Differential regulation of ciliary neurotrophic factor receptor-α expression in all major neuronal cell classes during development of the chick retina

Ciliary neurotrophic factor (CNTF) exerts a multiplicity of effects on a broad spectrum of target cells, including retinal neurons. To investigate how this functional complexity relates to the regulation of CNTF receptor α (CNTFRα) expression, we have studied the developmental expression of the receptor protein in chick retina by using immunocytochemistry. During the course of development, the receptor is expressed in all retinal layers, but three levels of specificity can be observed. First, the expression is regulated temporally with immunoreactivity observed in ganglion cells (embryonic day 8 [E8] to adult), photoreceptor precursors (E8–E12), amacrine cells (E10 to adult), bipolar cells (E12–E18), differentiated rods (E18 to adult), and horizontal cells (adult). Second, expression is restricted to distinct subpopulations of principal retinal neurons: preferentially, large ganglion cells; subpopulations of amacrine cells, including a particular type of cholinergic neuron; a distinctly located type of bipolar cell; and rod photoreceptors. Third, expression exhibits subcellular restriction: it is confined largely to dendrites in mature amacrine cells and is restricted entirely to outer segments in mature rods. These data correlate with CNTF effects on the survival of ganglion cells and mature photoreceptors, the in vitro differentiation of photoreceptor precursors and cholinergic amacrine cells, and the number of bipolar cells in culture described here or in previous studies. Thus, our results demonstrate an exceptional degree of complexity with respect to the regulation of neuronal CNTFRα expression in a defined model system. This suggests that the same signaling pathway is used to mediate a variety of regulatory influences, depending on the developmental stage and cell type. J. Comp. Neurol. 400:244–254, 1998. © 1998 Wiley-Liss, Inc.     

Cellular localization of cyclooxygenase-1 and cyclooxygenase-2 in the normal mouse,rat, and human retina

Prostaglandins, synthesized by cyclooxygenase (COX), regulate diverse neurophysiological actions such as regulation of autonomic responses, transmission of pain, generation of fever, control of sleep-wake cycle, synaptic signaling, and cross-talk between neurons and glia in the central nervous system. Although prostaglandins have been widely studied in the anterior segment tissues of the eye, relatively little is known about prostaglandins in the neural retina. By using immunohistochemistry, we have compared the cellular expression and localization of COX-1 and COX-2 in the normal mouse, rat, and human retina. In the normal mouse retina, COX-1 immunoreactivity is present in the outer segments of photoreceptor cells, horizontal cells, microglia, retinal ganglion cells, and displaced amacrine cells. In the normal rat retina, COX-1 immunoreactivity is present in microglia, retinal ganglion cells, and displaced amacrine cells. In the normal human retina, COX-1 immunoreactivity is present in microglia, astrocytes, retinal ganglion cells, and displaced amacrine cells. In the normal mouse and rat retina, COX-2 immunoreactivity is present in processes of the outer plexiform layer and in certain amacrine cells and retinal ganglion cells. In the normal human retina, COX-2 immunoreactivity is only present in processes of the outer plexiform layer. These results suggest that prostaglandins, synthesized by COX-1 or COX-2, may contribute to normal physiological and homeostatic functions in the retina.     

Spatial density and immunoreactivity of bipolar cells in the macaque monkey retina.

