Naturally occurring cell death and migration of microglial precursors in the quail retina during normal development.

We compared chronotopographical patterns of distribution of naturally occurring neuronal death in the ganglion cell layer (GCL) and the inner nuclear layer (INL) with patterns of tangential and radial migration of microglial precursors during quail retinal development. Apoptotic cells were identified by the terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end labeling technique, and microglial precursors were identified by immunocytochemistry with an antibody recognizing quail microglial cells (QH1 antibody). Apoptotic cells were first detectable in the GCL at the seventh day of incubation (E7), were most abundant at E10, and were absent after E13. In the INL, apoptotic cells first appeared at E7, were most abundant at E12, and disappeared entirely after the third posthatching day (P3). In both retinal layers, cell death first appeared in a small central area of the retina and subsequently spread along three gradients: central-to-peripheral, temporal-to-nasal, and dorsal-to-ventral. The chronology of tangential (between E7 and E16) and radial migration (between E8 and P3) of microglial precursors was highly coincident with that of cell death in the GCL and INL. Comparison of the chronotopographical pattern of distribution of apoptotic nuclei in the GCL with the patterns of tangential and radial migration of microglial precursors neither supported nor refuted the hypothesis that ganglion cell death is the stimulus that triggers the entry and migration of microglial precursors in the developing retina. However, microglial cells in most of the retina traversed the INL only after cell death had ceased in this layer, suggesting that cell death in the INL does not attract microglial precursors migrating radially. Dead cell debris in this layer was phagocytosed by Müller cells, whereas migrating microglial cells were seen phagocytosing apoptotic bodies in the nerve fiber layer and GCL but not in the INL.     

Reelin expression in the retina and optic tectum of developing common brown trout

Reelin (RELN) is an extracellular matrix protein largely related with laminar organization in several brain areas. The development of RELN immunoreactivity in the retina and the optic tectum of the brown trout are analyzed with a monoclonal (142) antibody against RELN whose suitability has been ascertained by western blot. In the retina of embryos and alevins, RELN immunoreactivity is detected in cells of the ganglion cell layer (GCL) and inner nuclear layer (INL), and in the inner plexiform layer (IPL), where it appears as "diffuse" material confined to the ON-sublayer. In juveniles, RELN expression becomes restricted to a stripe of cells in the INL. RELN-immunoreactive (RELN-ir) cells are absent from the outer nuclear layer (ONL) at any developmental stage. The developmental pattern of RELN expression in the trout retina shows many similarities with that of amniotes: (a) RELN expression parallels the vitreal to scleral progression of differentiation of the retina and, within each cell layer, RELN immunoreactivity appears confined to a subpopulation of postmitotic cells; (b) at early stages RELN expression is exclusively observed in the central retina and as maturation progresses from the center to the periphery, more RELN-ir cells are observed following the same spatial pattern. Differences with amniotes are noted regarding the absence of RELN expression in the GCL and INL in adulthood, and in the ONL at any developmental stage. In the optic tectum (OT) of trout, as in amniotes, RELN immunoreactivity increases within specific cell layers as lamination proceeds, and decreases when it is complete, except in the stratum opticum (SO), where RELN-ir cells are observed throughout life. Time-course expression of RELN in the OT suggests a role in the early modeling of synaptic contacts and the accommodation of new retinal arriving axons throughout life.     

Photoreceptor cell rescue at early and late RPE-cell transplantation periods during retinal disease in RCS dystrophic rats

