Horizontal Cell Layer Research Articles

Comparative ultrastructure of the olfactory system in the East African root rat (Tachyoryctes splendens) and the naked mole rat (Heterocephalus glaber).

The ultrastructure of the olfactory system of most fossorial rodents remains largely unexplored. This study sought to investigate the functional structure of the olfactory mucosa and olfactory bulb of two species of fossorial rodents that have distinct behaviour and ecology, the East African root rat (RR) and the naked mole rat (NMR). Transmission electron microscopy and scanning electron microscopy were employed. The basic ultrastructural design of the olfactory system of the two species was largely comparable. In both species, the olfactory mucosa comprised an olfactory epithelium and an underlying lamina propria. The olfactory epithelium revealed olfactory knobs, cilia and microvilli apically and sustentancular cells, olfactory receptor neurons and basal cells in the upper, middle and basal zones, respectively. The lamina propria was constituted by Bowman's glands, olfactory nerve bundles and vasculature supported by loose connective tissue. Within the olfactory bulb, intracellular and extracellular structures including cell organelles, axons and dendrites were elucidated. Notable species differences were observed in the basal zone of the olfactory epithelium and on the luminal surface of the olfactory mucosa. The basal zone of the olfactory epithelium of the RR consisted of a single layer of flattened electron-dense horizontal basal cells while the NMR had juxtaposed electron-dense and electron-lucent heterogenous cells, an occurrence seen as being indicative of quiescent and highly proliferative states of the olfactory epithelia in the two species, respectively. The olfactory epithelial surface of the NMR comprised an elaborate cilia network that intertwined extensively forming loop-like structures whereas in the RR, the surface was rugged and consisted of finger-like processes and irregular masses. With gross and histological studies showing significant differences in the olfactory structures of the two species, these findings are a further manifestation that the olfactory system of the RR and the NMR have evolved differently to reflect their varied olfactory functional needs.

Amoeboid-like migration ensures correct horizontal cell layer formation in the developing vertebrate retina.

Migration of cells in the developing brain is integral for the establishment of neural circuits and function of the central nervous system. While migration modes during which neurons employ predetermined directional guidance of either preexisting neuronal processes or underlying cells have been well explored, less is known about how cells featuring multipolar morphology migrate in the dense environment of the developing brain. To address this, we here investigated multipolar migration of horizontal cells in the zebrafish retina. We found that these cells feature several hallmarks of amoeboid-like migration that enable them to tailor their movements to the spatial constraints of the crowded retina. These hallmarks include cell and nuclear shape changes, as well as persistent rearward polarization of stable F-actin. Interference with the organization of the developing retina by changing nuclear properties or overall tissue architecture hampers efficient horizontal cell migration and layer formation showing that cell-tissue interplay is crucial for this process. In view of the high proportion of multipolar migration phenomena observed in brain development, the here uncovered amoeboid-like migration mode might be conserved in other areas of the developing nervous system.

Senescence-associated β-galactosidase activity in the developing avian retina.

Senescence-associated β-galactosidase (SA-β-GAL) histochemistry is the most commonly used biomarker of cellular senescence. These SA-β-GAL-positive cells are senescent embryonic cells that are usually removed by apoptosis from the embryo, followed by macrophage-mediated clearance. Some authors have proposed that SA-β-GAL activity in differentiated neurons from young and adult mammals cannot be uniquely attributed to cell senescence, whether in vivo or in vitro. Using the developing visual system of the chicken as a model, the present study found that SA-β-GAL detected in the developing retina corresponded to lysosomal β-galactosidase activity, and that SA-β-GAL activity did not correlate with the chronotopographical distribution of apoptotic cells. However, SA-β-GAL staining in the undifferentiated retina coincided with the appearance of early differentiating neurons. In the laminated retina, SA-β-GAL staining was concentrated in the ganglion, amacrine, and horizontal cell layers. The photoreceptors and pigment epithelial cells also exhibited SA-β-GAL activity throughout retinal development. We have also found that SA-β-GAL staining strongly correlated p21 immunoreactivity. In conclusion, the results clearly show that SA-β-GAL activity cannot be regarded as a specific marker of senescence during retinal development, and that it is mainly expressed in subpopulations of postmitotic neurons, which are nonproliferative cells, even at early stages of cell differentiation.

Stochastic single cell migration leads to robust horizontal cell layer formation in the vertebrate retina.

