Somatostatinergic neurones of the developing human and cat retinae

We have examined somatostatin-immunoreactive (S-IR) neurones in developing retinae of the human and cat. At 14 and 16 weeks' gestation (G14 and G16) in the human, S-IR cells were only found close to the putative fovea centralis, but by 18 weeks' gestation (G18), they were located in all retinal regions. By adulthood, the majority of S-IR cells were restricted to inferior retina. In the developing cat retina, two classes of S-IR cells were recognized. S1-IR cells were similar in morphology and distribution to adult cells: they had small round somata which were only found in inferior retina and gave rise to beaded processes which traversed the inner plexiform layer (IPL) and nerve fibre layer (NFL). S2-IR cells had larger somata located in the ganglion cell layer (GCL) and the label was compartmentalized within their cytoplasm. Most S2-IR cells had lost immunoreactivity by P (postnatal day) 25 and may have been alpha-ganglion cells transiently expressing somatostatin in association with their retention of plasticity into postnatal life.     

Distinct patterns of distribution among NADPH-diaphorase neurones of the guinea pig retina

We have examined the morphology and distribution of nicotinamide adenine dinucleotide phosphate diaphorase (NADPH-diaphorase) cells in the retina of the guinea pig. Two morphologically distinct classes of labelled cells were detected, one with larger, darkly labelled somata commonly located in the inner nuclear layer (INL: NDa cells) and the other with smaller, lightly labelled somata in the ganglion cell layer (GCL: NDb cells). The somata of NDb cells did not vary in diameter with eccentricity, whereas those of the NDa cells were smallest in the visual streak. The number of NDa cells was approximately 3,500, with a mean density of 26/mm2 and NDb cells numbered approximately 4,400, with a mean density of 33 mm2. NDa cells were distributed relatively uniformly across the retina, whereas NDb cells concentrated in the visual streak and were restricted to the superior half of the retina. In these features of morphology and distribution. NADPH-diaphorase neurones of the guinea pig retina are distinct from those observed in other species. It remains to be elucidated whether the diversity in the morphology and distribution of NADPH-diaphorase neurones between species reflects a diversity in their function.     

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.     

Localization of glycine uptake and receptors in the cat retina

Immunocytochemistry using a monoclonal antibody against glycine receptors revealed that these receptors in cat retina are confined to the inner plexiform layer (IPL). The outer half of that layer showed strong patchy labelling with some indication of two bands corresponding to the on- and off-sublamina. High-affinity uptake of [3H]glycine followed by autoradiography labelled amacrine cells, bipolar cells and the outer portion of the IPL. Silver grains in the IPL were patchily distributed. These results indicate in the cat retina a close match between presynaptic glycinergic elements labelled by high-affinity uptake and the postsynaptic receptor sites for glycine revealed by immunocytochemistry.     

Demonstration of a neuropeptide Y (NPY)-like immunoreactivity in the pigeon retina

The distribution of neuropeptide Y (NPY)-like immunoreactivity in the pigeon retina was investigated by fluorescence immunohistochemistry. NPY-positive cells were found in central and peripheral retina. NPY somata were located in the proximal portion of the inner nuclear layer and their processes directed to the inner plexiform layer where they ramified in 3 immunoreactive bands. NPY might play a role as a neurotransmitter or neuromodulator in the pigeon retina.     

Localization of substance P-like immunoreactivity in the adult and developing goldfish retina

Substance P-like immunoreactivity was localized to amacrine cells in both adult and developing goldfish retina using immunohistochemical techniques. These studies utilized a well-characterized monoclonal antiserum directed to substance P. Specificity was established by absorption of the anti-serum with 10 μm synthetic substance P. Specific substance P-like immunoreactivity was localized within a seemingly distinct population of unistratified amacrine cells which were distributed in both central and peripheral retinal regions. The immunoreactive somata were located at the border of the inner nuclear layer and inner plexiform layer and were characterized by a round or ovoid somata which measured about 9μm in diameter. These immunoreactive amacrine cells typically had a single process which descended to and ramified within lamina 3 of the inner plexiform layer.Specific substance P-like immunoreactivity first appeared 60 h after hatching (stage 27) within both somata and processes located in differentiated retinal regions. No substance P-like immunoreactive somata or processes were observed in undifferentiated retinal regions. In retinas from stage 27 to 14 days after hatching, the immunoreactive somata were characterized by an ellipsoidal soma and a large nucleus devoid of immunoreactivity. These immunoreactive cells were also characterized by a single process that descended to and ramified within lamina 3 of the differentiated inner plexiform layer. At 30 days after hatching, the substance P-containing cells were identical in appearance to these same cell types observed within the adult retina.     

