Failure of neural tube closure in the prosencephalon after administering caffeine to chick embryo.

A: The control embryo was treated with physiological saline solution, and then photographed as a whole-mount embryo and transverse sections at the head and trunk levels, which is indicated by the dotted line (A1 and A2). The black arrowheads indicate the point of neural tube closure, hence neural tube formation. The green arrowheads indicate the development of somites. B–D: 0.5, 1 and 1.5 mg/ml caffeine-treated embryos photographed as whole mount embryos. Transverse sections were taken at the head ad trunk levels as indicated by the dotted lines (B1–D2). The transverse sections show the failure of neural tube closure and thickening of the head mesenchymal layer in the prosencephalon (B1–D1), which are indicated by the black arrowheads. Also, the cells of the somites appear more scattered in comparison to the control, as indicated by the green arrows (B2–D2). The process of neurulation is completely inapparent in the high caffeine concentration groups, as indicated by the flatness of the neural plates (C–D). The black horizontal lines on neural tubes (A2–D2) indicate the width of neural tubes. Scale bar = 1 mm in A–D and 100 µm in A1–D2. Abbreviation: Nt, neural tube.