THE DEVELOPMENTAL ORIGIN OF PRIMORDIAL GERM-CELLS AND THE TRANSMISSION OF THE DONOR-DERIVED GAMETES IN MIXED-SEX GERMLINE CHIMERAS TO THE OFFSPRING IN THE CHICKEN

A novel system has been developed to determine the origin and developm ent of primordial germ cells (PGCs) in avian embryos directly. Approxi mately 700 cells were removed from the center of the area pellucida, t he outer of the area pellucida, and the area opaca of the stage X blas toderm (Eyal-Giladi and Kochav, 1976; Dev Biol 49:321-337). When the c ells were removed from the center of the area pellucida, the mean numb er of circulating PGCs per 1 mu l of blood was significantly decreased to 13 (P < 0.05) in the embryo at stage 15 (Hamburger and Hamilton, 1 951: J Morphol 88:49-92) as compared to intact embryos of 51. When the removed recipient cells from the center of the area pellucida were re plenished with 500 donor cells, no reduction in the PGC number was obs erved. The removal of cells from the outer of area pellucida or from t he area opaca had no effect on the number of PGCs. When another set of the manipulated embryos were cultured ex vivo to hatching and reared to sexual maturity, the absence of germ cells and the degeneration of seminiferous tubules were observed in resulting chickens derived from the blastoderm from which the cells were removed from the center of th e area pellucida. Chimeric embryos produced by the male donor cells an d the female recipient contained the female-derived cells at 97.2% in the whole embryo and 94.3% in the erythrocytes at 5 days of incubation . At 5-7 days of incubation, masculinization was observed in about one half of the mixed-sex embryos. The proportions of the female-derived cells in the whole embryo and in the erythrocytes were 76.5% and 80.2% at 7 days to 55.7% and 62.5% at 10 days of incubation, respectively. When the chimeras reached their sexual maturity, they were test mated to assess donor contribution to their germline. Five of six male chime ras (83%) and three of five female chimeras (60%) from male donor cell s and a female recipient embryo from which 700 cells at the center of area pellucida were removed were germline chimeras. Three of the five male germline chimeras (60%) and one of the three female germline chim eras (33%) transmitted exclusively (100%) donor-derived gametes into t he offspring. When embryonic cells were removed from the outer of area pellucida or area opaca, regardless of the sex combination of the don or and the recipient, the transmission of the donor-derived gametes wa s essentially null. The findings in the present studies demonstrated, both in vivo and in vitro, that the PGCs originate in the central part of the area pellucida and that the developmental fate to germ cell (P GCs) had been destined at stage X blastoderm in chickens. (C) 1997 Wil ey-Liss, Inc.