Transfilter evidence for a zone of polarizing activity participating in limb morphogenesis

Barriers were inserted into stage-20 HH chick embryo wing buds to separate the zone of polarizing activity from the anterior two-thirds of the wing bud with its overlying apical ectodermal ridge. Half of the barrier length projected out of the wing bud at insertion. Sham-operated wing buds developed only occasionally into wings with cartilage deletions. After insertion of an impermeable membrane (Cellophane), the typical wing skeleton contained only a humerus and a radius. In order to differentiate between diffusion, cell contact and cell penetration, Nuclepore filters with pore sizes of 0.05 micrometer 1.0 micrometer and 8.0 micrometer, respectively, were inserted. The typical wing skeleton after Nucleopore filter insertion was one with post-axial deletions. None, however, developed with complete distal deletions as after Cellophane. Deletions in the wing skeletons after Nuclepore insertion were the least with 1.0 micrometer filters and the most with 8.0 micrometer filters. Elevation of the apical ectodermal ridge was noted until 18 h after the insertions. In none of the groups did the ridge flatten. The results suggest that the zone of polarizing activity does have a role in normal limb morphogenesis. The mechanism by which its morphogen spreads is diffusion rather than being mediated via cell contacts.