Expression analysis of green fluorescent protein in retinal neurons of four transgenic mouse lines

Transgenic mice that express enhanced green fluorescent protein (EGFP) under the control of a cell-specific promoter have been used with great success to identify and label specific cell types of the retina. We studied the expression of EGFP in the retina of mice making use of four transgenic mouse lines. Expression of EGFP driven by the calretinin promoter was found in amacrine, displaced amacrine and ganglion cells. Comparison of the EGFP expression and calretinin immunolabeling showed that many but not all cells appear to be double labeled. Expression of EGFP under the control of the choline acetyltransferase promoter was found in amacrine cells; however, the cells did not correspond to the well known cholinergic (starburst) cells of the mouse retina. The expression of EGFP under the control of the parvalbumin promoter was restricted to amacrine cells of the inner nuclear layer and to cells of the ganglion cell layer (displaced amacrine cells and ganglion cells). Most of the cells were also immunoreactive for parvalbumin, however, differences in labeling intensity were observed. The expression of EGFP driven by the promoter for the 5-HT3 A receptor (5-HTR3A) was restricted to type 5 bipolar cells. In contrast, immunostaining for 5-HTR3A was found in synaptic hot spots in sublamina 1 of the inner plexiform layer and was not related to type 5 bipolar cells. The results show that these transgenic mice are very useful for future electrophysiological studies of specific types of amacrine and bipolar cells that express EGFP and thus permit directed microelectrode targeting under microscopic control.     

Characterization of green fluorescent protein-expressing retinal cells in CD 44-transgenic mice

Sensory information in the retina is transferred from rod and cone photoreceptors to higher visual centers via numerous parallel circuits that sample the photoreceptor mosaic independently. Each circuit consists of a unique combination of ganglion cell, bipolar and amacrine cell types. The morphology and physiological responses of many amacrine cells have been characterized. However, the synaptic connections and retinal circuits in which they participate are only rarely understood. A major problem that has prevented fuller characterization of retinal circuitry is the need for specific cellular markers for the more than 50 inner retinal cell types. One potential strategy for labeling cells is to use transgenic expression of a reporter gene in a specific cell type. In a recent study of cluster of differentiation 44 (CD44)-enhanced green fluorescent protein (EGFP) transgenic mice, we observed that the green fluorescent protein (GFP) was expressed in a population of amacrine and ganglion cells in the inner nuclear layer (INL) and the GCL. To characterize the morphology of the GFP-labeled cells, whole mount preparations of the retina were used for targeted iontophoretic injections of Lucifer Yellow and Neurobiotin. Furthermore, immunocytochemistry was used to characterize the antigenic properties of the cells. We found that many GFP-expressing cells were GABAergic and also expressed calretinin. In addition to the somatic staining, there was a strong GFP(+)-band located about 50-60% depth in the inner plexiform layer (IPL). Double labeling with an antibody to choline acetyltransferase (ChAT) revealed that the GFP-band was located at strata 3 inner retina. The best-labeled GFP-expressing cell type in the INL was a wide-field amacrine cell that ramified in stratum 3. The GFP-expressing cells in the GCL resemble the type B1, or possibly A2 ganglion cells. The CD44-EGFP mice should provide a valuable resource for electrophysiological and connectivity studies of amacrine cells in the mouse retina.     

Substance P-immunoreactive neurons in the retina of two lizards.