The anatomical substrates of spatial and color vision in the primate retina are investigated by measuring the immunoreactivity and spatial density of bipolar, amacrine and horizontal cells in the inner nuclear layer of the macaque monkey retina. Bipolar cells can be distinguished from amacrine and horizontal cells by their differential immunoreactivity to antisera against glutamate, glycine, GABA, parvalbumin, calbindin (CaBP D-28K), and the L7 protein from mouse cerebellum. The spatial density of bipolar cells is compared to the densities of photoreceptors and ganglion cells at different retinal eccentricities. In the centralmost 2 mm, cone bipolar cells outnumber ganglion cells by about 1.4:1. The density of cone bipolar cells is thus high enough to allow for input to different (parasol and midget) ganglion cell classes by different (diffuse and midget) bipolar cell classes. The density gradient of cone bipolar cells follows closely that of ganglion cells in central retina but falls less steeply in peripheral retina. This suggests that the convergence of cone signals to the receptive fields of ganglion cells in the peripheral retina occurs in the inner plexiform layer. The density of cone bipolar cells is 2.5-4 times that of cones at all eccentricities studied, implying that cone connectivity to bipolar cells remains constant throughout the retina. Different subgroups of bipolar cells are distinguished by their relative immunoreactivity to the different antisera. All rod and cone bipolar cells show moderate to strong glutamate-like immunoreactivity. The bipolar cells that show weak to moderate GABA-like immunoreactivity are also labeled with the antiserum to the L7 protein and are thus identified as rod bipolar cells. Nearly half of all cone bipolar cells showed glycine-like immunoreactivity. The results suggest that the inhibitory neurotransmitter candidates GABA and glycine are segregated respectively in rod and cone bipolar cell pathways. A diffuse, cone bipolar cell type can be identified by the anti-parvalbumin and the anti-calbindin antisera. All horizontal cells show parvalbumin-like immunoreactivity. Nearly all amacrine cells show GABA-like or glycine-like immunoreactivity; a variety of subpopulations also show immunoreactivity to one or more of the other markers used.     

Quantitative analysis of neuronal morphologies in the mouse retina visualized by using a genetically directed reporter

An alkaline phosphatase (AP) reporter has been used to visualize detailed morphologies for all major classes of retinal neurons in the adult mouse. The analysis was performed on retinas in which AP expression was activated by Cre-mediated DNA recombination in a small fraction of cells. Recombination was controlled pharmacologically and, to a first approximation, appears to have occurred randomly. The morphologies of 794 inner retinal neurons have been analyzed by measuring arbor area, stratification level, and neurite branching patterns. When analyzed in this multidimensional parametric space, the cells can be clustered into subgroups by visual inspection and by using the Ward's and K-means algorithms. One application of this cell morphology data set and cluster analysis is as a standard for comparison with the retinas of genetically altered mice. This work illustrates the utility and feasibility of genetically directed marking methods for large-scale surveys of neuronal morphology.     

Retinal anatomy and visual performance in a diurnal cone-rich laboratory rodent, the Nile grass rat (Arvicanthis niloticus)

Unlike laboratory rats and mice, muridae of the Arvicanthis family (A. ansorgei and A. niloticus) are adapted to functioning best in daylight. To date, they have been used as experimental models mainly in studies of circadian rhythms. However, recent work aimed at optimizing photoreceptor-directed gene delivery vectors (Khani et al. [2007] Invest Ophthalmol Vis Sci 48:3954-3961) suggests their potential usefulness for studying retinal pathologies and therapies. In the present study we analyzed the retinal anatomy and visual performance of the Nile grass rat (A. niloticus) using immunohistofluorescence and the optokinetic response (OKR). We found that approximately 35-40% of photoreceptors are cones; that many neural features of the inner retina are similar to those in other diurnal mammals; and that spatial acuity, measured by the OKR, is more than two times that of the usual laboratory rodents. These observations are consistent with the known diurnal habits of this animal, and further support its pertinence as a complementary model for studies of structure, function, and pathology in cone-rich mammalian retinae.     