Developmental programs that arrange cells and tissues into patterned organs are remarkably robust. In the developing vertebrate retina, for example, neurons reproducibly assemble into distinct layers giving the mature organ its overall structured appearance. This stereotypic neuronal arrangement, termed lamination, is important for efficient neuronal connectivity. Although retinal lamination is conserved in many vertebrates, including humans, how it emerges from single cell behaviour is not fully understood. To shed light on this issue, we here investigated the formation of the retinal horizontal cell layer. Using in vivo light sheet imaging of the developing zebrafish retina, we generated a comprehensive quantitative analysis of horizontal single cell behaviour from birth to final positioning. Interestingly, we find that all parameters analysed, including cell cycle dynamics, migration paths and kinetics, as well as sister cell dispersal, are very heterogeneous. Thus, horizontal cells show individual non-stereotypic behaviour before final positioning. Yet these initially variable cell dynamics always generate the correct laminar pattern. Consequently, our data show that the extent of single cell stochasticity in the lamination of the vertebrate retina is underexplored.

Comparative localization of cystathionine beta synthases and cystathionine gamma lyase in canine, non-human primate and human retina

Chronic exposure of the retina to light and high concentrations of polyunsaturated fatty acid in photoreceptor cells make this tissue susceptible to oxidative damage. As retinal degenerative diseases are associated with photoreceptor degeneration, the antioxidant activity of both hydrogen sulfide (H2S) and glutathione (GSH) may play an important role in ameliorating disease progression. H2S production is driven by cystathionine-γ-lyase (CSE) and cystathionine-β–synthase (CBS), the key enzymes that also drive transsulfuration pathway (TSP) necessary for GSH production. As it is currently unclear whether localized production of either H2S or GSH contributes to retinal homeostasis, we undertook a comparative analysis of CBS and CSE expression in canine, non-human primate (NHP) and human retinas to determine if these antioxidants could play a regulatory role in age-related or disease-associated retinal degeneration. Retinas from normal dogs, NHPs and humans were used for the study. Laser capture microdissection (LCM) was performed to isolate individual layers of the canine retina and analyze CBS and CSE gene expression by qRT-PCR. Immunohistochemistry and western blotting were performed for CBS and CSE labeling and protein expression in dog, NHP, and human retina, respectively. Using qRT-PCR, western blot, and immunohistochemistry (IHC), we showed that CBS and CSE are expressed in the canine, NHP, and human retina. IHC results from canine retina demonstrated increased expression levels of CBS but not CSE with post-developmental aging. IHC results also showed non-overlapping localization of both proteins with CBS presenting in rods, amacrine, horizontal, and nerve fiber cell layers while CSE was expressed by RPE, cones and Mϋller cells. Finally, we demonstrated that these enzymes localized to all three layers of canine, NHP and human retina: photoreceptors, outer plexiform layer (OPL) and notably in the ganglion cells layer/nerve fiber layer (GCL/NFL). QRT-PCR performed using RNA extracted from tissues isolated from these cell layers using laser capture microdissection (LCM) confirmed that each of CBS and CSE are expressed equally in these three layers. Together, these findings reveal that CSE and CBS are expressed in the retina, thereby supporting further studies to determine the role of H2S and these proteins in oxidative stress and apoptosis in retinal degenerative diseases.

Notch Signaling Regulates Lgr5+ Olfactory Epithelium Progenitor/Stem Cell Turnover and Mediates Recovery of Lesioned Olfactory Epithelium in Mouse Model.

The Notch signaling pathway regulates stem cell proliferation and differentiation in multiple tissues and organs, and is required for tissue maintenance. However, the role of Notch in regulation of olfactory epithelium (OE) progenitor/stem cells to maintain tissue function is still not clear. A recent study reported that leucine-rich repeat-containing G-protein-coupled receptor 5 (Lgr5) is expressed in globose basal cells (GBCs) localized in OE. Through lineage tracing in vivo, we found that Lgr5+ cells act as progenitor/stem cells in OE. The generation of daughter cells from Lgr5+ progenitor/stem cells is delicately regulated by the Notch signaling pathway, which not only controls the proliferation of Lgr5+ cells and their immediate progenies but also affects their subsequent terminal differentiation. In conditionally cultured OE organoids in vitro, inhibition of Notch signaling promotes neuronal differentiation. Besides, OE lesion through methimazole administration in mice induces generation of more Notch1+ cells in the horizontal basal cell (HBC) layer, and organoids derived from lesioned OE possesses more proliferative Notch1+ HBCs. In summary, we concluded that Notch signaling regulates Lgr5+ GBCs by controlling cellular proliferation and differentiation as well as maintaining epithelial cell homeostasis in normal OE. Meanwhile, Notch1 also marks HBCs in lesioned OE and Notch1+ HBCs are transiently present in OE after injury. This implies that Notch1+ cells in OE may have dual roles, functioning as GBCs in early development of OE and HBCs in restoring the lesioned OE. Stem Cells 2018;36:1259-1272.