Synapsin I expression in the rat retina during postnatal development

Summary The expression of the synapsin I gene was studied during postnatal development of the rat retina at the mRNA and protein levels. In situ hybridization histochemistry showed that synapsin I mRNA was expressed already in nerve cells in the ganglion cell layer of the neonatal retina, while it appeared in neurons of the inner nuclear layer from postnatal day 4 onward. Maximal expression of synapsin I mRNA was observed at P12 in ganglion cells and in neurons of the inner nuclear layer followed by moderate expression in the adult. At the protein level a shift of synapsin I appearance was observed from cytoplasmic to terminal localization during retinal development by immunohistochemistry. In early stages (P4 and P8), synapsin I was seen in neurons of the ganglion cell layer and in neurons of the developing inner nuclear layer as well as in the developing inner plexiform layer. In the developing outer plexiform layer synapsin I was localized only in horizontal cells and in their processes. Its early appearance at P4 indicated the early maturation of this cell type. A shift and strong increase of labelling to the plexiform layers at P12 indicated the localization of synapsin I in synaptic terminals. The inner plexiform layer exhibited a characteristic stratified pattern. Photoreceptor cells never exhibited synapsin I mRNA or synapsin I protein throughout development.Abbreviations GCL ganglion cell layer - INB inner neuroblast layer - INL inner nuclear layer - IPL inner plexiform layer - ONB outer neuroblast layer - ONL outer nuclear layer - OPL outer plexiform layer     

Distribution of GABA immunoreactivity in kainic acid-treated rabbit retina

Ischaemic retinal cell degeneration seems to involve both NMDA and non-NMDA receptor over stimulation. However, different retinal cell types differ largely in their susceptibility to excitatory amino acid induced neurotoxicity. We have investigated the vulnerability of GABAergic cells in the rabbit retina to the non-NMDA receptor agonist kainic acid (KA). The distribution of GABA immunoreactivity (GABA-IR) was examined in the central inferior retina at different survival times (5 h–6 days) following an intra-ocular injection of 140 nmol KA and compared to that of control and untreated retinas. In the normal retina, the majority of GABA-positive cells (79%) were located in the inner nuclear layer (INL), in one to four cell rows next to the inner plexiform layer (IPL), and in one cell row next to the outer plexiform layer (OPL). The remainder (21%) were found in the ganglion cell layer (GCL). Dense immunoreactivity was seen throughout the IPL. In the OPL, stained dots and occasional immunoreactive large processes could be seen. KA-exposed retinas processed for GABA immunocytochemistry 5 and 24 h after the injection showed an 85% reduction in the number of GABA immunoreactive cells. About the same degree of depletion was seen among GABA-IR cells located at different retinal levels. However, at these survival times, immunostaining was observed in three distinct bands in the IPL, indicating that the vulnerability to KA is not uniformly distributed among all GABAergic cells. At 48 h, an additional decrease in the number of labelled cells was noted, but immunoreactive cells were still found both in the INL and GCL. Even 6 days after KA treatment, a few stained cell bodies were seen in the INL next to the IPL, as well as a few processes in the IPL. The study shows that KA receptor overstimulation induces a marked depletion of the endogenous cellular GABA pools of the central rabbit retina, most likely as a result of GABAergic cell loss. However, a small population of GABAergic cells located in the INL appears to be less vulnerable to the toxic effects of 140 nmol KA.     

Peptide histidine-isoleucine (PHI)-immunoreactive amacrine cells in the retina of the rat

Peptide histidine-isoleucine (PHI) immunoreactivity was located in amacrine-like cells in adult rat retina by use of immunohistochemical techniques. Immunoreactive somata were found in the proximal part of the inner nuclear layer. From these somata, processes could be followed into the inner plexiform layer. The terminals of these processes were mainly found in the sublayers, 1, 2, and 3, but a few terminals were also present in the other inner plexiform sublaminae. The distribution of PHI-immunoreactive somata and processes corresponds with the cellular distribution of vasoactive intestinal peptide (VIP) and indicates a possible co-localization with this peptide.     

