Basic research in pattern formation is concerned with the generation of phenotypes and tissues. It can therefore lead to new tools for medical research. These include phenotypic screening assays, applications in tissue engineering, as well as general advances in biomedical knowledge. Our aim here is to discuss this emerging field with special reference to tools based on zebrafish developmental biology. We describe phenotypic screening assays being developed in our own and other labs. Our assays involve: (i) systemic or local administration of a test compound or drug to zebrafish in vivo; (ii) the subsequent detection or "readout" of a defined phenotypic change. A positive readout may result from binding of the test compound to a molecular target involved in a developmental pathway. We present preliminary data on assays for compounds that modulate skeletal patterning, bone turnover, immune responses, inflammation and early-life stress. The assays use live zebrafish embryos and larvae as well as adult fish undergoing caudal fin regeneration. We describe proof-of-concept studies on the localised targeting of compounds into regeneration blastemas using microcarriers. Zebrafish are cheaper to maintain than rodents, produce large numbers of transparent eggs, and some zebrafish assays could be scaled-up into medium and high throughput screens. However, advances in automation and imaging are required. Zebrafish cannot replace mammalian models in the drug development pipeline. Nevertheless, they can provide a cost-effective bridge between cell-based assays and mammalian whole-organism models. KEY WORDS: Danio rerio, zebrafish, high-throughput screening, high-content screeningThis article is part of a special journal issue on "Pattern Formation". In this context, it seems appropriate to ask: 'what benefits to society can come from research into pattern formation?' The answer depends on how "pattern formation" is defined. For our purposes, it includes developmental mechanisms that generate Int. J. Dev. Biol. 53: 835-850 (2009) doi: 10.1387/ijdb.082615sb One is the field of human congenital malformations (the pres-836 S. A. Brittijn et al. ence at birth of anatomical abnormalities). Good examples are human malformations of fingers and toes, some cases of which are linked to mutations in the Hox family of developmental patterning genes (Akarsu et al., 1996;Goodman et al., 1997;Goodman, 2002). Teratology -the study of agents that cause congenital malformations -can be seen as applied pattern formation research. Tissue engineering, a medical research discipline that aims to create replacement tissues for patients who have suffered injury or surgical removal of tissues, is another area where knowledge of pattern formation can be applied. To date, the commonest approach to tissue engineering is to culture autologous cells on a synthetic template (Silva and Mooney, 2004). In the future, however, it may be possible to promote pattern formation by these cells so that they generate selforganised tissues or structures ...

Biologists measure developmental time by dividing development into arbitrary time blocks called "stages." This is a reasonable approach, provided that developmental timing is precisely controlled within a species. However, the degree of this precision is unknown. This is unfortunate because precision in developmental timing at the population level is a central issue to the whole research program of heterochrony. To examine this issue, we apply Ontogenetic Sequence Analysis to 261 embryos of the Lake Victoria cichlid Haplochromis piceatus. The result of our analysis can be mapped as a complex web of 26,880 equally parsimonious developmental sequences. This topology reflects timing polymorphism (intraspecific heterochrony) among embryos of this species. Because of this timing polymorphism, it is not possible to define discrete "stages" in this cichlid (although there is sufficient sequence signal to assess the maturity of embryos). More generally, we show that sequence polymorphism creates uncertainty about how a given embryo will develop implying that the mechanisms controlling developmental timing in embryos lack precision. For this reason, it is imperative to consider patterns of embryonic variability when measuring developmental time.

Because little is known about embryonic developmental stages in any haplochromine cichlid, we describe here a series of normal stages of the Lake Victoria cichlid Haplochromis piceatus. We collected 273 embryos and scored them for 47 morphological characters. The result was an illustrated series of 12 stages from embryonic shield until hatching based on live, fixed, and histological material. We defined each stage according to a single ''key'' character that applied to all embryos of that stage. Other characters forming part of the stage descriptions were not necessarily present in all embryos of a stage. We compare our findings with published studies of other freshwater teleosts and find wide variation in staging systems. Our data will form a baseline for further research on cichlid development.