The Zebrafish Anillin-eGFP Reporter Marks Late Dividing Retinal Precursors and Stem Cells Entering Neuronal Lineages.

Monitoring cycling behaviours of stem and somatic cells in the living animal is a powerful tool to better understand tissue development and homeostasis. The tg(anillin:anillin-eGFP) transgenic line carries the full-length zebrafish F-actin binding protein Anillin fused to eGFP from a bacterial artificial chromosome (BAC) containing Anillin cis-regulatory sequences. Here we report the suitability of the Anillin-eGFP reporter as a direct indicator of cycling cells in the late embryonic and post-embryonic retina. We show that combining the anillin:anillin-eGFP with other transgenes such as ptf1a:dsRed and atoh7:gap-RFP allows obtaining spatial and temporal resolution of the mitotic potentials of specific retinal cell populations. This is exemplified by the analysis of the origin of the previously reported apically and non-apically dividing late committed precursors of the photoreceptor and horizontal cell layers.

Specific connectivity between photoreceptors and horizontal cells in the zebrafish retina.

The functional and morphological connectivity between various horizontal cell (HC) types (H1, H2, H3, and H4) and photoreceptors was studied in zebrafish retina. Since HCs are strongly coupled by gap junctions and feedback from HCs to photoreceptors depends strongly on connexin (Cx) hemichannels, we characterized the various HC Cxs (Cx52.6, Cx52.7, Cx52.9, and Cx55.5) in Xenopus oocytes. All Cxs formed hemichannels that were conducting at physiological membrane potentials. The Cx hemichannels differed in kinetic properties and voltage dependence, allowing for specific tuning of the coupling of HCs and the feedback signal from HCs to cones. The morphological connectivity between HC layers and cones was determined next. We used zebrafish expressing green fluorescent protein under the control of Cx promoters. We found that all HCs showed Cx55.5 promoter activity. Cx52.7 promoter activity was exclusively present in H4 cells, while Cx52.9 promoter activity occurred only in H1 cells. Cx52.6 promoter activity was present in H4 cells and in the ventral quadrant of the retina also in H1 cells. Finally, we determined the spectral sensitivities of the HC layers. Three response types were found. Monophasic responses were generated by HCs that contacted all cones (H1 cells), biphasic responses were generated by HCs that contacted M, S, and UV cones (H2 cells), and triphasic responses were generated by HCs that contacted either S and UV cones (H3 cells) or rods and UV cones (H4 cells). Electron microscopy confirms that H4 cells innervate cones. This indicates that rod-driven HCs process spectral information during photopic and luminance information during scotopic conditions.

A Retinal Mechanism Inspired Color Constancy Model.

In this paper, we propose a novel model for the computational color constancy, inspired by the amazing ability of the human vision system (HVS) to perceive the color of objects largely constant as the light source color changes. The proposed model imitates the color processing mechanisms in the specific level of the retina, the first stage of the HVS, from the adaptation emerging in the layers of cone photoreceptors and horizontal cells (HCs) to the color-opponent mechanism and disinhibition effect of the non-classical receptive field in the layer of retinal ganglion cells (RGCs). In particular, HC modulation provides a global color correction with cone-specific lateral gain control, and the following RGCs refine the processing with iterative adaptation until all the three opponent channels reach their stable states (i.e., obtain stable outputs). Instead of explicitly estimating the scene illuminant(s), such as most existing algorithms, our model directly removes the effect of scene illuminant. Evaluations on four commonly used color constancy data sets show that the proposed model produces competitive results in comparison with the state-of-the-art methods for the scenes under either single or multiple illuminants. The results indicate that single opponency, especially the disinhibitory effect emerging in the receptive field's subunit-structured surround of RGCs, plays an important role in removing scene illuminant(s) by inherently distinguishing the spatial structures of surfaces from extensive illuminant(s).