Ionotropic purinergic receptors P2X in frog and turtle retina: Glial and neuronal localization

Purinergic signaling is represented in both the peripheral and central nervous system (CNS), and in particular in the retina, which may be regarded as a part of the CNS. While purigenic signaling is relatively well studied in mammalian retinas, little is known about it in retinas of lower vertebrates. The aim of present study was to investigate, using immunocytochemistry, the distribution of purinoreceptors P2X in retinas of frog and turtle, which are appropriate models of the brain neuron-to-glia interactions. The results showed widespread expression of all seven ionotropic purinoreceptors (P2X1–P2X7) in both frog and turtle retinas. They were predominantly expressed in Müller cells, the principal glial cells in the retina. All structures typical of Müller cells: the outer and the inner limiting membranes, the cells bodies in the inner nuclear layer, the radial processes in the inner plexiform layer (IPL), and the so called endfeet (frog) or the orthogonal arrays of particles (turtle) in the ganglion cells layer were immunostained. Colocalizations between P2X1–P2X7 and the glial cell marker Vimentin proved that the immunostaining was in the Müller cells. In addition to the glial staining, neuronal staining was also seen as fine puncta in the inner plexiform layer and by small dots and patches in the outer plexiform layer. Some cell bodies of horizontal, amacrine and ganglion cells were also stained. The results obtained imply that the purinergic P2X receptors may significantly contribute to the neuron-to-glia signaling in retinas of the lower vertebrates.     

Displaced retinal ganglion cells project to the accessory optic system in the chameleon ( Chamaeleo calyptratus)

Retinal ganglion cells were successfully labelled in the chameleon by retrograde axonal transport of dextran amines that were applied to the nucleus of the basal optic root (nBOR) in an in vitro preparation. Labelled ganglion cells were restricted to the contralateral eye. Many cells were completely stained including their dendritic trees. With few exceptions, all cells had displaced somata that were located at the inner margin of the inner nuclear layer. The labelled ganglion cells had two to six primary dendrites that branched frequently and formed large unistratified dendritic trees within sublamina 1 of the inner plexiform layer. There was extensive overlap of the dendritic trees of neighbouring cells leading to an estimated coverage factor of 2-4. The dendritic field areas varied in size according to the retinal position of the cells and were highest in the central retina around the fovea with a maximum of 0.14 mm(2) and reached a second maximum at the retinal margin with values of 0.08-0.1 mm(2). The smallest dendritic areas (0.04-0.06 mm(2)) were measured midway between the fovea and retinal margin. The size of the soma area was not correlated to the dendritic field size and increased from 100 to 150 microm(2) near the fovea to 150-300 microm(2) at the retinal margin. There was no evidence for a retinotopic organisation of ganglion cell fibres within the nBOR. All cells were of uniform morphology that was identical to the type of nBOR-projecting displaced ganglion cell (DGC) described previously for the bird retina. Similar to birds, the labelled DGCs were the only source of retinal projection to the nBOR. A small fraction of cells had orthotopic somata located in the ganglion cell layer but were otherwise identical to the labelled DGCs. The similarity of chameleon nBOR-projecting ganglion cells to those described in avian retinas mirrors the close phylogenetic relationship of birds and lizards.     

游离锌离子在小鼠视网膜的定位研究

目的研究游离锌离子在小鼠视网膜的定位分布。方法应用ZnSe金属自显影技术(AMG)检测硒酸钠注射40 m in后小鼠视网膜内的锌离子。结果注射硒酸钠40 m in后发现游离锌离子主要分布于小鼠视网膜的色素上皮细胞层、光感受器的内节、外核层、外网层、内核层、内网层和神经节细胞层。在色素上皮细胞层、光感受器的内节和内核层与内网层交界处AMG阳性反应最为明显,在光感受器外节和神经纤维层几乎没有AMG阳性反应产物。结论小鼠视网膜内锌离子,在视网膜神经元视觉信息的传导和形成过程中可能起着重要作用。     

Tyrosine hydroxylase immunoreactive interplexiform cells in the lamprey retina

The distribution of tyrosine hydroxylase immunoreactivity was investigated in retinae of metamorphic, postmetamorphic and adult lampreys. Immunoreactive cell bodies were located mainly in the innermost part of the inner nuclear layer, with a few cells scattered throughout the inner plexiform layer. The processes of these neurons ran preferentially in the inner plexiform layer. Additionally, dense plexus of labelled processes were observed in the outer plexiform and nuclear layers. These findings suggest that most of the tyrosine hydroxylase-immunoreactive cells in the lamprey retina are interplexiform cells.     