Bipolar cells of the mouse retina: a gene gun, morphological study

One of the key elements concerning our understanding of the organization of the mouse retina is the complete classification of the various types of bipolar cells. With the present study, we tried to contribute to this important issue. Unfortunately, most of the antibodies that stain specifically bipolar cells in the retina of other mammals hardly work for the retina of the mouse. We succeeded in overcoming this limitation by using a relatively novel technique based on the gene gun transfer of fluorescent dyes to cells. Hence, we were able to stain a considerable number of bipolar cells that could be characterized according to morphological and comparative criteria. We also performed a complete morphometric analysis of a subset of bipolar cells stained by anti-neurokinin-3 receptor antibodies. We found nine types of cone bipolar cells and one type of rod bipolar cell; these data are consistent with the findings of previous studies on the retinas of other mammals, such as rabbits, rats, and monkeys and with a recent study based on the mouse retina (Ghosh et al. [2004] J Comp Neurol 469:70-82). Our results also confirm the existence of a common structural similarity among mammalian retinas. It remains to be elucidated what is exactly the functional role of the various types of cone bipolar cells and what is the specific contribution they provide to the perception of a given visual stimulus. Most probably, each bipolar cell type constitutes a specialized channel for the computation of a selected component of the visual stimulus. More complex signal coding, involving the coordinated activity of various types of bipolar cells, could also be postulated, as it has been shown for ganglion cells (Meister [1996] Proc Natl Acad Sci U S A 93:609-614).     

The dopamine D2 receptor subfamily in rat retina: ultrastructural immunogold and in situ hybridization studies

Dopamine, a major neurotransmitter in the vertebrate retina, is released from interplexiform cells and a restricted subset of amacrine cells. Dopamine effects vary between different retinal cell types, most likely due to differences in cell-specific receptor subtype expression. Identification of cells expressing receptors of the D2-subfamily (D2R, D3R, D4R) on a light microscopical level has rendered equivocal results, and no information is as yet available concerning the subcellular distribution of receptor protein. In the present study, D2R and D2/3R subtype-specific antisera, and D2R-, D3R- and D4R-specific oligonucleotide probes were used for ultrastructural and in situ hybridization analyses of the receptor subtype distribution in the rat retina. Light and electron microscopy showed that in addition to the known localization of intense D2R-immunoreactivity in all dopaminergic cells immunoreactive for tyrosine hydroxylase (TH), homogeneous, less intense D2R-immunoreactivity was also seen throughout the inner plexiform layer (IPL). Ultrastructurally, many additional amacrine cell processes devoid of TH-immunoreactivity at all levels of the inner plexiform layer were immunoreactive. D2R-immunoreactivity was found mainly on intracellular vesicles, and immunoreactivity associated with the plasma membrane was always extrasynaptic. No D2R-immunoreactivity was found in amacrine cell somata postsynaptic to the so-called dopaminergic 'ring endings'. Many D2R-mRNA reactive cells were observed throughout the inner nuclear layer. Morphologically, labelled cells resemble amacrines and bipolars but not horizontal cells. Reactivity with splice variant-specific oligonucleotide probes suggested that the D2LR variant is the predominant if not the only D2R isoform in the rat retina. D2R-mRNA reactivity was not observed in other retinal layers, in particular not in photoreceptor inner segments, which displayed D4R-mRNA reactivity. D3R-mRNA reactivity was not detected. The results indicate that D2-like responses are mediated through the D2R subtype, by an autoreceptor mechanism in dopaminergic cells, and by volume transmission in non-dopaminergic cells of the inner retina. D2-like responses in photoreceptors probably represent D4R activation.     