Normal retinal pigment epithelial (RPE) cells were transplanted into retinas of Royal College of Surgeons (RCS) dystrophic rats at different stages of the retinal disease process. RPE-cell transplantation at 10, 17 and 26 days resulted in rescue of photoreceptor cells, such that at 4 months the outer nuclear layer (ONL) was 8-10 cells in thickness as shown in retinas of age-matched control rats. Of these transplantation times, day 17 appeared to affect the best rescue of photoreceptor cells. Nongrafted retinas of 4 month-old RCS dystrophic rats exhibited scattered PRC's, most prevalent in the peripheral retina. In addition, a small, but significant increase in the ONL thickness was detected in vehicle-injected retinas (sham control) of 17 day-old RCS dystrophic rats at 2 months; however, at 3 months, the ONL thickness was reduced to control levels. A normal distribution of (Na+ + K+)-ATPase immunostain was demonstrated beneath grafted RPE cells in retinas of 4 month-old RCS dystrophic rats. Dense immunostaining was shown along rescued photoreceptor cell inner segments (IS), within the inner (IPL) and outer (OPL) plexiform layers and on plasmalemma of cell bodies in the inner nuclear layer (INL). In nongrafted retinas of age-matched RCS dystrophic rats, immunostaining for (Na+ + K+)-ATPase was observed only in the INL and IPL. Under RPE-cells transplants in retinas of 4 month-old RCS dystrophic rats, opsin immunostaining was detected along both rescued photoreceptor cell inner and outer (OS) segments and on plasmalemma of ONL cell bodies. However, immunostaining for opsin was restricted to a debris zone in nongrafted retinas of age-matched RCS dystrophic rats.(ABSTRACT TRUNCATED AT 250 WORDS)     

Birth and fate of proliferative cells in the inner nuclear layer of the mature fish retina

In teleost fish, unlike other vertebrates, the retina continues to grow throughout the animal's life both by stretching of the mature tissue and by the addition of new cells. Following larval development, new retinal cell birth is known to occur in a rim at the periphery of the mature retina and in the outer nuclear layer (ONL). We have now found that cell birth and proliferation also occurs in the inner nuclear layer (INL) of the mature fish retina. In rainbow trout (Onchoryncus mykiss), proliferative cells exist in the INL of fish of all ages, at least up to 2 years posthatching. The proliferative cells form clusters in the INL that align in radial columns, reaching from the inner to the outer plexiform layers. The density of proliferative cell clusters changes along the equatorial plane of the retina and is highest near both the nasal and temporal poles. Our data suggest that, after birth, the proliferative cells migrate away from the INL and into the ONL, with a half-time of about 3 days, and their cell bodies can be seen in the outer plexiform layer. Once they are in the ONL, the proliferative cells continue to divide and likely give rise to the precursor cells that differentiate into new rod photoreceptors. J. Comp. Neurol. 394:271–282, 1998. © 1998 Wiley-Liss, Inc.     

Developmental neuronal death is not a universal phenomenon among cell types in the chick embryo retina

The goal of this study was to investigate whether all the cell types present in the chick embryo retina undergo developmental neuronal death. Apoptosis was investigated in retinal sections at different developmental stages, processed either with propidium iodide, which stains pyknotic nuclei intensely, or with terminal transferase-mediated deoxyuridine triphosphate (d-UTP)-biotin nick-end labeling (TUNEL). Internucleosomal DNA fragmentation was investigated in tissue extracts by agarose gel electrophoresis. TUNEL-positive (T+) cells and pyknotic nuclei were first detectable in the ganglion cell layer (GCL) around embryonic day (ED) 8 and peaked at ED 10. In the inner nuclear layer (INL), T+ and pyknotic cells first appeared on ED 8, reached maximum frequency on ED 11, and were largely absent after ED 14. DNA ladders were observed at all the stages, when T+ and pyknotic cells were abundant, but not on ED 4, when only scattered dead cells were observed histologically. Dying cells were virtually never detected in the outer nuclear layer (ONL) from ED 4 to postnatal day 2. After unilateral midbrain ablation on ED 5, there was a striking increase in the number of pyknotic and T+ cells in both the GCL and in the INL of the contralateral eye but not in the ONL. The absence of apoptotic cell death in the ONL during normal development and after tectal ablation shows that developmental death is not universal among the various cell populations present in the chick embryo retina and raises questions regarding mechanisms controlling both photoreceptor survival and the matching of pre- and postsynaptic elements in the outer plexiform layer of this species. J. Comp. Neurol. 396:12–19, 1998. © 1998 Wiley-Liss, Inc.     

