STAGING OF COMMITMENT AND DIFFERENTIATION OF AVIAN CARDIAC MYOCYTES

The present study establishes the earliest time point for commitment o f cardiac myocyte progenitors at gastrulation and determines the effec ts of bromodeoxyuridine (BrdU) on postgastrulated committed cardiac pr ogenitor cells at a molecular level. Using immunochemical and reverse transcription/polymerase chain reaction assays for cardiac muscle-spec ific gene expression, we found that while both pre- and postgastrulate d embryonic cells were capable of cardiogenic differentiation at high cell density, only postgastrulated cells exhibited the ability to diff erentiate at clonal density. These data indicate that while cardiac my ocyte differentiation of pregastrulated cells can occur in vitro, cell -cell interactions are necessary for this to happen. Only gastrulated cardiac progenitor cells are able to differentiate in the absence of c ell-cell interactions and are therefore both specified and committed. Next, by exposing postgastrulated committed cardiac progenitor cells f rom embryos at various stages to BrdU, we demonstrated that these cell s from the lateral-plate mesoderm vary in their ability to differentia te into cardiac myocytes in vitro. Differentiation of cardiac myocyte progenitor cells from stages 4 and 5 was completely blocked by BrdU, w hereas it was not blocked in cells from stages 7 and 8 and cells from stage 6 were varied in their reaction. Analysis of cardiac myogenesis in vivo revealed that cardiac progenitors acquire BrdU resistance as t hey migrate, postgastrulation, anteriorly along a rostrocaudal axis. T he results from these two experiments suggest that while pregastrulate d cells exhibit a limited ability to undergo cardiac myocyte different iation, only postgastrulated anterior lateral-plate mesoderm contains committed cardiac myocyte progenitors and that these committed progeni tors are not equivalent in their ability to differentiate. (C) 1994 Ac ademic Press, Inc.