Pax6 expression during retinogenesis in sharks: comparison with markers of cell proliferation and neuronal differentiation

Pax6 is a highly conserved transcription factor that appears involved in the entire process of retinogenesis, including maintenance of proliferation of retinal progenitors and differentiation of particular neuron fates. To gain insight into the retinogenesis in fish, we study the dynamics of Pax6 expression in the developing and mature retina of two sharks that inhabit in particular environments, and compare it with the dynamics of a marker of cell proliferation (proliferating cell nuclear antigen, PCNA) and markers of neuronal differentiation, such as glutamic acid decarboxylase (GAD), calretinin (CR), tyrosine-hydroxylase, and serotonin (5-HT). Our results reveal that Pax6 is expressed in PCNA-immunoreactive cells within the nonlayered retina, suggesting a role for Pax6 in proliferating progenitors. Pax6 expression decays as development proceeds and eventually remains in some postmitotic cells, which points to additional roles of Pax6 following neurogenesis. Double immunofluorescence reveals Pax6/CR colocalization in the ganglion cell layer, Pax6/5-HT in the inner part of the inner nuclear layer (INLi), and Pax6/GAD in the INLi and horizontal cell layer. Our results suggest that Pax6 may contribute to neuron diversification in the neural retina.

Genesis of rods in the zebrafish retina occurs in a microenvironment provided by polysialic acid‐expressing Müller glia

Polysialic acid (polySia) is a posttranslational modification of the neural cell adhesion molecule NCAM, which in the vertebrate brain is dynamically regulated during development and crucially involved in developmental and adult neurogenesis. In the fish retina, new neurons are persistently generated, but the possible contribution of polySia has not yet been addressed. Here we used immunohistochemistry with NCAM- and polySia-specific antibodies to study spatiotemporal expression patterns of NCAM and polySia in the developing and mature zebrafish retina. As early as 2.3 days postfertilization (dpf), NCAM but not polySia was detected on cell somata and fibers of the developing retina. At 4.3 dpf polySia immunoreactivity first appeared in the ventral retina and was localized to the nascent outer nuclear layer (ONL). In mature zebrafish, polySia immunoreactivity in the ONL extended to the entire retina. Colocalization with rhodopsin-EGFP in transgenic zebrafish or the Müller glia-specific protein cellular retinaldehyde-binding protein (CRALBP) revealed that polySia immunoreactivity was confined to the compartment of radial Müller glia processes crossing the ONL and to a small band of processes positioned proximal to the horizontal cell layer of the mature retina. As shown by 5-bromo-2-deoxyuridine (BrdU) labeling, both newly generated rod precursors within the mature ONL and precursors of the marginal zone were polySia-negative. Thus, polySia-negative rod precursors of the mature zebrafish retina face a polySia-NCAM-positive microenvironment presented by radial Müller glia. In view of the prominent role of polySia in other neurogenic systems, this pattern indicates that polySia provides environmental cues that are relevant for the generation of new rods.

Preoptic FMRF-amide-like immunoreactive projections to the retina in the lamprey ( Lampetra fluviatilis)

A centrifugal visual system showing FMRF-amide-like immunoreactivity has been demonstrated in Lampetra fluviatilis by using immunocytochemical and hodological techniques. From 50 to 60 immunoreactive neurons, labelled after contralateral intraocular injection of rhodamine β-isothiocyanate, form a small, clearly defined, nucleus in the lateral neural plate of the magnocellular preoptic nucleus. These cells give rise to immunoreactive axons which have been observed at the base of the nucleus, in the optic chiasma and in the optic nerve, to project into the intermediate plexiform layer of the retina, which separates the layer of internal horizontal cells from the layer of external horizontal cells. This FMRF-amide-like immunoreactive centrifugal visual system is compared to that described in Gnathostomes.

Horizontal cell progenitors arrest in G2-phase and undergo terminal mitosis on the vitreal side of the chick retina

We have addressed the question when horizontal cells in the chick retina are generated and undergo their terminal mitosis. Horizontal cell progenitors replicate their DNA early and migrate bi-directionally to the horizontal cell layer. It was hypothesized that the cells undergo mitosis directly after replication and migrate as post-mitotic transition cells before differentiating to horizontal cells. However, our results show that cells expressing markers for the axon-bearing and the axon-less subtypes of horizontal cells undergo terminal mitosis while residing on the vitreal side of the retina. By combining horizontal cell transcription factors Lim1, Isl1 and Prox1 labeling with phospho-histone H3, a marker for mitosis, we demonstrate that all or a clear majority of vitreal mitoses are undertaken by the horizontal cell committed progenitors. The pattern of cells that incorporated the thymidine analogue EdU implied that the progenitors replicated their genome while migrating towards the vitreal side. Upon arrival to the vitreal retina they become arrested for about two days prior to mitosis. Hence, cells expressing horizontal cell markers are arrested in G2-phase on the vitreal side of the retina. These results support the existence of committed progenitors that give rise to horizontal cells and that those cells become arrested in G2-phase before undergoing terminal mitosis on the vitreal side of the retina followed by migration to the horizontal cell layer. The results also indicate that the regulation of the transition from G2-phase to mitosis is important for the development of these committed progenitor cells.