The light microscopic localization of substance P- and somatostatin-like immunoreactive cells in the larval tiger salamander retina

Summary Light microscopic immunocytochemistry was utilized to localize the populations of substance P (SP)- and somatostatin (SOM)-like immunoreactive cells in the larval tiger salamander retina. Of 104 SP-immunostained cells observed, 82% were Type 1 amacrine cells. Another 8% of the SP-cells were classified as Type 2 amacrine cells, while 10% of the SP-cells had their cell bodies located in the ganglion cell layer and were designated as displaced amacrine cells. Each type of SP-like immunoreactive cell was observed in the central and peripheral retina. SP-immunopositive processes were observed in the inner plexiform layer as a sparse plexus in sublamina 1 and as a denser network of fibers in sublamina 5. Seventy-eight percent of the 110 somatostatin-immunopositive cells observed were designated as Type 1 amacrine cells. Another 12% of SOM-cells were classified as displaced amacrine cells, while only two SOM-immunopositive Type 2 amacrine cells were observed. Nine percent of the SOM-cells were designated as interplexiform cells, based on their giving rise to processes distributing in the outer plexiform layer as well as processes ramifying in the inner plexiform layer. Each type of SOM-immunoreactive cell was observed in the central and peripheral retina, with the exception of the Type 2 amacrine cells, whose somas were only found in the central retina. Lastly, SOM-immunopositive processes in the inner plexiform layer appeared as a fine plexus in sublamina 1 and as a somewhat denser network of fibers in sublamina 5.     

Immunocytochemical labelling of horizontal cells in mammalian retina using antibodies against calcium-binding proteins

Using antibodies against calcium-binding proteins immunocytochemistry revealed quantitative staining of horizontal cells in whole mount preparations. In cat both A- and B-type horizontal cells, and in monkey probable H1- and H2-horizontal cells were labelled with antibodies against parvalbumin. In rabbit and ox two types of horizontal cell were labelled with antibodies against calcium-binding protein (CaBP-28K). In ox retina processes of horizontal cells were observed descending into the inner plexiform layer (IPL).     

Displaced retinal ganglion cells in normal frogs and those with regenerated optic nerves

Summary We have analysed the number and spatial distribution of displaced retinal ganglion cells in the frog Litoria (Hyla) moorei. A series of normal animals was compared with one in which the optic nerve was crushed and allowed to regenerate. Ganglion cells were labelled with horseradish peroxidase (HRP) applied to the optic nerve, and retinae were examined as sections or whole mounts. We analysed separately ganglion cells with somata displaced to the inner nuclear (Dogiel cells, DGCs) and to the inner plexiform layer (IPLGCs). These findings were related to data for the orthotopic ganglion cells (OGCs). The mean number of DGCs in the normal series was 2,550 (±281) and fell to 1,630 (±321) after regeneration, representing a mean loss of 36%. This reduction was not significantly different from the mean loss of 43% from the OGC population in which mean values fell from 474,700 (±47,136) to 268,700 (±54,395). In both the normal and the regenerate series, DGCs were estimated to represent means of only 0.6% of the OGC population. Densities of DGCs were highest in the nasoventral and temporo-dorsal peripheries; densities of both DGCs and OGCs were lower after optic nerve regeneration. We conclude that the factors which affect ganglion cell death during optic nerve regeneration, do so to similar extents amongst the DGC and the OGC populations. The IPLGCs were very rare in normal animals with a mean of 420 (±95). However, their numbers increased after regeneration to a mean of 3,350 (±690), estimated to be 1.2% of the OGC population. These cells normally favoured peripheral retina but became pan-retinal after regeneration. The primary dendrites of the majority of IPLGCs were oriented in the same direction as those of OGCs. We conclude that most IPLGCs were OGCs which had relocated their somata to the inner plexiform layer.     