Bipolar cell-photoreceptor connectivity in the zebrafish (Danio rerio) retina

Bipolar cells convey luminance, spatial, and color information from photoreceptors to amacrine and ganglion cells. We studied the photoreceptor connectivity of 321 bipolar cells in the adult zebrafish retina. 1,1′‐Dioctadecyl‐3,3,3′,3′‐tetramethylindocarbocyanine perchlorate (DiI) was inserted into whole‐mounted transgenic zebrafish retinas to label bipolar cells. The photoreceptors that connect to these DiI‐labeled cells were identified by transgenic fluorescence or their positions relative to the fluorescent cones, as cones are arranged in a highly ordered mosaic: rows of alternating blue‐ (B) and ultraviolet‐sensitive (UV) single cones alternate with rows of red‐ (R) and green‐sensitive (G) double cones. Rod terminals intersperse among cone terminals. As many as 18 connectivity subtypes were observed, 9 of which—G, GBUV, RG, RGB, RGBUV, RGRod, RGBRod, RGBUVRod, and RRod bipolar cells—accounted for 96% of the population. Based on their axon terminal stratification, these bipolar cells could be further subdivided into ON, OFF, and ON–OFF cells. The dendritic spread size, soma depth and size, and photoreceptor connections of the 308 bipolar cells within the nine common connectivity subtypes were determined, and their dendritic tree morphologies and axonal stratification patterns compared. We found that bipolar cells with the same axonal stratification patterns could have heterogeneous photoreceptor connectivity whereas bipolar cells with the same dendritic tree morphology usually had the same photoreceptor connectivity, although their axons might stratify on different levels. J. Comp. Neurol. 520:3786–3802, 2012. © 2012 Wiley Periodicals, Inc.     

Structural and functional remodeling in the retina of a mouse with a photoreceptor synaptopathy: plasticity in the rod and degeneration in the cone system

Knowledge about the plastic and regenerative capacity of the retina is of key importance for therapeutic approaches to restore vision in patients who suffer from degenerative retinal diseases. In the retinae of mice, mutant for the presynaptic scaffolding protein Bassoon, signal transfer at photoreceptor ribbon synapses is disturbed due to impaired ribbon attachment to the active zone. In a long-term study we observed, with light and electron microscopic immunocytochemistry and electroretinographic recordings, two overlapping events in the Bassoon mutant retina, i.e. loss of photoreceptor synapses in the outer plexiform layer, and structural remodeling and formation of ectopic photoreceptor synapses in the outer nuclear layer, a region usually devoid of synapses. Formation of ectopic synaptic sites starts around the time when photoreceptor synaptogenesis is completed in wild-type mice and progresses throughout life. The result is a dense plexus of ectopic photoreceptor synapses with significantly altered but considerable synaptic transmission. Ectopic synapse formation is led by the sprouting of horizontal cells followed by the extension of rod bipolar cell neurites that fasciculate with and grow along the horizontal cell processes. Although only the rod photoreceptors and their postsynaptic partners show structural and functional remodeling, our study demonstrates the potential of the retina for long-lasting plastic changes.     

Immunocytochemical localization of three vesicular glutamate transporters in the cat retina

Vesicular transporters play an essential role in the packaging of glutamate for synaptic release and so are of particular importance in the retina, where glutamate serves as the neurotransmitter for photoreceptors, bipolar cells, and ganglion cells. In the present study, we have examined the distribution of the three known isoforms of vesicular glutamate transporter (VGLUT) in the cat retina. VGLUT1 was localized to all photoreceptor and bipolar cells, whereas VGLUT2 was found in ganglion cells. This basic pattern of complementary distribution for the two transporters among known populations of glutamatergic cells is similar to previous findings in the brain and spinal cord. However, the axon terminals of S-cone photoreceptors were found to express both VGLUT1 and VGLUT2 and some ganglion cells labeled for both VGLUT2 and VGLUT3. Such colocalizations suggest the existence of dual modes of regulation of vesicular glutamate transport in these neurons. Staining for VGLUT2 was also present in a small number of varicose processes, which were seen to ramify throughout the inner plexiform layer. These fibers may represent axon collaterals of ganglion cells. The most prominent site of VGLUT3 immunoreactivity was in a population of amacrine cells; the axon terminals of B-type horizontal cells were also labeled at their contacts with rod spherules. The presence of the VGLUT3 transporter at sites not otherwise implicated in glutamate release may indicate novel modes of glutamate signaling or additional roles for the transporter molecule.     

