math5 is a murine orthologue of atonal, a bHLH proneural gene essential for the formation of photoreceptors and chordotonal organs in Drosophila. The expression of math5 coincides with the onset of retinal ganglion cell (RGC) differentiation. Targeted deletion of math5 blocks the initial differentiation of 80% of RGCs and results in an increase in differentiated amacrine cells. Furthermore, the absence of math5 abolishes the retinal expression of brn-3b and the formation of virtually all brn-3b-expressing RGCs. These results imply that math5 is a proneural gene essential for RGC differentiation and that math5 acts upstream to activate brn-3b-dependent differentiation processes in RGCs. The mammalian retina is the peripheral portion of the visual system containing six major neuronal cell types and one glial cell type organized in a laminar structure. The visual information process in the retina follows a general pathway from photoreceptors to bipolar cells to retinal ganglion cells (RGCs). Horizontal cells and amacrine cells act to mediate lateral interactions among photoreceptors, bipolar cells, and RGCs. The latter serve as the sole output neurons in the retina to send the visual information down the optic nerve to the rest of brain. Both birthdating experiments using 3 H-thymidine labeling and cell lineage analysis using retroviral and tracermediated approaches demonstrate that vertebrate retinal neurons are generated from common progenitors through sequential differentiation and ordered migration to form the laminar retinal structure (Cepko et al. 1996). Current models for retinal neuron differentiation suggest that the formation of a specific retinal neuron is determined by the intrinsic properties of the retinal progenitor and the extrinsic cues from the retinal environment (Cepko et al. 1996). Among the likely intrinsic factors, the basic helix-loop-helix (bHLH) class of proneural transcription factors appears to play an essential role in regulating the differentiation of retinal neurons (Cepko 1999). In Drosophila, expression of proneural genes of the achaete-scute complex (AS-C) and atonal (ato) in proneural clusters endow cells with neural competence (Jan and Jan 1993). Loss-of-function mutation of genes in AS-C causes the cells in the would-be proneural clusters to adopt epidermal fates rather than neuronal precursor fates. Conversely, gain-of-function mutations in the proneural genes leads to the ectopic formation of sensory neurons (Campuzano and Modolell 1992). The expression of ato is found in the optic furrow of the eye-antennal disc in addition to the ectodermal proneural clusters and sensory organ precursors, which give rise to the chordotonal organs. Deletion of ato causes the absence of the chordotonal organ and the lack of photoreceptors (Jarman et al. 1995), and the reduced expression of ato results in defects in axonal pathfinding of photoreceptors (White and Jarman 2000), suggesting that ato plays dual roles in determining neuronal potential and regulating specific neuronal differentiation...

In Brn3b(-/-) mice, where 80% of retinal ganglion cells degenerate early in development, the remaining 20% include most or all ganglion cell types. Cells of the same type cover the retinal surface evenly but tile it incompletely, indicating that a regular mosaic and normal dendritic field size can be maintained in the absence of contact among homotypic cells. In Math5(-/-) mice, where only approximately 5% of ganglion cells are formed, the dendritic arbors of at least two types among the residual ganglion cells are indistinguishable from normal in shape and size, even though throughout development they are separated by millimeters from the nearest neighboring ganglion cell of the same type. It appears that the primary phenotype of retinal ganglion cells can develop without homotypic contact; dendritic repulsion may be an end-stage mechanism that fine-tunes the dendritic arbors for more efficient coverage of the retinal surface.

Opsin 4 (Opn4)/melanopsin-expressing intrinsically photosensitive retinal ganglion cells (ipRGCs) play a major role in non-imageforming visual system. Although advances have been made in understanding their morphological features and functions, the molecular mechanisms that regulate their formation and survival remain unknown. Previously, we found that mouse T-box brain 2 (Tbr2) (also known as Eomes), a T-box-containing transcription factor, was expressed in a subset of newborn RGCs, suggesting that it is involved in the formation of specific RGC subtypes. In this in vivo study, we used complex mouse genetics, single-cell dye tracing, and behavioral analyses to determine whether Tbr2 regulates ipRGC formation and survival. Our results show the following: (1) Opn4 is expressed exclusively in Tbr2-positive RGCs; (2) no ipRGCs are detected when Tbr2 is genetically ablated before RGC specification; and (3) most ipRGCs are eliminated when Tbr2 is deleted in established ipRGCs. The few remaining ipRGCs display abnormal dendritic morphological features and functions. In addition, some Tbr2-expressing RGCs can activate Opn4 expression on the loss of native ipRGCs, suggesting that Tbr2-expressing RGCs may serve as a reservoir of ipRGCs to regulate the number of ipRGCs and the expression levels of Opn4.

Retinal ganglion cells (RGCs) are the first cell type to differentiate during retinal histogenesis. It has been postulated that specified RGCs subsequently influence the number and fate of the remaining progenitors to produce the rest of the retinal cell types. However, several genetic knockout models have argued against this developmental role for RGCs. Although it is known that RGCs secrete cellular factors implicated in cell proliferation, survival, and differentiation, until now, limited publications have shown that reductions in the RGC number cause significant changes in these processes. In this study, we observed that Math5 and Brn3b double null mice exhibited over a 99% reduction in the number of RGCs during development. This severe reduction of RGCs is accompanied by a drastic loss in the number of all other retinal cell types that was never seen before. Unlike Brn3b null or Math5 null animals, mice null for both alleles lack an optic nerve and have severe retinal dysfunction. Results of this study support the hypothesis that RGCs play a pivotal role in the late phase of mammalian retina development.