TrkA, TrkB and p75 mRNA expression is developmentally regulated in the rat retina

We examined the cellular distribution of mRNAs coding for the neurotrophin receptors TrkA, TrkB and p75 in the rat retina during early postnatal development. At PO (postnatal day 0), mRNAs coding for each of the three receptors were detected in the ganglion cell layer (GCL) and in the inner plexiform layer (IPL), the latter structure essentially containing retinal ganglion cell processes at this developmental stage. At P5, the innermost part of the inner nuclear layer (INL) also expressed TrkA, TrkB and p75 mRNAs. Finally, the GCL, IPL and the whole INL of P10 retinae were labeled by the three probes. The developmentally regulated expression of these receptors underlies a possible role for neurotrophins in the differentiation and survival of retinal cells.     

Nonuniform distribution of cell proliferation in the adult teleost retina

Teleost fish continue to grow throughout life, and their eyes enlarge correspondingly. Within the eye, the retina grows by stretching existing tissue and adding new cells. Cell addition occurs in two ways: First, all cell types except rod photoreceptors are added circumferentially at the edge of the eye where the retina meets the iris; second, rod photoreceptors are generated from a population of rod progenitor cells which divide throughout the outer nuclear layer (ONL). To determine the spatial distribution of rod progenitor cells across the teleost retina, we labeled dividing cells with an antibody to proliferating cell nuclear antigen (PCNA) throughout a 24 h period. We found a significantly higher density of dividing rod precursor cells at the nasal and temporal margins than in the central retina throughout the 24 h cycle. At night, the density of dividing cells is significantly greater at the nasal pole of the eye. The difference between cell division at the center and the margin was reduced at night when the density of cell division in the central retina increased significantly. Taken together, these data suggest that the eye grows asymmetrically, with more cells added at the nasal pole. Possible developmental causes and functional consequences of the reported distribution of cell divisions in time and location are presented.     

Neuronal addition and retinal expansion during growth of the crucian carp eye

Using standard paraffin technique the addition of new cells in crucian carp retinas was examined. Between eye diameters 4.4 and 10.0 mm the number of ganglion cells increases from 103,000 to 205,000, INL cells from 1.5 to 3 million, cones from 250,000 to 900,000, and rods from 2 to 9 million. Concomitantly retinal area increases fivefold and the cell densities decrease by 37% for the cones, 57% for th e INL cells, and 58% for the ganglion cells, while the rod density remains stable. In relation to the rods the cell ratios at different retinal loci undergo marked changes during growth. The contributions to retinal growth by addition of new neurons and by expansion of the retina have been determined for the different retinal layers. The layer of rods grows exclusively by addition of new rod mosaic. In the cone layer 81% of growth is due to addition of new cone mosaic. In the inner nuclear layer (INL) 56% of growth is due to addition of new cells and in the ganglion cell layer 52% is due to cell addition. In each case retinal expansion accounts for the remainder of increase in retinal area. On morphological grounds six cone types can be found in the crucian carp retina. Their ratios are constant during retinal growth and at different retinal loci.     

Novel organotypic culture model of adult mammalian neurosensory retina in co-culture with retinal pigment epithelium