Development of cholinergic amacrine cells is visual activity-dependent in the postnatal mouse retina

In the present study, we used immunocytochemistry to study the temporal and spatial arrangement of mouse cholinergic amacrine cells during postnatal retinal development under normal light/dark cycles and during visual deprivation. Choline acetyltransferase (ChAT)-immunolabeled cells were detected in the neuroblastic layer (NBL) and in the ganglion cell layer (GCL) at postnatal day 0 (P0). Between P3-5, two characteristic cholinergic bands were clearly identified in the inner plexiform layer (IPL). The signal intensity of somas and processes progressively increased over the first 2 postnatal weeks. Around eye opening at P12, cholinergic neurons were mature-like. This early developmental process was not altered by visual deprivation. After eye opening, the space between the two cholinergic bands increased continuously and the spatial regularity index changed constantly, indicating that the cholinergic neurons possibly underwent refinement during later postnatal development. The changes occurring following eye opening were retarded by visual deprivation. The morphologies of photoreceptors, horizontal cells, recoverin-positive OFF-cone bipolar cells, rod bipolar cells, dopaminergic amacrine cells, and Müller cells appeared normal. Their stratification in the outer plexiform layer (OPL) and the IPL was not affected by visual deprivation. However, glial cells grew vertically across the entire thickness of dark-reared retinas. Our results suggest that the development of cholinergic neurons before eye opening is independent of the lighting conditions. Their development after eye opening is greatly impeded by visual deprivation. This visual activity-dependent phase of development may be a critical period for the maturation and synaptic wiring of cholinergic amacrine cells in the mammalian retina.     

Manipulation of synaptic sign and strength with divalent cations in the vertebrate retina: pushing the limits of tonic, chemical neurotransmission

At the first synaptic level of the vertebrate retina, photoreceptor light responses are transmitted to second order neurones through a chemical synapse based on a tonic release of neurotransmitter modulated by graded changes of presynaptic potential. The possibility that such synapses could work through a Ca2+-independent process had been proposed by previous authors, based on the persistence of transmission process in low Ca2+ media containing Co2+ or Ni2+ ions. Recently, we were able to explain these results within the framework of the classical calcium-hypothesis of synaptic transmission by taking into account the modifications of presynaptic surface potential brought about by changes of divalent cation concentrations. Here we report data showing how a surface-charge hypothesis could account for several apparently paradoxical effects of divalent cation manipulations such as: the enhancement of neurotransmitter release induced by low Ca2+ media; the transmission "unblocking" effect of Zn2+, Co2+ and Ni2+; and the reversal of transmission polarity induced by application of low Ca2+ media containing Cd2+ or Mg2+ ions.     

GABA-like immunoreactivity in the macaque monkey retina: a light and electron microscopic study

The distribution of GABA-like immunoreactivity in the macaque monkey retina was studied by using postembedding techniques on semithin and ultrathin sections. At the light microscopic level, both inner and outer plexiform layers showed strong GABA-like immunoreactivity in the central retina. All the horizontal cells, some bipolar cells, 30-40% of amacrine cells, occasional interplexiform cells, and practically all displaced amacrine cells were labeled. In the peripheral retina (beyond 5 mm eccentricity), the outer plexiform layer and the horizontal cells were not labeled, but all other cell types showed the same labeling pattern as in the central retina. Synapses of the inner plexiform layer involving a pre- or postsynaptic GABA-labeled process were studied electron microscopically. Synapses involving a GABA-labeled presynaptic amacrine cell process made up 80% of the synapses observed. These GABA-labeled amacrine processes synapsed onto amacrine, bipolar, and ganglion cell processes as well as onto amacrine and ganglion cell bodies. Synapses involving a postsynaptic GABA-labeled process made up 20% of the synapses studied. The GABA-like immunoreactive processes were postsynaptic to bipolar cells at the dyads and to amacrine cells at conventional synapses.     