The dendritic morphology and retinal distribution of substance P(SP)-immunoreactive neurons was determined in two Australian lizard species Pogona vitticeps and Varanus gouldii, by using immunohistochemistry on retinal wholemounts and sectioned materials. In both species, two classes of SP-immunoreactive neurons were described in the inner nuclear layer (INL) and classified as amacrine cells (types A and B). Type A amacrine cells had large somata and wide-field, bistratified dendrites branching in sublaminas 1 and 5 of the inner plexiform layer (IPL). Their morphology and retinal distribution differed between the two species. Type B amacrine cells in both species had small somata and small-field dendritic branching. A population of SP-immunoreactive neurons with classical ganglion cell morphology were identified in the ganglion cell layer (GCL). Immunostained ganglion cells occurred in larger numbers of Varanus gouldii than in Pogona vitticeps. In both species type B SP cells were the most numerous and were estimated to be about 60,000-70,000. They were distributed non-uniformly with a high density band across the horizontal meridian of the retina, from where the density decreased towards the dorsal and ventral retinal margins. In both species type A amacrine cells occurred in small numbers distributed sparsely in the peripheral retina. The faint immunostaining of SP-immunoreactive neurons in the GCL, did not allow us to reliably determine their numbers and retinal distribution. The functional significance of SP-immunoreactive amacrine and ganglion cells in the lizard retina remains to be determined.     

Characterization of histamine projections and their potential cellular targets in the mouse retina

The vertebrate retina receives histaminergic input from the brain via retinopetal axons that originate from perikarya in the posterior hypothalamus. In the nervous system, histamine acts on three G-protein-coupled receptors, histamine receptor (HR) 1, HR2 and HR3. In order to look for potential cellular targets of histamine in the mouse retina, we have examined the retina for the expression of histamine and the presence of these three receptors. Consistent with studies of retina from other vertebrates, histamine was only found in retinopetal axons, which coursed extensively through the ganglion cell and inner plexiform layers. mRNA for all three receptors was expressed in the mouse retina, and immunohistochemical studies further localized HR1 and HR2. HR1 immunoreactivity was observed on dopaminergic amacrine cells, calretinin-positive ganglion cells and axon bundles in the ganglion cell layer. Furthermore, a distinct group of processes in the inner plexiform layer was labeled, which most likely represents the processes of cholinergic amacrine cells. HR2 immunoreactivity was observed on the processes and cell bodies of the primary glial cells of the mammalian retina, the Müller cells. This distribution of histamine and its receptors is consistent with a brain-derived source of histamine acting on diverse populations of cells in the retina, including both neurons and glia.     

Functional circuitry for peripheral suppression in Mammalian Y-type retinal ganglion cells

A retinal ganglion cell receptive field is made up of an excitatory center and an inhibitory surround. The surround has two components: one driven by horizontal cells at the first synaptic layer and one driven by amacrine cells at the second synaptic layer. Here we characterized how amacrine cells inhibit the center response of on- and off-center Y-type ganglion cells in the in vitro guinea pig retina. A high spatial frequency grating (4-5 cyc/mm), beyond the spatial resolution of horizontal cells, drifted in the ganglion cell receptive field periphery to stimulate amacrine cells. The peripheral grating suppressed the ganglion cell spiking response to a central spot. Suppression of spiking was strongest and observed most consistently in off cells. In intracellular recordings, the grating suppressed the subthreshold membrane potential in two ways: a reduced slope (gain) of the stimulus-response curve by approximately 20-30% and, in off cells, a tonic approximately 1-mV hyperpolarization. In voltage clamp, the grating increased an inhibitory conductance in all cells and simultaneously decreased an excitatory conductance in off cells. To determine whether center response inhibition was presynaptic or postsynaptic (shunting), we measured center response gain under voltage-clamp and current-clamp conditions. Under both conditions, the peripheral grating reduced center response gain similarly. This result suggests that reduced gain in the ganglion cell subthreshold center response reflects inhibition of presynaptic bipolar terminals. Thus amacrine cells suppressed ganglion cell center response gain primarily by inhibiting bipolar cell glutamate release.     

