In situ hybridization to detect spatial gene expression in medaka.
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 7d 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 100microg/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.
Tompsett
A R
AR
Department of Zoology, Center for Integrative Toxicology, Michigan State University, East Lansing, MI, USA. amber.tompsett@usask.ca
Park
J W
JW
Zhang
X
X
Jones
P D
PD
Newsted
J L
JL
Au
D W T
DW
Chen
E X H
EX
Yu
R
R
Wu
R S S
RS
Kong
R Y C
RY
Giesy
J P
JP
Hecker
M
M
D000818:Animals; D001345:Autoradiography; D001835:Body Weight / Q000187:drug effects; D017931:DNA Primers; D018076:DNA, Complementary / Q000096:biosynthesis / Q000235:genetics; D004965:Estrogen Antagonists / Q000494:pharmacology; D017316:Fadrozole / Q000494:pharmacology; D005260:Female; D005618:Fresh Water / Q000032:analysis; D015870:Gene Expression / Q000502:physiology*; D006128:Growth / Q000187:drug effects; D007091:Image Processing, Computer-Assisted; D007150:Immunohistochemistry; D017403:In Situ Hybridization; D008297:Male; D009990:Oryzias / Q000502:physiology*; D010053:Ovary / Q000378:metabolism / Q000473:pathology; D015347:RNA Probes; D012333:RNA, Messenger / Q000032:analysis / Q000096:biosynthesis; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013737:Testis / Q000378:metabolism / Q000473:pathology
D017931:DNA Primers; D018076:DNA, Complementary; D004965:Estrogen Antagonists; D015347:RNA Probes; D012333:RNA, Messenger; D017316:Fadrozole