Pituitary defects ranging from panhypopituitarism (total loss of function) to loss of a single hormone secreting population are not uncommon in fetal development, with the most frequent single hormone deficiency, loss of growth hormone, affecting 1 in 4000 newborns. Studies on the origin and patterning mechanisms of the early presumptive adenohypophysis (anterior pituitary gland) in chick and other model systems, prior to formation of Rathke's pouch (the pituitary anlage), are highly limited, inconclusive and contradictory. The developing chick adenohypophysis is an excellent model system to study the complex temporal and spatial patterning events, tissue interactions and signaling mechanisms that occur in ectodermal primordia during embryogenesis. High-resolution fate mapping, using multiple fluorescent dyes and time-lapse videomicroscopy, together with detailed spatial and temporal analyses of gene expression patterns, using in situ hybridization (ISH), will be used to determine conclusively the neural/non-neural origins of the ectodermal cells that form the adenohypophysis. Then, to establish the timing and extent of specification of the populations of ectodermal cells that form the adenohypophysis, and the inductive tissue interactions that lead to specification, individual candidate tissues will be cultured in collagen gels. ISH using a number of marker genes following culture will elucidate the state of specification of the contributing tissue populations in embryos from the beginning of neuralization. Tissue recombinants cultured in collagen gels then will be used to investigate the inductive tissue interactions leading to specification. Lastly, the molecular mechanisms responsible for the specification of cells to an adenohypophyseal fate will be investigated. Preliminary studies implicate a novel pathway involved in adenohypophyseal cell specification, the Insulin/Insulin-like Growth Factor pathway. Following detailed analyses of gene expression and protein distribution, by ISH and immunocytochemistry, respectively, of Insulin family members, we will conducts experiments using both gain and loss of function approaches to begin to elucidate the molecular mechanisms involved in the sequential signaling events underlying specification of the adenohypophysis.