We have developed an Object Segmentation, Counter and Analysis Resource (OSCAR) that is designed specifically to quantify densely packed biological samples with reduced signal-to-background ratio. OSCAR uses as input three dimensional images reconstructed from confocal 2D sections stained with dies such as nuclear marker and immunofluorescence labeling against specific antibodies to distinguish the cell types of interest. Taking advantage of a combination of arithmetic, geometric and statistical algorithms, OSCAR is able to reconstruct the objects in the 3D space bypassing segmentation errors due to the typical reduced signal to noise ration of biological tissues imaged in toto. When applied to the zebrafish developing retina, OSCAR is able to locate and identify the fate of each nuclei as a cycling progenitor or a terminally differentiated cell, providing a quantitative characterization of the dynamics of the developing vertebrate retina in space and time with unprecedented accuracy.

Morphogens of the Hh-family trigger gene expression changes of receiving cells in a concentration-dependent manner. The outputs of the pathway include regulation of cell identity, proliferation, death or metabolism, depending on the tissue or organ. This variety of responses relies on a conserved signaling pathway. Its internal logic includes a negative feedback loop involving the Hh receptor Ptc. In this paper, we use experiments and computational models to study and compare the different spatial signaling profiles downstream of Hh in several developing Drosophila organs. We show that the spatial distribution of Ptc and the activator form of the Gli transcription factor, CiA, in wing, antenna and ocellus show similar features, but markedly different from that in the compound eye (CE). We show that these two profile types represent two time points along the signaling dynamics, and that the interplay between the spatial displacement of the Hh source in the CE and the negative feedback loop maintains the receiving cells effectively in an earlier stage of signaling. These results indicate that the dynamics of the Hh source strongly influences the signaling profile Hh elicits in receiving cells, and show how the interaction between spatial and temporal dynamics of signaling and differentiation processes can contribute to the informational versatility of the conserved Hh signaling pathway.