Early development of GABAergic cells of the retina in sharks: An immunohistochemical study with GABA and GAD antibodies

We studied the ontogeny and organization of GABAergic cells in the retina of two elasmobranches, the lesser-spotted dogfish ( Scyliorhinus canicula) and the brown shyshark ( Haploblepharus fuscus) by using immunohistochemistry for gamma-aminobutyric acid (GABA) and glutamic acid decarboxylase (GAD). Both antibodies revealed the same pattern of immunoreactivity and both species showed similar organization of GABAergic cells. GABAergic cells were first detected in neural retina of embryos at stage 26, which showed a neuroepithelial appearance without any layering. In stages 27–29 the retina showed similar organization but the number of neuroblastic GABAergic cells increased. When layering became apparent in the central retina ( stage-30 embryos), GABAergic cells mainly appeared organized in the outer and inner retina, and GABAergic processes and fibres were seen in the primordial inner plexiform layer (IPL), optic fibre layer and optic nerve stalk. In stage-32 embryos, layering was completed in the central retina, where immunoreactivity appeared in perikarya of the horizontal cell layer, inner nuclear layer and ganglion cell layer, and in numerous processes coursing in the IPL, optic fibre layer and optic nerve. From stage 32 to hatching ( stage 34), the layered retina extends from centre-to-periphery, recapitulating that observed in the central retina at earlier stages. In adults, GABA/GAD immunoreactivity disappears from the horizontal cell layer except in the marginal retina. Our results indicate that the source of GABA in the shark retina can be explained by its synthesis by GAD. Such synthesis precedes layering and synaptogenesis, thus supporting a developmental role for GABA in addition to act as neurotransmitter and neuromodulator.

Axon-bearing and axon-less horizontal cell subtypes are generated consecutively during chick retinal development from progenitors that are sensitive to follistatin.

BackgroundHorizontal cells are retinal interneurons that modulate the output from photoreceptors. A rich literature on the morphological classification and functional properties of HCs in different animals exists, however, the understanding of the events underlying their development is still limited. In most vertebrates including chicken, two main horizontal cell (HC) subtypes are identified based on the presence or absence of an axon.ResultsIn this work we have molecularly characterized three HC subtypes based on Lim1, Isl1, GABA and TrkA, a classification that is consistent with three chick HC subtypes previously defined by morphology. The axon-bearing and axon-less HC subpopulations molecularly defined by Lim1 and Isl1, are born consecutively on embryonic day (E) 3–4 and E4–5, respectively, and exhibit temporally distinguishable periods of migration. Their relative numbers are not adjusted by apoptosis. A sharp decrease of high endogenous levels of the activin-inhibitor follistatin at E3 coincides with the appearance of the Lim1 positive cells. Extending the follistatin exposure of the HC retinal progenitor cells by injection of follistatin at E3 increased the number of both Lim1- and Isl1 positive HCs when analysed at E9.ConclusionThe results imply that the axon-bearing and axon-less HC subgroups are defined early and are generated consecutively from a retinal progenitor cell population that is sensitive to the inhibitory action of follistatin. The results are consistent with a model wherein added follistatin causes HC-generating progenitors to proliferate beyond the normal period of HC generation, thus producing extra HCs of both types that migrate to the HC layer.

Somal positioning and dendritic growth of horizontal cells are regulated by interactions with homotypic neighbors

Retinal neurons extend their dendritic fields to achieve a degree of dendritic overlap with homotypic neighbors that is cell-type specific. How these neurons regulate their dendritic growth is unclear. The dendritic field of a retinal horizontal cell varies inversely with horizontal cell density across different strains of mice, suggesting that proximity to neighboring cells regulates dendritic growth. To test this directly, we have employed the Cre-loxP conditional gene targeting strategy to achieve inactivation of Lim1 function in developing horizontal cells. Through this approach, Lim1 function was prevented within a subset of horizontal cells that in turn fail to migrate to the horizontal cell layer and differentiate normally. For those remaining horizontal cells with Lim1 intact (about half of the normal population in these mice), we show that they spread themselves out tangentially and differentiate a dendritic morphology that is essentially normal but for the fact that it has nearly doubled in area. Such larger horizontal cells, sampling from an area of retina containing twice their normal afferent number, differentiate a dendritic field with nearly double the number of higher order branches and terminal clusters. These results demonstrate directly that positioning and dendritic growth are regulated by interactions with homotypic neighbors, whereas afferents instruct the differentiation of dendritic patterning.