Characterization of receptors for glutamate and GABA in retinal neurons

Glutamate and gamma-aminobutyric acid (GABA) are major excitatory and inhibitory neurotransmitters in the vertebrate retina, "a genuine neural center" (Ramón y Cajal, 1964, Recollections of My Life, C.E. Horne (Translater) MIT Press, Cambridge, MA). Photoreceptors, generating visual signals, and bipolar cells, mediating signal transfer from photoreceptors to ganglion cells, both release glutamate, which induces and/or changes the activity of the post-synaptic neurons (horizontal and bipolar cells for photoreceptors; amacrine and ganglion cells for bipolar cells). Horizontal and amacrine cells, which mediate lateral interaction in the outer and inner retina respectively, use GABA as a principal neurotransmitter. In recent years, glutamate receptors and GABA receptors in the retina have been extensively studied, using multi-disciplinary approaches. In this article some important advances in this field are reviewed, with special reference to retinal information processing. Photoreceptors possess metabotropic glutamate receptors and several subtypes of GABA receptors. Most horizontal cells express AMPA receptors, which may be predominantly assembled from flop slice variants. In addition, these cells also express GABAA and GABAC receptors. Signal transfer from photoreceptors to bipolar cells is rather complicated. Whereas AMPA/KA receptors mediate transmission for OFF type bipolar cells, several subtypes of glutamate receptors, both ionotropic and metabotropic, are involved in the generation of light responses of ON type bipolar cells. GABAA and GABAC receptors with distinct kinetics are differentially expressed on dendrites and axon terminals of both ON and OFF bipolar cells, mediating inhibition from horizontal cells and amacrine cells. Amacrine cells possess ionotropic glutamate receptors, whereas ganglion cells express both ionotropic and metabotropic glutamate receptors. GABAA receptors exist in amacrine and ganglion cells. Physiological data further suggest that GABAC receptors may be involved in the activity of these neurons. Moreover, responses of these retinal third order neurons are modulated by GABAB receptors, and in ganglion cells there exist several subtypes of GABAB receptors. A variety of glutamate receptor and GABA receptor subtypes found in the retina perform distinct functions, thus providing a wide range of neural integration and versatility of synaptic transmission. Perspectives in this research field are presented.     

Evidence for a columnar organization of cones, Müller cells, and neurons in the retina of a cichlid fish

In the retina of many lower vertebrates, the arrangement of cells, in particular of cone photoreceptors, is highly regular. The data presented in this report show that in the retina of a cichlid fish (Astatotilapia burtoni) the regular arrangement is not restricted to cone photoreceptors and their synaptic terminals but can be found in elements of the inner retina as well. A variety of immunocytochemical and other markers was used in combination with confocal microscopy on whole-mount preparations and tangential sections. Nearest neighbor analysis was performed and density recovery profiles as auto- and cross-correlograms were generated. Cells displaying a regular arrangement of their synaptic processes in matching radial register to each other were identified for each major retinal neuronal cell type except ganglion cells (i.e. photoreceptors, horizontal cells, bipolar cells, and amacrine cells). The precise location of some of the corresponding cell bodies was not as regular but still non-random, however there was no spatial cross-correlation between cell bodies of different types. The radial processes of Müller glial cells displayed a distribution correlating to the arrangement of photoreceptors and neurons. Thus, for one Müller glial cell I found two PKC-positive cone bipolar cells, a spatially corresponding grid of parvalbumin-positive amacrine cell processes, one H1 horizontal cell, and two pairs of double cones. There was no evidence among ganglion cells matching this pattern, possibly due to the lack of suitable markers. Although many other cell types do not follow this matching regular mosaic arrangement, a basic columnar building block can be postulated for the retina at least in cichlid fish. This suggests a functional radial unit from photoreceptors to the inner plexiform layer.     