Specificity of the horizontal cell‐photoreceptor connections in the zebrafish (Danio rerio) retina

Horizontal cells (HCs) are involved in establishing the center‐surround receptive field organization of photoreceptor and bipolar cells. In many species, HCs respond differentially to colors and may play a role in color vision. An earlier study from our laboratory suggested that four types of HCs exist in the zebrafish retina: three cone HCs (H1, H2 and H3) and one rod HC. In this study, we describe their photoreceptor connections. Cones are arranged in a mosaic in which rows of alternating blue (B)‐ and ultraviolet (UV)‐sensitive single cones alternate with rows of red (R)‐ and green (G)‐sensitive double cones; the G cones are adjacent to UV cones and B cones adjacent to R cones. Two small‐field (H1 and H2) and two large‐field (H3 and rod HC) cells were observed. The cone HC dendritic terminals connected to cones with single boutons, doublets, or rosettes, whereas the rod HCs connected to rods with single boutons. The single boutons/doublets/rosettes of cone HCs were arranged in double rows separated by single rows for H1 cells, in pairs and singles for H2 cells, and in a rectilinear pattern for H3 cells. These connectivity patterns suggest that H1 cells contact R, G, and B cones, H2 cells G, B, and UV cones, and H3 cells B and UV cones. These predictions were confirmed by applying the DiI method to SWS1‐GFP retinas whose UV cones express green fluorescent protein. Each rod HC was adjacent to the soma or axon of a DiI‐labeled cone HC and connected to 50–200 rods. J. Comp. Neurol. 516:442–453, 2009. © 2009 Wiley‐Liss, Inc.     

Expression of vesicular glutamate transporter 1 in the mouse retina reveals temporal ordering in development of rod vs. cone and ON vs. OFF circuits

Glutamatergic transmission is crucial to the segregation of ON and OFF pathways in the developing retina. The temporal sequence of maturation of vesicular glutamatergic transmission in rod and cone photoreceptor and ON and OFF bipolar cell terminals is currently unknown. Vesicular glutamate transporters (VGLUTs) that load glutamate into synaptic vesicles are necessary for vesicular glutamatergic transmission. To understand better the formation and maturation of glutamatergic transmission in the rod vs. cone and ON vs. OFF pathways of the retina, we examined the developmental expression of VGLUT1 and VGLUT2 immunocytochemically in the mouse retina. Photoreceptor and bipolar cell terminals showed only VGLUT1-immunoreactivity (-IR); no VGLUT2-IR was present in any synapses of the developing or adult retina. VGLUT1-IR was first detected in cone photoreceptor terminals at postnatal day 2 (P2), several days before initiation of ribbon synapse formation at P4-P5. Rod terminals showed VGLUT1-IR by P8, when they invade the outer plexiform layer (OPL) and initiate synaptogenesis. Developing OFF bipolar cell terminals showed VGLUT1-IR around P8, 2-3 days after bipolar terminals were first identified in the inner plexiform layer (IPL) by labeling for the photoreceptor and bipolar cell terminal marker, synaptic vesicle protein 2B. Although terminals of ON bipolar cells were present in the IPL by P6-P8, most did not show VGLUT1-IR until P8-P10 and increased dramatically from P12. These data suggest a hierarchical development of glutamatergic transmission in which cone circuits form prior to rod circuits in both the OPL and IPL, and OFF circuits form prior to ON circuits in the IPL.     

GABA-like immunoreactivity in the cat retina: light microscopy

Semithin sections of the cat retina were stained with antibodies against GABA conjugated to bovine serum albumin with glutaraldehyde. Labelled cells were visualized by means of the peroxidase-antiperoxidase method. In the outer plexiform layer both A- and B-type horizontal cells and the B-type axon terminal system expressed GABA-like immunoreactivity. Approximately 25-30% of all amacrine cells and the whole inner plexiform layer were heavily labelled. Two types of putative GABA-ergic interplexiform cells could be distinguished. One of them also expressed tyrosine-hydroxylase-like immunoreactivity. A few bipolar cells were also GABA-immunolabelled. GABA-like immunoreactivity and 3H-muscimol uptake were colocalized in 90% of the amacrine cells labelled. However, horizontal cells did not accumulate 3H-muscimol.     

