Stages of embryonic development from zygote to hatching in the viviparous fish, Gambusia holbrooki

Developmental staging is indispensable to study developmental pathways, genetics and evolution among taxa. Life-history stages have been described for few Poeciliid species, however, proposed classifications are not generally applicable due to divergent morphology among the different Poeciliid species. In Poeciliid, with placentation developmental strategy, embryos receive the maternal investment throughout gestation. Therefore, normal morpho-physiological characteristics are often compromised by applying previously employed in vitro culture and embryo fixation approaches. Here, we describe the early development of the embryonic process in a livebearing fish, Gambusia holbrooki, based on detailed morphological diagnostics and overlay cardiac rate in developing embryos under normal conditions. Light microscopy observations of live embryos obtained from gravid wild captured females, allowed assignment of the development process into seven periods: zygote, cleavage, blastula, gastrula, segmentation, Pharyngula, and hatching. Thirty-seven stages were determined and assigned within each of these periods applying a numbered staging system. The novelty of the present study is the real-time, in vivo quantification of heart rhythm variations as a marker for embryonic staging from early gastrulation to late hatching. Moreover, at least one morphologic landmark for each of the embryonic stages was defined to maximize the accuracy of staging.