ORIGIN OF PRIMORDIAL GERM-CELLS THE PRESTREAK CHICK-EMBRYO

The temporal and spatial pattern of segregation of the avian germline from the formation of the area pellucida to the beginning of primitive streak formation (stages VII-XIV, EG&K) was investigated using the cu lture of whole embryos and central and peripheral embryo fragments on vitelline membranes at stages VII-IX, immunohistological analysis of w hole mount embryos and sections with monoclonal antibodies MC-480 agai nst stage-specific embryonic antigen-1 (SSEA-1) and EMA-1, and with th e culture of dispersed blastoderms at stages IX-XIV with and without a n STO feeder layer. Whole embryos at intrauterine stages developed up io the formation of the primitive streak despite the absence of area p ellucida expansion. Primordial germ cells (PGCs) appeared in the cultu res of whole embryos and only in central fragments containing a partia lly formed area pellucida al stages VII-IX. When individual stage IX-X IV embryos were dispersed and cultured without a feeder layer, 25-45 P GCs/embryo were detected only with stage X-XIV, but not with stage IX blastoderms. However, the culture of dispersed cells from the area pel lucida of stages IX-XIII on STO feeder layers yielded about 150 PGCs/e mbryo. The carbohydrate epitopes recognized by anti-SSEA-1 and EMA-1 f irst appeared at siege X on cells in association with polyingressing c ells on the ventral surface of the epiblast and later on the dorsal su rface of the hypoblast. The SSEA-1-positive hypoblast cells gave rise to chicken PGCs when cultured on a feeder layer of quail blastodermal cells. From these observations, we propose that the segregation and de velopment of avian germline is a gradual, epigenetic process associate d with the translocation of SSEA-1/EMA-1-positive cells from the ventr al surface of the area pellucida at stage X to the dorsal side of the hypoblast at stages XI-XIV. (C) 1996 Wiley-Liss, Inc.