This proposal examines the origin, differentiation, and morphogenesis of the ossified craniofacial skeleton in the oriental Fire-bellied Toad, Bombina orientalis. It entails (1) documenting the extent of cranial neural crest contribution to the osteocranium, and (2) identifying the roles of four factors that influence the timing and sequence of bone formation, and subsequent skeletal growth and morphogenesis, and other vertebrates. These factors are temperature, epithelial-mesenchymal tissue interactions, thyroid hormone, and the eye. Methods include: identifying results of ablation of restricted portions of the embryonic neural crest on subsequent cranial osteogenesis; quantifying differences in the timing and sequence of ossification among specimens reared at different constant temperatures; documenting the ability of cranial mesenchyme to initiate osteogenesis both in the absence of overlying epidermis and in the presence of trunk (i.e., noncranial) epidermis; examining patterns of ossification in larvae reared in various concentrations of thyroxine; analyzing effects of unilateral removal of the eye on bone growth and morphogenesis, and on adult skull size and shape. Some of these analyses will consider the entire complement of skull bones; others will focus primarily on the frontoparietal, an intramembranous bone that is homologous to the frontal and parietal of other vertebrates. Analytical procedures include whole-mount and serial-section histology of preserved specimens, in vivo bone labelling with Alizarin Red S, and computer-aided, three-dimensional reconstruction of skeletal morphology. Diagnosis of the developmental bases of human congenital skull malformations relies largely on the study of nonhuman vertebrates. An understanding of the mechanisms that control cranial ossification during anuran metamorphosis can serve as a """"""""model system"""""""" for the study of craniofacial skeletal development in vertebrates, including man.