Morphological analysis of the blue cone pathway in the retina of a New World monkey,the marmoset Callithrix jacchus

We have studied the components of the short wavelength-sensitive (SWS or “blue”) cone pathway in the retina of a New World primate, the marmoset Callithrix jacchus. Of particular interest was the small bistratified ganglion cell, which has been identified in macaque monkey to be the morphological substrate of the blue-ON cell (Dacey and Lee [1994] Nature 367:731–735). Small bistratified cells were intracellularly filled with Neurobiotin in an in vitro retinal wholemount preparation. Their morphology, size, and level of dendritic stratification were similar to their homologues in macaque and human retina. A number of different antibodies were applied to vertical cryostat sections, some of which were cut through the processes of injected small bistratified or parasol ganglion cells. We used antibodies against cholecystokinin (CCK) to label blue cone bipolar cells, and antibodies against the human SWS cone photopigment to label SWS cones. Double-labelled preparations showed that blue cone bipolar cell dendrites contact SWS cone pedicles, and the inner dendrites of the small bistratified cell are costratified with the axon terminals of blue cone bipolar cells. A monoclonal antibody against calbindin was used to label a subpopulation of bipolar cells that stratifies in the outer half of the inner plexiform layer. The axon terminals of these bipolar cells occasionally cross the outer dendrites of small bistratified cells and show extensive costratification with the dendrites of OFF parasol cells. We conclude that an SWS cone pathway with similar connectivity is a preserved feature of the primate visual system. J. Comp. Neurol. 379:211–225, 1997. © 1997 Wiley-Liss, Inc.     

Synaptogenesis and synaptic protein localization in the postnatal development of rod bipolar cell dendrites in mouse retina

Retinal responses to photons originate in rod photoreceptors and are transmitted to the ganglion cell output of the retina through the primary rod bipolar pathway. At the first synapse of this pathway, input from multiple rods is pooled into individual rod bipolar cells. This architecture is called convergence. Convergence serves to improve sensitivity of rod vision when photons are sparse. Establishment of convergence depends on the development of a proper complement of dendritic tips and transduction proteins in rod bipolar cells. How the dendrites of rod bipolar cells develop and contact the appropriate number of rods is unknown. To answer this question we visualized individual rod bipolar cells in mouse retina during postnatal development and quantified the number of dendritic tips, as well as the expression of transduction proteins within dendrites. Our findings show that the number of dendritic tips in rod bipolar cells increases monotonically during development. The number of tips at P21, P30, and P82 exceeds the previously reported rod convergence ratios, and the majority of these tips are proximal to a presynaptic rod release site, suggesting more rods provide input to a rod bipolar cell. We also show that dendritic transduction cascade members mGluR6 and TRPM1 appear in tips with different timelines. These finding suggest that (a) rod bipolar cell dendrites elaborate without pruning during development, (b) the convergence ratio between rods and rod bipolar cells may be higher than previously reported, and (c) mGluR6 and TRPM1 are trafficked independently during development.     

Types of bipolar cells in the mouse retina

We studied the morphology of bipolar cells in fixed vertical tissue sections (slices) of the mouse retina by injecting the cells with Lucifer Yellow and Neurobiotin. Nine different cone bipolar cell types and one rod bipolar cell type were distinguished. The major criteria for classifying the cells were the branching pattern and stratification level of their axon terminals in the inner plexiform layer (IPL). To assess this, the IPL was subdivided into five strata of equal width. The slices were immunostained for calretinin, which labels three horizontal bands serving as a standard measure for the precise localization of the axon terminals. Immunostaining the retina with antibodies against the G-protein Ggamma13, a marker for ON-bipolar cells, made it possible to separate OFF- and ON-bipolar cells. At least two OFF-cone bipolar cells (Types 1 and 2) were immunolabeled with antibodies against the neurokinin 3 receptors (NK3R). A further OFF- and an ON-cone bipolar cell (Types 3 and 5) were immunostained with antibodies against the calcium-binding protein CaB5. The bipolar cell types described here were compared with previous schemes of rat and primate bipolar cells. Homologous types between the three species are discussed.