PURPOSE: The purpose of this study was to assess survival of adult mammalian neurosensory retina cultured in contact with the layer of a choroid-retinal pigment epithelium (RPE) explant. METHODS: The entire adult porcine neurosensory retina and RPE-choroid layer were placed in tissue culture by juxtaposing both tissues in their original orientation. Culture of the neurosensory retina alone and freshly prepared retina were used as control. After 3 days in culture retinal explants were fixed and processed for immunohistochemistry and TUNEL technique. RESULTS: We observed limited nuclei loss and significant reduction in apoptotic cells in nuclear cell layers (GCL, INL, and ONL) and decreased Muller cell hypertrophy in retina-RPE cultures compared to retinal cultures alone. In addition, cultures were characterized by reduced upregulation of GFAP, vimentin as well as S100 and increased glutamine synthetase expression. CONCLUSIONS: As any tissue culture model, retinal tissue culture is a short-term system and since degenerative processes begin quite early it may be a good model to investigate degenerative processes in the retina. However, our model of culture of retina adjacent to the RPE-choroid layer improves the maintenance of neural retina as evidenced by reduced apoptosis in nuclear cell layers (GCL, INL, and ONL) and reduced gliosis as indicated by the diminished expression of glial-specific proteins and increased glutamine synthetase compared to cultures of retina alone. Thus the retina-RPE-choroid culture system can enable the evaluation of interactions between RPE and neural retina, the role of signaling molecules as well the effect of pharmaceuticals on retinal biology.     

Presence of calpain-induced proteolysis in retinal degeneration and dysfunction in a rat model of acute ocular hypertension

The purpose of this study was to determine if calpain-induced proteolysis was associated with retinal degeneration or dysfunction in the rat acute ocular hypertensive model. Acute glaucoma was produced by elevation of IOP to 120 mm Hg for 1 hr. Retinal degeneration was evaluated by H&E staining and apoptosis was determined by TUNEL staining in histologic sections of retina. Electroretinogram (ERG) was carried out to evaluate changes in functionality. Activation of calpains was determined by casein zymography and immunoblotting. Total calcium in retina was measured by atomic absorption spectrophotometry. Proteolysis of alpha-spectrin, tau, cdk5, and p35 (a regulator of cdk5) were evaluated by immunoblotting. The thickness of inner plexiform layer (IPL) and inner nuclear layer (INL), and the number of cells in the ganglion cell layer (GCL) decreased after ocular hypertension. Numerous cells in the INL stained positive for TUNEL and some cells in the outer nuclear layer (ONL) showed TUNEL staining. The a-wave in ERG was temporarily decreased after ocular hypertension and then recovered to normal. In contrast, the b-wave was completely lost. Calpains were activated after ocular hypertension. Activation of calpains was associated with increased calcium in retina. Calpain-dependent proteolysis of alpha-spectrin, tau, and p35 were observed in retina after ocular hypertension. The results suggested that increased calcium and subsequent proteolysis by activated calpains was associated with the death of inner retinal cells due to acute ocular hypertension in the rat model. Calpain inhibitors may be candidate drugs for treatment of retinal degeneration and dysfunction resulting from glaucoma.     

Cell-specific regulation of neuronal production in the larval frog retina

We have previously postulated the existence of a feedback mechanism from differentiated neurons that regulates the production of new neurons. Evidence for such regulatory feedback comes from experiments in which dopamine-containing amacrine cells, ablated in the developing retina by 6-hydroxydopamine (6-OHDA), were up-regulated in their production. To determine whether this is a general phenomenon of the developing retina, the neurotoxin kainic acid (KA) was injected intraocularly in midlarval-stage Rana pipiens tadpoles to produce selective lesions of certain retinal cell types. After periods of 1-21 d, the animals received intraperitoneal injections of 3H-thymidine. Animals were then allowed to survive for periods of up to 3 weeks and were then fixed, the eyes embedded in plastic, sectioned at 3 micron, and processed for autoradiography by standard methods. At the dosage used, the KA produced a 52% decline in the cell density of the inner nuclear layer (INL), a 37% decline in the retinal ganglion cell layer (RGC), and no significant change in the density of cells in the outer nuclear layer (ONL). The 3H-thymidine allowed us to detect any changes in the number of new cells added to the retina after the KA lesion. Within the first week after the KA injection, there was a decrease in the number of 3H-thymidine (3H-Thy)-labeled cells in the lesioned eye as compared to in the control retina; however, KA treatment of slice cultures demonstrated that the toxin does not affect proliferating neuroblasts directly.(ABSTRACT TRUNCATED AT 250 WORDS)     