Lamina formation in the Mongolian gerbil retina (Meriones unguiculatus)

Retinae of nocturnal rodents, such as mice and rats, are almost exclusively rod-dominated. The gerbil, in contrast, shows active periods during day and night and uses both rod- and cone-based vision. However, its retina has not been studied in detail, except for one developmental study analysing its prenatal period (Wikler et al. 1989). Here, the formation of the laminar structure of the gerbil retina was studied from birth until late adult stages. At birth, the retina consisted of a wide neuroblastic layer, with 30% of cells still dividing, a rate decreasing to nearly zero by P6. Shortly after birth, segregation of a ganglion cell layer began. All retinal layers reached their final size around P20, as determined from DAPI-stained cryosections. Müller glial cells developed their typical structure from P1 onwards, e.g. announcing an outer plexiform layer (OPL) at P5, as analysed by the Ret-G7 and glutamine synthetase antibodies. The analyses of the inner retina were performed by antibodies to calretinin (CR) and calbindin (CB). CR is expressed in ganglion cells followed by amacrine cells from P1 onwards; their processes formed four subbands in the inner plexiform layer (IPL) and appeared sequentially after P5 until P20. CB stained a subtype of horizontal cells with their processes into the OPL from P14 onwards. The rod-specific antibody rho4D2 announced photoreceptors at P4, showing signs of outer segments from P10 onwards. The study shows that the formation of all retinal layers in the gerbil occurs postnatally. This and the fact that the gerbil retina is not exclusively rod-dominated could render the gerbil a valuable model for in vitro studies of retinogenesis in rodents.     

An immunocytochemical marker for hamster retinal ganglion cells

Summary We examined the specificity and developmental time course of the labelling of retinal ganglion cells in Syrian hamsters by a monoclonal antibody AB5. In adult hamsters, AB5 selectively labelled somata in the ganglion cell layer, dendrites in the inner plexiform layer and axons in the nerve fibre layer. When retinal ganglion cells were retrogradely labelled with Dil prior to AB5 immunocytochemistry, all of the retrogradely labelled retinal ganglion cells in the ganglion cell layer were AB5 immunoreactive, indicating that AB5 labels all classes of ganglion cell in that layer. In retinae depleted of retinal ganglion cells by neonatal optic nerve transections, AB5 did not label any somata or processes, indicating that AB5 specifically labels retinal ganglion cells. During development, AB5 labelling first appeared as a weak staining of cell bodies in the ganglion cell layer on postnatal day 12 (P12; PO=first 24 h following birth) and acquired the staining pattern seen in the adult by postnatal day 14. From the onset of AB5 immunoreactivity, AB5-labelled somata of varying sizes were present across the entire retinal surface. Although AB5 labelled retinal ganglion cell axons in the nerve fibre layer of the retina it did not label the optic nerve or retinal ganglion cell axons in the brain at any age examined. AB5 labelling was also found to be compatible with bromodeoxyuridine immunocytochemistry and, therefore, useful for determining the time of generation of hamster retinal ganglion cells.     

Neuronal and glial localization of homocysteate-like immunoreactivity in the rat retina

Summary To study the distribution ofl-homocysteate in the rat retina, specific polyclonal and monoclonal anti-homocysteate antibodies have been used in combination with a highly sensitive postembedding method for light microscopic immunocytochemistry. In central and peripheral retina, the most strongly immunoreactive cell bodies lay in the inner nuclear layer. They represented about 17% of the total neuronal cell population of the layer and were identified as bipolar cells (19–20% of cells in the outer half of the inner nuclear layer) and amacrine cells (15% of cells in the inner half of the inner nuclear layer). A third cell type showing heavy homocysteate-like immunoreactivity was identified as Müller glial cells. Characteristically, their descending processes formed three immunoreactive bands in the inner plexiform layer. Furthermore, the outer and inner limiting membranes as well as glia around and between ganglion cell axons and in the vicinity of blood vessels were labelled intensely. Photoreceptors and their terminals, and ganglion cells, were not immunostained. These findings indicate the presence of homocysteate in some bipolar and amacrine cells of the inner nuclear layer and support a role for this sulphur-containing excitatory amino acid as a neurotransmitter candidate in the retina.