GABAergic circuitry in the opossum retina: a GABA release induced by l-aspartate

Glutamate and γ-amino butyric acid (GABA) are the major excitatory and inhibitory neurotransmitters, respectively, in the central nervous system (CNS), including the retina. Although in a number of studies the retinal source of GABA was identified, in several species, as horizontal, amacrine cells and cells in the ganglion cell layer, nothing was described for the opossum retina. Thus, the first goal of this study was to determine the pattern of GABAergic cell expression in the South America opossum retina by using an immunohistochemical approach for GABA and for its synthetic enzyme, glutamic acid decarboxylase (GAD). GABA and GAD immunoreactivity showed a similar cellular pattern by appearing in a few faint horizontal cells, topic and displaced amacrine cells. In an effort to extend the knowledge of the opossum retinal circuitry, the possible influence of glutamatergic inputs in GABAergic cells was also studied. Retinas were stimulated with different glutamatergic agonists and aspartate (Asp), and the GABA remaining in the tissue was detected by immunohistochemical procedures. The exposure of retinas to NMDA and kainate resulted the reduction of the number of GABA immunoreactive topic and displaced amacrine cells. The Asp treatment also resulted in reduction of the number of GABA immunoreactive amacrine cells but, in contrast, the displaced amacrine cells were not affected. Finally, the Asp effect was totally blocked by MK-801. This result suggests that Asp could be indeed a putative neurotransmitter in this non-placental animal by acting on an amacrine cell sub-population of GABA-positive NMDA-sensitive cells.     

Lucifer yellow stains displaced amacrine cells of the chicken retina during embryonic development

We have used Lucifer Yellow for histological tracing of displaced amacrine cells within the ganglion cell layer (GCL) during the embryonic development of the chicken retina. Incubating whole eyes in the dye leads to bright staining of all displaced amacrine cells, whereas ganglion cells and glial cells are not stained. A subpopulation of cells of the inner part of the inner nuclear layer (INL) are also stained (for further details see ref. 13). Kainic acid, which is known to interfere with and kill amacrine cell systems, blocks the staining of these cells fully. This in addition to histological evidence confirms that the LY-stained cells in the GCL are displaced amacrine cells. Of the cells in the GCL, 23% (+/- 3%) are of the displaced amacrine type. Further, we find that the cytoarchitectural arrangement of these cells changes significantly during development.     

Developmental expression of calretinin immunoreactivity in the human retina and a comparison with two other EF-hand calcium binding proteins.

This paper reports the localization pattern of calretinin, a calcium-binding protein, in the human retina during development, as studied by immunohistochemistry. A comparison is made of the cellular distribution of calretinin with two other calcium-binding proteins, calbindin and parvalbumin, recently reported by us in the human retina, and by parallel labeling with both antisera in the same tissues. At 11-12 weeks of gestation, calretinin immunoreactivity was expressed in many prospective ganglion cells of the central inner neuroblastic zone. At 16-17 weeks of gestation, the immunoreactivity was localized in the ganglion cell layer, inner plexiform layer, and in most differentiated amacrine, horizontal and cone cells located in the central (1-2 mm temporal from optic disc) to midperipheral parts of the retina. By midgestation (20-21 weeks), calretinin immunoreactivity was strongly developed in the cone photoreceptors. Parallel labeling with calbindin and parvalbumin antisera revealed that the calretinin-positive horizontal cells were somewhat smaller and less frequent and less intense than the calbindin- and parvalbumin-positive counterparts, at 16-21 weeks of gestation. No horizontal cells were calretinin immunopositive in the postnatal (four-month-old infant) and adult retinas examined. Also, at both stages, a few bipolar and cone cells were weakly immunoreactive. These observations suggest a critical role for calretinin in the development and maturation of a select class of horizontal cells. The widespread expression of immunoreactivity in the early ganglion cells indicates that calretinin may be involved in their differentiation. The weak immunoreactivity pattern noted in the adult photoreceptor and bipolar cells, and an apparent lack of immunoreactivity in the mature horizontal cells, tends to indicate that, unlike calbindin and parvalbumin, calretinin plays little role in the transport and physiological buffering of Ca2+ in these neurons of the human retina. It appears, however, that calretinin is predominantly involved in both processes in amacrine cells.     

