With the increased use of mutant mice and zebrafish in vision research there is a growing need for methods to: (a) visualize individual neuronal morphologies as part of the phenotypic analysis of mutant animals, (b) to mark cells in a noninvasive manner for lineage studies, and (c) to visualize subcellular structures in a sparse subset of living neurons. This proposal has two overall goals. The first goal is to develop, test, and optimize a set of reagents - principally, plasmids and mouse and zebrafish lines - that would be easy to use, provide high quality visualization of cellular morphology and subcellular structures, and be freely available to the research community. In mice, the methodology that we will use is based on a pharmacologically controlled Cre recombinase-estrogen receptor fusion protein that irreversibly activates (by DNA recombination) the production of one or more enzymatic or fluorescent reporter proteins in a sparse subset of cells. In zebrafish, sparse labeling will be achieved by taking advantage of the variegated expression that typically accompanies transgenesis. The second goal is to use these reagents for (a) mapping of retinal ganglion cell morphologies in two lines of transcription factor gene knock-out mice (the POU-homeodomain genes Brn3b and Brn3c), and (b) identifying the spatial distribution of pre- and post-synaptic sites in single identified neurons in both living and fixed retinas in mice and zebrafish. Although the methodology and reagents that we develop will be tested in and applied to the retina, they will be generally useful for the analysis of any part of the nervous system.
| Badea, Tudor C; Cahill, Hugh; Ecker, Jen et al. (2009) Distinct roles of transcription factors brn3a and brn3b in controlling the development, morphology, and function of retinal ganglion cells. Neuron 61:852-64 |