Vimentin isoform expression in the human retina characterized with the monoclonal antibody 3CB2

The antigen recognized by the monoclonal antibody 3CB2 (3CB2-Ag and 3CB2 mAb) is expressed by radial glia and astrocytes in the developing and adult vertebrate central nervous system (CNS) of vertebrates as well as in neural stem cells. Here we identified the 3CB2-Ag as vimentin by proteomic analysis of human glial cell line U-87 extracts (derived from a malignant astrocytoma). Indeed, the 3CB2 mAb recognized three vimentin isoforms in glial cell lines. In the human retina, 3CB2-Ag was expressed in Müller cells, astrocytes, some blood vessels, and cells in the horizontal cell layer, as determined by immunoprecipitation and immunofluorescence. Three populations of astrocytes were distinguishable by double-labeling immunohistochemistry: vimentin+/GFAP+, vimentin-/GFAP+, and vimentin+/GFAP-. Hence, we conclude that 1) the 3CB2-Ag is vimentin; 2) vimentin isoforms are differentially expressed in normal and transformed astrocytes; 3) human retinal astrocytes display molecular heterogeneity; and 4) the 3CB2 mAb is a valuable tool to study vimentin expression and its function in the human retina.

Morphogenesis in the retina of a slow-developing teleost: Emergence of the GABAergic system in relation to cell proliferation and differentiation

Gamma-aminobutyric acid (GABA) has been implicated in cell proliferation and differentiation during development. In the present study, immunohistochemical techniques were used to investigate the development of the GABAergic system in the retina of the trout and its relation to markers of differentiation [calretinin (CR), and tyrosine hydroxylase (TH)]. The expression of Pax6, an eye-patterning protein involved in the proliferation and emergence of specific retinal cell types, was also studied. Retinal layering was observed to begin centrally in prehatching embryos, as the first GABAergic cells appeared in the ganglion cell layer (GCL) and inner part of the inner nuclear layer (INL). At hatching, GABAergic cells were also observed in the horizontal cell layer (HCL). In alevins, GABAergic cells and processes spread laterally following retinal growth although they did not invade neuroblastic retinal regions. CR- and Pax6-immunoreactive (ir) cells were first seen in the GCL and the inner part of the INL, whereas sparse TH-ir cells appeared in the INL. In juveniles, GABAergic cells were observed in the GCL, inner part of the INL and HCL, whereas CR-ir cells spread to the outer part of the INL and HCL. A subset of CR-ir in the GCL and of Pax6-ir cells in the GCL and INL showed colocalization with GABAergic markers. This study provides further comparative knowledge about the development of GABAergic system of the retina in teleosts and shows differences and similarities with that reported in fast-developing species such as zebrafish, in which retinal expression of GABA was transient in some populations.

Localization and Diurnal Expression of Melanopsin, Vertebrate Ancient Opsin, and Pituitary Adenylate Cyclase-Activating Peptide mRNA in a Teleost Retina

Localization and Diurnal Expression of Melanopsin, Vertebrate Ancient Opsin, and Pituitary Adenylate Cyclase-Activating Peptide mRNA in a Teleost Retina

Nonapical Symmetric Divisions Underlie Horizontal Cell Layer Formation in the Developing Retina In Vivo

Symmetric cell divisions have been proposed to rapidly increase neuronal number late in neurogenesis, but how critical this mode of division is to establishing a specific neuronal layer is unknown. Using in vivo time-lapse imaging methods, we discovered that in the laminated zebrafish retina, the horizontal cell (HC) layer forms quickly during embryonic development upon division of a precursor cell population. The precursor cells morphologically resemble immature, postmitotic HCs and express HC markers such as ptf1a and Prox1 prior to division. These precursors undergo nonapical symmetric division at the laminar location where mature HCs contact photoreceptors. Strikingly, the precursor cell type we observed generates exclusively HCs. We have thus identified a dedicated HC precursor, and our findings suggest a mechanism of neuronal layer formation whereby the location of mitosis could facilitate rapid contact between synaptic partners.