Selective neuronal survival and upregulation of PCNA in the rat inner retina following transient ischemia

In this study we investigated the extent and time course of neuronal cell death and the regulation of the proliferating cell nuclear antigen (PCNA) in the different retinal cell layers following ischemia-reperfusion injury. Retinal ischemia was induced by controlled elevation of the intraocular pressure for a duration of 60 min. Changes in thickness and cell numbers in the retinal cell layers were analyzed at various time points (1 h to 4 weeks) after reperfusion. In parallel, apoptotic cell death was determined by the TUNEL method and the expression of PCNA analyzed by immunocytochemistry. In addition, we tested whether PCNA is expressed in neurons by double immunocytochemistry. The reduction in thickness was found to be less pronounced in the inner nuclear layer (INL). Correspondingly, cell numbers decreased by only 33% in the inner retina, but by more than 80% in the outer nuclear layer (ONL). Alterations in glial cell numbers did not contribute significantly to postischemic changes in the INL and ONL as assessed by using immunocytochemical markers for microglial and Müller cells. The time course of cell death determined by the TUNEL technique also differed markedly in the retinal layers being rapid and transient in the inner retina but delayed and prolonged in the ONL. PCNA immunoreactivity was undetectable in the normal retina, but was specifically induced in neurons of the inner retina within 1 h after reperfusion and was sustained for at least 4 weeks. We conclude that in contrast to photoreceptors in the ONL, a significant proportion of inner retinal neurons is resistant to ischemic insult induced by transiently increased intraocular pressure and that PCNA may possibly play a role in the selective postischemic survival of these cells.     

Spatial patterning of cholinergic amacrine cells in the mouse retina

The two populations of cholinergic amacrine cells in the inner nuclear layer (INL) and the ganglion cell layer (GCL) differ in their spatial organization in the mouse retina, but the basis for this difference is not understood. The present investigation examined this issue in six strains of mice that differ in their number of cholinergic cells, addressing how the regularity, packing, and spacing of these cells varies as a function of strain, layer, and density. The number of cholinergic cells was lower in the GCL than in the INL in all six strains. The nearest neighbor and Voronoi domain regularity indexes as well as the packing factor were each consistently lower for the GCL. While these regularity indexes and the packing factor were largely stable across variation in density, the effective radius was inversely related to density for both the GCL and INL, being smaller and more variable in the GCL. Consequently, despite the lower densities in the GCL, neighboring cells were more likely to be positioned closer to one another than in the higher-density INL, thereby reducing regularity and packing. This difference in the spatial organization of cholinergic cells may be due to the cells in the GCL having been passively displaced by fascicles of optic axons and an expanding retinal vasculature during development. In support of this interpretation, we show such displacement of cholinergic somata relative to their dendritic stalks and a decline in packing efficiency and regularity during postnatal development that is more severe for the GCL.     

Dlx1, Dlx2, Pax6, Brn3b,and Chx10 homeobox gene expression defines the retinal ganglion and inner nuclear layers of the developing and adult mouse retina

