Mouse Development II: Alkaline phosphatase staining for germ cells and fetal alcohol exposure Part 1. Germ cell staining 1. Carefully remove fixative from gonads fixed last week. Discard it in waste container. 2. Wash gonads twice with alkaline phosphatase (AP) buffer for 20 minutes. 3. Add AP substrate. Check for staining periodically under dissecting microscope. 4. Stop reaction with PBS. Part 2. Fetal alcohol exposure. Maternal exposure to teratogens can cause developmental defects in the developing embryos. One of the most widespread human environmental teratogens is alcohol. Exposure in utero to excess alcohol can result in mental retardation, and a characteristic pattern of craniofacial defects known as fetal alcohol syndrome. The developing craniofacial regions undergo specific patterns of programmed cell death which are required for normal development. Exposure to alcohol has been shown to enlarge these regions, which accounts for the observed developmental defects. 1. Examine pregnant uteruses from control mice and from females that were given 2 or 3 injections of 0.5 ml 25% ethanol in saline on embryonic day 7.5. Make a note of resorption sites which indicate that an embryo has died. 2. Dissect out fetuses and examine for developmental defects, especially in the craniofacial region. References: Sulik, K.K., M.C. Johnson, and M.A. Webb (1981) Fetal alcohol syndrome: embryogenesis in a mouse model. Science 214: 936- 938. Sulik, K.K., C.S. Cook and W.S. Webster (1988) Teratogens and craniofacial malformations: relationships to cell death. Development 103Supplement: 213-232. Carlson, Chapter 15 Moore and Presaud, Chapter11