The effect of trypan blue on the growth of the rat embryo in vivo.

In a previous paper (Berry, 1968) it has been demonstrated, by comparison of in vivo and in vitro growth of the rat foetus, that it is possible to dissociate growth and differentiation in the rat. In a series now exceeding 1000 embryos it is evident that for a given somite number the protein content of an animal developing in vitro might be less than half that of the normal control, suggesting that a considerable reduction in growth rate might be found without necessarily inducing death or malformation. The rate of somite formation is not reduced in vitro in the rat; it has been shown by Herrman & Schultz (1958) that the rate of somite formation is not interfered with by different expiant conditions in the chick embryo. These findings suggest that the increase in somite number is more rigidly determined than the increase in growth rate. In an attempt to determine whether other methods of interfering with growth alter the rate of somite formation, the effects of trypan blue on the growth and development of the rat embryo in vivo were examined. The dye is localized in the yolk sac after maternal injection (Wilson, Shepard & Gennaro, 1963) and at this site has an inhibitory effect on lysosomal hydrolytic enzymes in the cells of this membrane (Beck, Lloyd & Griffiths, 1967). The compound is ineffective after the 10th day of pregnancy (Wilson, Beaudoin & Free, 1959)—counting the day on which a vaginal plug is found as day 1— presumably due to the development of effective placentation at this stage in the rat. Since foetal death following maternal trypan blue administration is secondary to malformation (Beck & Lloyd, 1963, 1966) it appears that trypan blue will produce malformation and death by an effect on embryonic nutrition via the yolk sac. This argument has been subsequently extended by Lloyd & Beck (1968). If embryonic death and malformation are related to defects in yolk sac function, it seems likely that all animals exposed to the compound in the susceptible period, whether morphologically abnormal or not, should show evidence of growth retardation. The embryos should be examined soon after 1 Author's address: Department of Morbid Anatomy, Department of Child Health, 30 Guildford Street, London W.C. 1.