Distal-less homeobox genes are expressed in the developing forebrain. We assessed Dlx gene expression in the developing and adult mouse retina. Dlx1 and Dlx2 are detected in retinal neuroprogenitors by embryonic day (E) 12.5 (Eisenstat et al. [1999] J. Comp. Neurol. 217-237). At E13.5, the expression of four homeodomain proteins, DLX2, BRN3b, PAX6, and CHX10, define distinct yet overlapping domains in the retinal neuroepithelium. By postnatal day (P) 0, DLX2 is expressed in the neuroblastic layer and the ganglion cell layer (GCL) consisting of ganglion and displaced amacrine cells. DLX1 expression resembles DLX2 to P0 but decreases postnatally. In the adult, DLX2 is localized to ganglion, amacrine, and horizontal cells as determined by coexpression with retinal cell-specific markers. There is coincident expression of DLX2 with gamma-aminobutyric acid (GABA), glutamic acid decarboxylase (GAD)65, and GAD67 in the inner nuclear layer (INL) and GCL. In the adult, DLX2 is coexpressed with BRN3b in ganglion cells; PAX6 in amacrine, horizontal, and ganglion cells; and Chx10 in some bipolar cells. We predict that a combinatorial code of these homeobox genes and others specify retinal cell fate. Our results support a possible role for Dlx1 and Dlx2 in inner retinal development and in the terminal differentiation and/or maintenance of INL interneurons and ganglion cells in the adult. The correlation of DLX2 with GABA expression in the mouse retina closely mirrors the relationship of DLX2 to GABAergic neuronal differentiation in the embryonic forebrain, including neocortex, olfactory bulb and hippocampus, signifying a conservation of function of Dlx genes in the developing central nervous system.     

Choline acetyltransferase-immunoreactive neurons in the developing rat retina

The development of cholinergic cells in the rat retina has been examined with immunocytochemistry by using antisera against choline acetyltransferase (ChAT). ChAT-immunoreactive (IR) cells were first detected at embryonic day 17 (E17) in the transitional zone between the neuroblastic layer (NBL) and ganglion cell layer (GCL). At E20, ChAT-IR cells are located exclusively in the GCL. At postnatal day 0 (P0), ChAT immunoreactivity appeared for the first time in cells at the distal margin of the NBL. Two prominent bands of labeled processes were first visible at P3, and by P15, these two bands resembled those of the adult retina. In addition, ChAT immunoreactivity appeared transiently in horizontal cells from P5 to P10. The number of ChAT-IR cells increased steadily up to P15. This resulted in a 93.8-fold increase between E17 and P15 (680-63,800 cells). However, after P15, the number declined by 19% from 63,800 cells at P15 to 51,800 in the adult. At all ages, the spatial density of each ChAT-IR cell population in the central retina was higher than in the periphery. In both central and peripheral regions, the peak density of ChAT-IR cells in the GCL was attained at E20. However, in the INL, the peak densities occurred at P3 in the central region and at P5 in the peripheral region. Up to P15, the soma diameter of ChAT-IR cells in the INL and GCL in each region increased continuously, reaching peak values at P15. Our results demonstrate that ChAT immunoreactivity is expressed in early developmental stages in the rat retina, as in other mammals, and that acetylcholine released from ChAT-IR cells may have neurotrophic functions in retinal maturation.     

Tracer coupling of neurons in the rat retina inner nuclear layer labeled by Fluorogold

A subpopulation of neurons in the inner nuclear layer (INL) of the rat retina were labeled 9-13 weeks after application of Fluorogold (FG) to the superior colliculus. Neurobiotin injection of FG-labeled cells in the INL of flatmounted living retina revealed that these cells consisted of both displaced ganglion cells and a subset of amacrine cells. Fluorogold-labeled amacrine cells in the INL showed tracer coupling to other presumptive amacrine cells in the INL, but there was no evidence of coupling to neurons in the ganglion cell layer (GCL). As the labeling of amacrine cells by FG may be due to gap junction coupling between ganglion and amacrine cells, these data add to the evidence that tracer coupling between these cells can be unidirectional. Some of the FG-labeled displaced ganglion cells in the INL injected with Neurobiotin also showed tracer coupling to neurons in the INL or GCL.     

