4 axon branches. Axons were located either in the optic fiber layer or just beneath it in the ganglion cell layer, or near the border of the ganglion cell layer and the inner plexiform layer. This study shows that in the human retina, intraretinal axon collaterals are associated with a morphologically distinct ganglion cell type. The synaptic connections and functional role of these cells are not yet known. Since distinct ganglion cell types with intraretinal axon collaterals have also been found in monkey, cat, and turtle, this cell type may be common to all vertebrate retinas.