大鼠,金黄地鼠和家兔视网膜内一氧化氮合酶分布的比较

用ＮＡＤＰＨ黄递酶组织化学染色法观察了正常成年大鼠、金黄地鼠和家兔视网膜内一氧化氮合酶的分布，并比较了３种不同动物的区别。结果显示，在视网膜内ＮＯＳ阳性神经元主要为分布于内核层的无长突细胞、节细胞层的移位无长突细胞和少数节细胞，不同种类动物的视网膜内，ＮＯＳ阳性细胞的配布、密度和细胞形态均有差异。大鼠视网膜内ＮＯＳ阳性细胞多尾于内核层无长突细胞和节细胞层移位无长细胞，偶见于视网膜节细胞。金黄地鼠视     

Generation of retinal cells in the wallaby, Setonix brachyurus (quokka)

We have examined the generation of retinal cells in the wallaby, Setonix brachyurus (quokka). Animals received a single injection of tritiated thymidine between postnatal days 1-85 and retinae were examined at postnatal day 100. Retinae were sectioned, processed for autoradiography and stained with Cresyl Violet. Ganglion cells were labelled by injection of horseradish peroxidase into the optic tracts and primary visual centres. Other cells were classified according to their morphology and location. Retinal cell generation takes place in two phases. During the first phase, which concludes by postnatal day 30, cells destined to lie in all three cellular layers of the retina are produced. In the second phase, which starts by postnatal day 50, cell generation is almost entirely restricted to the inner and outer nuclear layers. Cells produced in the first phase are orthotopic and displaced ganglion cells, displaced and orthotopic amacrine cells, horizontal cells and cones. Glia in the ganglion cell layer, orthotopic amacrine cells, bipolar and horizontal cells. Muller glia, and rods are generated in the second phase. Cells became heavily labelled with tritiated thymidine in the central retina before postnatal day 7, over the entire retina (panretinal) by postnatal day 7 and from postnatal day 18, only in the periphery. The second phase of cell generation is initiated at P50, in a region extending from the optic nerve head to mid-temporal retina. Subsequently, cells are generated in annuli, centred on mid-temporal retina, which are seen at progressively more peripheral locations. Therefore, cell addition to the inner and outer nuclear layers continues for longer in peripheral than in mid-temporal retina. We suggest that such later differential cell addition to the inner and outer nuclear layers contributes to an asymmetric increase in retinal area. This non-uniform growth presumably results in more expansion of the ganglion cell layer peripherally than in mid-temporal retina and may play a role in establishing density gradients of ganglion cells.     

Protein gene product 9.5-immunoreactive neurons in the retina of striped dolphin (Stenella coeruleoalba) and Fraser dolphin (Lagenodelphis hosei).

This study demonstrates immunocytochemically that protein gene product 9.5 (PGP 9.5), a neuronal marker, is expressed by various populations of retinal cells in Stenella coeruleoalba (striped dolphin) and Lagenodelphis hosei (Fraser dolphin): one in the retinal ganglion cells and the other in the inner nuclear layer, resembling horizontal and amacrine cells. The specific distribution of PGP 9.5 in a dolphin closely resembles that in rodents and carnivores; however, some differences arise among these animals. In a dolphin's retina, for example, only a few of giant ganglion cells are immunoreacted while almost all the small ganglion cells are stained strongly. The processes of horizontal cells, identified according to their localization, appear not to connect entirely in a dolphin. Instead, PGP 9.5 positive cells are widely distributed in the small to moderate ganglion cells and have distinct processes which are ramified extensively in the outer plexiform layer in rodents and carnivores. The high levels of PGP 9.5 expressing in the inner part of dolphin retina, including ganglion cells and their axons as well as distinct sublamination in the inner plexiform layer, indicate that this molecule markedly influences the retinal system, possibly in visual connection. Although mammals have various visual behavior, i.e., living marine vs. terrestrial environment, and active during daytime vs. in the night, the retina is a common model to characterize the neurochemical properties.     