Two phases of increased cell death in the inner retina following early elimination of the ganglion cell population

Neurons in the inner nuclear layer (INL) of the vertebrate retina undergo considerable programmed cell death during development, but the determinants of this cell death remain largely unknown. The present study examines the role of retinal ganglion cells in support of INL neurons in the developing ferret retina. The retinal ganglion cell population was eliminated by optic nerve transection at postnatal day (P) 2, and the incidence of cell death was examined using terminal deoxytransferase dUTP nick-end labelling (TUNEL) at various ages during the first 3 postnatal weeks. Significant increases in TUNEL-positive cells were observed in the neuroblast layer (NBL) as early as P3, prior to synapse formation within the inner plexiform layer (IPL), and again in the INL at P22, the normal peak of naturally occurring cell death within the ferret's INL. A decrease in TUNEL-positive cells was found in the NBL at P8. These results show three phases of response to the loss of retinal ganglion cells and suggest that cells in the NBL/INL are normally dependent on retinal ganglion cells for their survival. Recent studies have shown that certain populations of retinal neurons are reduced in adult animals that had lost the population of ganglion cells during early development, so the present study also examined when this reduction could first be detected. The number of parvalbumin-immunoreactive amacrine cells was decreased significantly in the NBL of the manipulated eye as early as P8, when we could first label this population, and this difference persisted through adulthood. The fact that cell death in the NBL has already increased within 24 hours of ganglion cell elimination, coupled with the specificity of this effect on the adult complement of INL cell types, shows that cell-cell interactions controlling survival are already highly specific for particular types of retinal neuron early in development Copyright 2001 Wiley-Liss, Inc.     

Development of neuropeptide Y-immunoreactive neurons in the rat retina

Neuropeptide Y-like immunoreactivity (NPY-LI) was examined in the rat retina by radioimmunoassay and immunocytochemistry during prenatal and postnatal development. NPY-LI appears late in gestation (embryonic day [E18]), at which time it is present in small quantities (0.038 +/- 0.005 pm/mg protein) and the NPY-LI is confined to cells in the ganglion cell layer. The concentration of NPY-LI rises steadily over pre- and postnatal development; and on postnatal day 6 (P6), immunoreactive cells first appear in the inner nuclear layer. At eye opening (P13), there is a large increase in NPY-LI (0.207 +/- 0.035 pm/mg protein), and immunoreactive cells can be seen in the innermost row of the inner nuclear layer (INL) as well as in the ganglion cell layer (GCL). As the retina matures, the levels of NPY-LI fall to adult levels (0.080 +/- 0.019 pm/mg protein) and the peptide is confined to two subpopulations of cells, one in the INL and one in the GCL. The transient increase in NPY-LI at eye opening suggests that it may have a role at this time in modulating developing retinal circuitry. This pattern is very different from that of somatostatin-like immunoreactivity which appears earlier in development in high quantities and decreases prior to synaptogenesis and eye opening.     

Distribution and synaptic connectivity of neuropeptide Y-immunoreactive amacrine cells in the rat retina

Neuropeptide Y (NPY) is a potent bioactive peptide that is widely expressed in the nervous system, including the retina. Here we show that specific NPY immunoreactivity was localized to amacrine and displaced amacrine cells in the rat retina. Immunoreactive cells had a regular distribution across the retina and an overall cell density of 280 cells/mm(2) in the inner nuclear layer (INL) and 90 cells/mm(2) in the ganglion cell layer (GCL). In the INL, most immunoreactive cells were characterized by small cell bodies and fine processes that appeared to ramify primarily in stratum 1 of the inner plexiform layer (IPL). A few cells in the INL also ramified in stratum 3 of the IPL. In the GCL, small to medium immunoreactive cells appeared to ramify primarily in stratum 5 of the IPL. A few immunoreactive processes, originating from somata in the INL and processes in the IPL, ramified in the OPL. NPY-immunoreactive cells contained GABA immunoreactivity, and some amacrine cells also contained tyrosine hydroxylase immunoreactivity. NPY-immunostained processes were most frequently presynaptic to nonimmunostained amacrine and ganglion cell processes and postsynaptic to nonimmunostained amacrine cell processes and cone bipolar cell axonal terminals. These findings indicate that NPY immunoreactivity is present in two populations of amacrine cells, one located in the INL and the other in the GCL, and that these cells mainly form synaptic contacts with other amacrine cells. These observations suggest that NPY-immunoreactive cells participate in multiple circuits mediating visual information processing in the inner retina.