In situ hybridization to detect spatial gene expression in medaka

Abstract

A whole-animal tissue section in situ hybridization (ISH) system with radio-labeled probes was developed to detect differential gene expression among tissues of the small, oviparous teleost fish, Japanese medaka ( Oryzias latipes). Because of its tissue- and gender-specific expression, gonadal aromatase (CYP19a) was selected as a model gene to demonstrate the potential of the system. The ISH system was validated with a 7 d exposure to the model aromatase inhibitor, fadrozole. Fadrozole did not affect the magnitude of gene expression in testes, but significantly up-regulated CYP19a gene expression in ovaries. These results were confirmed with quantitative real-time-polymerase chain reaction (RT-PCR). Histological evaluation revealed that females exposed to 100 μg/L fadrozole lacked mature oocytes. Male gonadal morphology was normal in all treatments. The ISH method developed in this study allowed tissue-specific resolution of gene expression in a whole animal model, as well as the ability to analyze cellular morphological detail in the same organism.