Histology of the ferret retina

The retina of the adult ferret, Mustelo furo, was studied with light and transmission electron microscopy to provide an anatomical basis for use of the ferret as a model for retinal research. The pigment epithelium is a simple cuboidal layer of cells characterized by a zone of basal folds, apical microvilli, and pigment granules at various stages of maturation. The distinction between rod and cone photoreceptor cells is based on their location, morphology, heterochromatin pattern and the electron density of their inner segments. The round, light-staining cone cell nuclei occupy the layer of perikarya along the apical border of the outer nuclear layer. The remainder of the outer nuclear layer consists of oblong, deeply-stained rod cell nuclei. Ribbon type synaptic complexes involving photoreceptor cell axons, horizontal cell processes, and bipolar cell dendrites characterize the outer plexiform layer. The inner nuclear layer is comprised of horizontal, bipolar, and amacrine cell perikarya as well as the perikarya of the Müller cells. The light-staining horizontal cell nuclei are prominent along the apical border of the inner nuclear layer. The light-staining amacrine cell nuclei form a more or less continuous layer along the basal border of the inner nuclear layer. Both conventional and ribbon-type synapses characterize the inner plexiform layer. The ganglion cells form a single cell layer. The optic fiber layer contains bundles of axons surrounded by Müller cell processes. Small blood vessels and capillaries are present in the basal portion of the retina throughout the region extending from the internal limiting membrane to the outer plexiform layer. The adult one-year-old retina is compared with the retina at the time of eye opening.     

Quantitative analysis of GABA-immunoreactive synapses in the inner plexiform layer of theBufo marinus retina: identification of direct output to ganglion cells and contacts with dopaminergic amacrine cells

Summary We have recently reported that about 50% of amacrine cells and some of the bipolar and ganglion cells are GABA-immunoreactive in the retina ofBufo marinus. Synapses formed by these elements in the inner plexiform layer were studied. GABA-immunoreactive amacrine cell processes were found most frequently in synaptic contact with non-immunoreactive amacrine cells. Double-label experiments showed that some of these non-GABA-immunoreactive elements contain tyrosine hydroxylase immunoreactivity. Another source of input to the GABA-immunoreactive amacrine cells were the bipolar cells; some of which were GABA-immunoreactive. GABA-immunoreactive amacrine cells synapsed also onto bipolar cell terminals, and ganglion cell dendrites that were identified by the retrograde transport of horseradish peroxidase from the optic nerve. Synapses between GABA-immunoreactive amacrine cells and bipolar and ganglion cells were non-uniformly distributed in the inner plexiform layer. Synaptic contacts with bipolar cells were more frequent in the OFF-sublamina, and those with ganglion cell dendrites in the ON-sublamina. These results demonstrate that GABA-immunoreactive amacrine cells (1) preferentially synapse with OFF-responding bipolar and ON-centre ganglion cells in the through-pathway, (2) synapse with tyrosine hydroxylase-immunoreactive amacrine cells in both the OFF- and ON-sublaminae, and (3) synapse directly with GABA-immunoreactive ganglion cells. The synapses between GABA-immunoreactive amacrine and GABA-immunoreactive ganglion cells may inhibit the centrally projecting inhibitory ganglion cells, causing disinhibition in the visual centres.On leave from Department of Zoology, Attila József University, Szeged, Hungary.     

牛蛙视网膜内γ-氨基丁酸免疫阳性反应的分布

用免疫组织化学ABC法．研究了GABA免疫阳性反应在牛蛙视网膜的分布。证明光感受器内段（主要是视锥细胞）呈棕褐色的GABA反应;在外核层未见GABA标记的胞体,但在靠近外网层处偶见GABA标记的终末;在内核层,大量无长突细胞呈GABA反应阳性,并可鉴别出胞体染色较深和淡的两个亚群,一些双极细胞和个别水平细胞的胞体及它们的突起呈较弱的GABA反应阳性,偶见双极细胞轴突终末以膨体紧密贴附在GABA标记的无长突细胞上。在节细胞层,一些神经节细胞和散在的移位无长突细胞呈GABA反应阳性。此外．外网层和内网层均呈GABA反应阳性。上述结果表明,GABA广泛分布于牛蛙视网膜的各层,提示它在视觉信号的传递过程中发挥着重要作用。     

