Dynamic expression of ganglion cell markers in retinal progenitors during the terminal cell cycle

Department of Human Genetics, University of Michigan, Ann Arbor, MI 48109, United States.
Molecular and Cellular Neuroscience (Impact Factor: 3.84). 05/2012; 50(2):160-8. DOI: 10.1016/j.mcn.2012.05.002
Source: PubMed


The vertebrate neural retina contains seven major cell types, which arise from a common multipotent progenitor pool. During neurogenesis, these cells stop cycling, commit to a single fate, and differentiate. The mechanism and order of these steps remain unclear. The first-born type of retinal neurons, ganglion cells (RGCs), develop through the actions of Math5 (Atoh7), Brn3b (Pou4f2) and Islet1 (Isl1) factors, whereas inhibitory amacrine and horizontal precursors require Ptf1a for differentiation. We have examined the link between these markers, and the timing of their expression during the terminal cell cycle, by nucleoside pulse-chase analysis in the mouse retina. We show that G2 phase lasts 1-2 h at embryonic (E) 13.5 and E15.5 stages. Surprisingly, we found that cells expressing Brn3b and/or Isl1 were frequently co-labeled with EdU after a short chase (<1 h) in early embryos (<E14), indicating that these factors, which mark committed RGCs, can be expressed during S or G2 phases. However, during late development (>E15), Brn3b and Isl1 were exclusively expressed in post-mitotic cells, even as new RGCs are still generated. In contrast, Ptf1a and amacrine marker AP2α were detected only after terminal mitosis, at all developmental stages. Using a retroviral tracer in embryonic retinal explants (E12-E13), we identified two-cell clones containing paired ganglion cells, consistent with RGC fate commitment prior to terminal mitosis. Thus, although cell cycle exit and fate determination are temporally correlated during retinal neurogenesis, the order of these events varies according to developmental stage and final cell type.

