Timing and topography of cell genesis in the rat retina

Abstract

To understand the mechanisms of cell fate determination in the vertebrate retina, the time course of the generation of the major cell types needs to be established. This will help define and interpret patterns of gene expression, waves of differentiation, timing and extent of competence, and many of the other developmental processes involved in fate acquisition. A thorough retinal cell "birthdating" study has not been performed for the laboratory rat, even though it is the species of choice for many contemporary developmental studies of the vertebrate retina. We investigated the timing and spatial pattern of cell genesis using 3H-thymidine (3H-TdR). A single injection of 3H-TdR was administered to pregnant rats or rat pups between embryonic day (E) 8 and postnatal day (P) 13. The offspring of prenatally injected rats were delivered and all animals survived to maturity. Labeled cells were visualized by autoradiography of retinal sections. Rat retinal cell genesis commenced around E10, 50% of cells were born by approximately P1, and retinogenesis was complete near P12. The first postmitotic cells were found in the retinal ganglion cell layer and were 9-15 microm in diameter. This range includes small to medium diameter retinal ganglion cells and large displaced amacrine cells. The sequence of cell genesis was established by determining the age at which 5, 50, and 95% of the total population of cells of each phenotype became postmitotic. With few exceptions, the cell types reached these developmental landmarks in the following order: retinal ganglion cells, horizontal cells, cones, amacrine cells, rods, bipolar cells, and Müller glia. For each type, the first cells generated were located in the central retina and the last cells in the peripheral retina. Within the sequence of cell genesis, two or three phases could be detected based on differences in timing, kinetics, and topographic gradients of cell production. Our results show that retinal cells in the rat are generated in a sequence similar to that of the primate retina, in which retinogenesis spans more than 100 days. To the extent that sequences reflect underlying mechanisms of cell fate determination, they appear to be conserved.

Controlled expression of transgenes introduced by in vivo electroporation

“…4, the clusterin promoter was cloned upstream of ER T2 CreER T2 , and the resulting plasmid was coelectroporated with CALNL-DsRed and CAG-GFP into P0 rat retinas. The retinas were stimulated with 4OHT at P14, the time point when retinogenesis is complete (24,25), and then analyzed at P16. Without 4OHT stimulation, DsRed expression was not detected (Fig.…”

Development and diversification of retinal amacrine interneurons at single cell resolution

“…Single cells, either genetically labeled or randomly chosen from a pool of dissociated cells, were isolated from mouse retinae at 6 developmental ages [supporting information (SI) Table S1] (8). Although these cells had not yet developed their specific morphologies, these time points were chosen to identify transcriptional programs active in amacrine cells at different stages of neuronal differentiation (9). In all, 32 single amacrine cells were isolated and profiled on Affymetrix microarrays.…”

“…To test this hypothesis, we performed classical birthdating studies of GABAergic and glycinergic amacrine cells (9,18,19). Pregnant mice were injected with [ 3 H]-thymidine at 1 of 4 time points during the window of amacrine cell production.…”

Cell density ratios in a foveal patch in macaque retina

“…Because the number of cell divisions that produce such a retinal clone is small (Rapaport et al, 1996), the number of cells of a given group may be just one or two, with later differentiation within each group into specific cell types. For example, we find close to one outer diffuse bipolar cell per foveal cone, but this group of cells is comprised of three cell types, DB1, DB2, and DB3 (Boycott & Wässle, 1991).…”