Consideration of the liver of embryonic lacZ @entity19 as an analogue of the XXXX coat colour spot test: preliminary data and technical problems.

@entity19 @entity331
Pregnant lacZ+ @entity19 (Muta @entity19 ) were treated with @entity331 (25 mg/kg) by oral gavage on day 10.5 of pregnancy. This dose of @entity331 is the optimal dose observed by other investigators for activity in the @entity19 coat colour spot test. Day 10.5 of pregnancy represents the stage when the embryonic liver first becomes visually discernable. By day 15.5, when the maternal and embryonic livers were analysed for lacZ- mutation frequency, the embryonic liver is the largest tissue in the embryo (approximately 25 mg). These experiments therefore represent treatment of a small pool of progenitor hepatocytes just as they are entering into an intense wave of cell division--optimum conditions for the fixation of mutations. Exposure to @entity331 led to an average fourfold, and a maximum tenfold increase in mutation frequency in the embryonic livers. This relatively weak response is consistent with lacZ- mutants not having a growth advantage; unlike in the @entity19 spot test, clonal amplification of lacZ- mutants cannot be separately scored. The level of mutation in the control embryonic livers was lower than that of the maternal control livers, but the group sizes were too small to conclude this definitively. Embryonic livers from each individual mother showed a range of mutation frequencies that were not obviously related to that of the maternal liver. On a treatment group basis, @entity331 was non-mutagenic to maternal livers. Half of the embryonic livers yielded DNA that failed to package, despite repeated attempts and re-isolation of the DNA from the liver. The cause of this unexpected finding is not clear. What is clear is that it was not due to the @entity331 treatment because eight of 22 control embryonic livers behaved similarly. These preliminary results suggest that further research is required in order to establish a practical transgenic analogue of the @entity19 coat colour spot test.