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Available from: Tom Glaser, May 19, 2015
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    • "Atoh7 was shown to be capable of inducing differentiation of ganglion cells if expressed in cells restricted to amacrine and photoreceptor cell fates. However, Atoh7 was not sufficient to alter differentiation significantly if expressed in cells committed to the photoreceptor lineage (Prasov and Glaser, 2012a; Mao et al., 2013). Together these findings suggest a differential susceptibility of RPCs and differentiating RPCs towards Atoh7 function. "
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    ABSTRACT: During vertebrate eye development retinal progenitor cells (RPCs) differentiate into all neural cell types of the retina. Retinal ganglion cells (RGCs) represent the first cell type to be generated. For their development, Atoh7, a basic Helix Loop Helix (bHLH) transcription factor is crucial. Atoh7 loss of function results in a massive reduction or even a total loss of RGCs. However, inconsistent results have been obtained in atoh7 gain of function experiments with respect to ganglion cell genesis, implying that the effect of Atoh7 is likely to be dependent on the competence state of the RPC. In this study we addressed the differential susceptibilities of early RPCs to Atoh7 in vivo, using medaka. Unexpectedly, we observed a largely normal development of the dorsal retina, although atoh7 was precociously expressed. However, the development of the retina close to the optic nerve head (part of the ventral retina) was disturbed severely. Photoreceptors were largely absent and the Müller glia cell number was reduced significantly. The majority of cells in this domain were ganglion cells and the abnormal development of this area affected the closure of the optic fissure resulting in coloboma.
    Full-text · Article · Aug 2014 · Mechanisms of Development
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    • "In the human retina, RGC genesis begins around fetal week (Fwk) 5 and ends around Fwk 18 (Pacal and Bremner, 2014), whereas in fish and amphibians, new RGCs can be added throughout life. Commitment of progenitors to a RGC fate occurs during, or just shortly after, the terminal cell division (McLoon and Barnes, 1989; Waid and McLoon, 1995; Prasov and Glaser, 2012; Pacal and Bremner, 2014) and is controlled by a combination of intrinsic factors and cell–cell signals. Differentiation of RGCs begins in the central retina and spreads peripherally in a roughly concentric fashion (Drager, 1985; Hu and Easter, 1999; McCabe et al., 1999; Neumann and Nuesslein-Volhard, 2000; Martinez-Morales et al., 2005; Hufnagel et al., 2010). "
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    ABSTRACT: The visual system is beautifully crafted to transmit information of the external world to visual processing and cognitive centers in the brain. For visual information to be relayed to the brain, a series of axon pathfinding events must take place to ensure that the axons of retinal ganglion cells, the only neuronal cell type in the retina that sends axons out of the retina, find their way out of the eye to connect with targets in the brain. In the past few decades, the power of molecular and genetic tools, including the generation of genetically manipulated mouse lines, have multiplied our knowledge about the molecular mechanisms involved in the sculpting of the visual system. Here, we review major advances in our understanding of the mechanisms controlling the differentiation of RGCs, guidance of their axons from the retina to the primary visual centers, and the refinement processes essential for the establishment of topographic maps and eye-specific axon segregation. Human disorders, such as albinism and achiasmia, that impair RGC axon growth and guidance and, thus, the establishment of a fully functioning visual system will also be discussed. The eyes together with their connecting pathways to the brain form the visual system. In the eye, the cornea bends light rays and is primarily responsible for focusing the image on the retina. The lens behind the cornea inverts the image top to bottom and right to left. The retina, the receptive surface inside the back of the eye, is the struc-ture that translates light into nerve signals, and enables us to see under conditions that range from dark to sunlight, discriminate colors, and provide a high degree of visual precision. The retina consists of three layers of nerve cell bodies separated by two layers containing synapses made by the axons and dendrites of these cells. The back of the retina comprises the photoreceptors, the rods, and cones. The medial retinal layer contains three types of nerve cells, bipolar, horizontal, and amacrine cells. Bipolar cells receive input from the photoreceptors, and many of them feed directly into the retinal ganglion cells (RGCs). Horizontal cells connect receptors and bipolar cells by relatively long connections that run parallel to the retinal layers. Amacrine cells link bipolar cells and RGCs, the cells located in the inner retina. RGC axons pass across the surface of the retina and are collected in a bundle at the optic disk to leave the eye and form the optic nerve. There are approximately 20 RGC types that can be classified by morphological, molecular, and func-tional criteria. Each RGC type participates in distinct retinal circuits and projects to a specific set of targets in the brain (Coombs et al., 2007; Schmidt et al., 2011), including the main image-forming nuclei such as the lat-eral geniculate nucleus (LGN), the visual part of the thal-amus, and the superior colliculus (SC), located in the roof of the midbrain, that coordinates rapid movement of the eye (Figure 1). The optic axons from both eyes meet at the optic chiasm, which is located at the base of the hypothalamus. There, RGC axons from the nasal retina cross over to the opposite side of the brain (contralateral or commissural axons) while axons from the temporal retina turn to
    Full-text · Article · Jan 2014
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    • "Integration of the curve gave an estimation of the total number of mitoses during the period st26–st33 with in total 130 Lim1+ basal mitoses/10 µm section. The length of the late G2/M-phase (PH3 staining) during the investigated stages was assumed to be approximately 1 h [54] and is shorter than the sampling frequency (12 h/st). The risk of counting a cell with a basal mitosis twice is very little and no compensation is required. "
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    ABSTRACT: Retinal progenitor cells undergo apical mitoses during the process of interkinetic nuclear migration and newly generated post-mitotic neurons migrate to their prospective retinal layer. Whereas this is valid for most types of retinal neurons, chicken horizontal cells are generated by delayed non-apical mitoses from dedicated progenitors. The regulation of such final cell cycle is not well understood and we have studied how Lim1 expressing horizontal progenitor cells (HPCs) exit the cell cycle. We have used markers for S- and G2/M-phase in combination with markers for cell cycle regulators Rb1, cyclin B1, cdc25C and p27Kip1 to characterise the final cell cycle of HPCs. The results show that Lim1+ HPCs are heterogenic with regards to when and during what phase they leave the final cell cycle. Not all horizontal cells were generated by a non-apical (basal) mitosis; instead, the HPCs exhibited three different behaviours during the final cell cycle. Thirty-five percent of the Lim1+ horizontal cells was estimated to be generated by non-apical mitoses. The other horizontal cells were either generated by an interkinetic nuclear migration with an apical mitosis or by a cell cycle with an S-phase that was not followed by any mitosis. Such cells remain with replicated DNA and may be regarded as somatic heteroploids. The observed heterogeneity of the final cell cycle was also seen in the expression of Rb1, cyclin B1, cdc25C and p27Kip1. Phosphorylated Rb1-Ser608 was restricted to the Lim1+ cells that entered S-phase while cyclin B1 and cdc25C were exclusively expressed in HPCs having a basal mitosis. Only HPCs that leave the cell cycle after an apical mitosis expressed p27Kip1. We speculate that the cell cycle heterogeneity with formation of heteroploid cells may present a cellular context that contributes to the suggested propensity of these cells to generate cancer when the retinoblastoma gene is mutated.
    Full-text · Article · Mar 2013 · PLoS ONE
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