Cytochemical localization of adenylate cyclase in the limb buds of Bufo bufo

The importance of cyclic nucleotides in the regulation of the processes of differentiation and embryonic development is known. The possible role that cyclic adenosine monophosphate (cAMP) plays during the development of the posterior limb of Bufo bufo is studied by the cytochemical localization of adenylate cyclase (AC), an enzyme that catalyzes the synthesis of the cyclic nucleotide. The method is based on the reaction between the enzyme AC and its specific substrate AMP-PNP (5'-adenylylimidodiphosphate) in the presence of lead. The lead precipitates that form as secondary reaction products are evidence of enzymatic activity. Reaction products are present only at the epithelial level in the limb bud; initially, such products are visible only at the base of the bud, particularly on the epithelial fascia located at the boundary with the body. During successive elongation and toe formation, AC activity is only present on the cells of the proximal portion of each new segment. Enzymatic activity is never present in correspondence to the ectodermal apical crest. cAMP is probably not involved in the processes of cellular proliferation but, rather, in the processes of inducing differentiation of the internal mesenchymal cells.