The egg of the Japanese medaka (Oryzias latipes) is enclosed in a transparent chorion, which makes it possible to observe the entire embryonic development non-invasively. This process -from fertilization to hatching- takes about 192-240 hours, a timeframe we believe is an ideal for many types of research. For comparison, development is quicker in zebrafish (Danio rerio, ~72 hours) and fathead minnows (Pimephales promelas, ~120 hours), while it is much slower in other freshwater species such as the rainbow trout (Oncorhynchus mykiss), which can take over three weeks (Knight, 1963). This moderate developmental pace makes medaka embryos a valuable model for a wide range of vertebrate studies. Research has used them to explore topics from fertilization (Yamamoto, 1944; Hishida and Nakano, 1954; Kageyama, 1977; Iwamatsu et al 1997, Shibata et al 2000; Hyodo et al., 2009; Li et al., 2011) to enzymatic activity in early (Fluck 1982, Inohaya et al 1999) and late stages (Yasumasu et al 1992, Nguyen et al 2001; Puangchit et al., 2017). Medaka embryos are also particularly sensitive to environmental agents and xenobiotics (e.g. Kimm-Brinson, 2001; Merino et al., 2010; Lee and Yang, 2010; Gonz, which has made this species a long-standing model in developmental toxicity research. They have been used in studies involving dioxin congeners (Chen and Cooper, 1999; Hanno et al., 2010), agricultural pesticides (Holcombe et al 1995, Teather et al 2001; Ji et al., 2021; Zhou et al., 2022), and environmental estrogens (Yokota et al 2000; Kawamura et al., 2002; Lee et al., 2012).
Notably, toxicity is not only influenced by chemical type, dose, and exposure duration, but also by the specific developmental window during which embryos are exposed. Embryos undergo rapid and dynamic changes, and even small timing differences can significantly alter the observed effects. As a result, medaka have also been used to study stage-specific responses to toxicants (Hamm and Hinton 2000, Owens and Baer 2000; Kataoka et al., 2018). To conduct any developmental study –particularly in toxicology- it is essential to accurately identify and classify stages in the embryonic phase. Having standardized criteria for normal development gives researchers a reliable baseline for detecting deviations, supports reproducibility, and improves cross-study comparisons.
Over the years, several Japanese researchers have documented medaka development in detail. To our knowledge, Matsui (1934, 1949) was the first to illustrate the embryogenesis of medaka, providing 75 drawings of 35 stages. His work was later revised by Usui (1962) and Gamo and Terajima (1963). Iwamatsu (1976), and Iraki and Iwamatsu (1979) added histological insights for early stages. Unfortunately, while valuable and carefully hand drawn, these early studies were published in Japanese, limiting access for international researchers. Yamamoto (1975) was the first to make a full description in English, presenting 76 drawings across 35 stages. Still, the most comprehensive effort was Iwamatsu´s later work (1994, 2002, 2011), which illustrated 40 stages—from the unfertilized egg to the eleutheroembryo—using both micro- and macro-anatomical observations from orange-red and wild type medaka. His figures, detailed and clear, have become a gold standard in medaka developmental biology.
Despite their quality, line drawings come with inherent limitations. Wile they are often meticulously detailed, they can be hard to apply in practice–especially for researchers trying to stage large groups of embryos using just a dissecting microscope. Idealized illustrations often include features that are not easily visible
in vivo or that require dechorionation, sectioning, or special imaging. Translating those idealized two-dimensional line drawing forms into real-life observations—on curved, three-dimensional embryos—isn’t always straightforward, especially for those without a strong background in embryology. From our experience, we saw the need for a simpler, more practical staging tool—one that uses real embryos, seen under conditions that most researchers have access to: a standard dissecting microscope.
The next logical step in medaka documentation was a photographic atlas. Kirchen and West (1976) created the first one, with 38 black and white photos covering 36 stages. We also came across online resources combining photos with Iwamatsu´s illustrations (e.g.
https://www.slideshare.net/guest124c20/slide-show-medaka1), and we recall another earlier site by Glase (1998), though it appears to be no longer accessible. Why has a full photographic atlas not been widely available until now? From our experience two challenges stand out:
- Developmental variability: Even under controlled laboratory conditions, we often struggle to match all the features of a published stage to a single embryo. Factors like oxygen levels, minor temperature shifts, and natural variation among embryos—even from the same mother—can result in noticeable differences. This variability becomes more pronounced over time.
- Image complexity: To capture all key features of a stage in one photograph is not easy. Line drawings can combine views and exaggerate clarity for teaching purposes. Real embryos, on the other hand, require manipulation, reorientation, and refocusing to bring structures into view across their curved surfaces and at different depths.
These challenges led us to produce a color photographic atlas covering 39 stages using an analog camera (González-Doncel et al., 2006). At the time of its production, digital imaging technology was not yet advanced or widely available in laboratories. Unfortunately, the later conversion from analog to digital format reduced image resolution and clarity. Now, with digital imaging standard in laboratories, we have revisited this effort. As a complement to our post-hatch histological atlas of the orange-red medaka strain, we present an updated
in vivo atlas of medaka embryo development. Each stage is shown as it would appear under a typical stereo dissecting microscope. We have included multiple angles for each stage to highlight easy-to-spot features that make quick and accurate staging possible. We hope this atlas serves as a practical, user-friendly resource alongside established references like Iwamatsu (2004), and that it proves valuable in both developmental and toxicological studies.
You can read more about our process in the next section, "How We Made It".