The neurons of the ground squirrel retina as revealed by immunostains for calcium binding proteins and neurotransmitters

Ground squirrel retinas were immunostained with antibodies against calcium binding proteins (CBPs) and classical neurotransmitters in order to describe neuronal phenotypes in a diurnal mammalian retina and to then compare these neurons with those of more commonly studied nocturnal retinas like cats' and rabbits'. Double immunostained tissue was examined by confocal microscopy using antibodies against the following: rhodopsin and the CBPs, calbindin, calretinin, parvalbumin, calmodulin and recoverin (CB, CR, PV, CM, RV), glycine, GABA, choline acetyltransferase (CHAT) and tyrosine hydroxylase (TOH).In ground squirrel retina, the traditional cholinergic mirror symmetric amacrine cells colocalize CHAT with PV and GABA and faintly with glycine. A second cholinergic amacrine cell type colocalizes glycine alone. CR is found in at least 3 different amacrine cell types. The CR-immunoreactive (IR) cell population is a mixture of glycinergic and GABAergic types. The dopamine cell type IR to tyrosine hydroxylase has the typical morphology of a wide field cell with dendrites in S1 but the rings seen in cat or rabbit retina are not as numerous. TOH-IR amacrine cells send large club-shaped processes to the outer plexiform layer. CB and CR are in bipolar cells, A- and B-type horizontal cells and several amacrine cell types. Anti-rhodopsin labels the low density rod photoreceptor population in this species. Anti-recoverin labels cones and some bipolar cells while PKC is found in several different bipolar cell types. One ganglion cell with dendritic branching in S3 is strongly CR-IR.We find no evidence for an AII amacrine cell in the ground squirrel, with either anti-CR or anti-PV. An amacrine cell with similarity to the DAP1-3 cell of rabbit is CR-IR and glycine-IR. We discuss this labeling pattern in relationship to other mammalian species. The differences in staining patterns and phenotypes revealed suggest a functional diversity in the populations of amacrine cells according to whether the retinas are rod or cone dominated.     

Cholinergic amacrine cells of the chicken retina: a light and electron microscope immunocytochemical study

Cholinergic amacrine cells of the chicken retina were detected by immunohistochemistry using an antiserum against affinity-purified chicken choline acetyltransferase. Three populations of cells were detected: type I cholinergic amacrine cells had cell bodies on the border of the inner nuclear and inner plexiform layers and formed a prominent laminar band in sublamina 2 of the inner plexiform layer, while type II cholinergic amacrine cells had cell bodies in the ganglion cell layer, and formed a prominent laminar band in sublamina 4 of the inner plexiform layer. Type III cholinergic amacrine cell bodies were located towards the middle of the inner nuclear layer, and their processes were more diffusely distributed in sublaminas 1 and 3-5 of the inner plexiform layer. Type I and type II cells were present at densities of over 7000 cells/mm2 in central areas declining to less than 2000 cells/mm2 in the temporal retinal periphery. The cells were organized locally in a non-random mosaic, with regularity indices ranging from 3 peripherally to over 5 centrally. Neither at the light nor electron microscopic levels was a lattice of cholinergic dendrites of the kind reported by Tauchi and Masland [J. Neurosci. 5, 2494-2501 (1985)] detectable. Within the two prominent dendritic plexuses, a major feature of the synaptic interactions of the type I and type II cholinergic cells was extensive synaptic interaction between cholinergic processes. Apart from this, there was little, if any, input to cholinergic processes from non-cholinergic amacrine cells, but there was input from bipolar cells. Output from the cholinergic amacrine cell processes was directed towards non-cholinergic amacrine cells as well as other cholinergic amacrine cells, and ganglion cells.