"Leaky" epidermal cells contain a complete receptor-mediated adenylate cyclase system with an accessible GTP...

Hormone sensitivity to epinephrine or histamine of the adenylate cyclase system in pig skin is very labile to homogenization. We have developed a new adenylate cyclase receptor-mediated assay system with trypsinized epidermal cells which are treated with hypotonic shock. This new assay system maintained the hormonal sensitivity, as both epinephrine and histamine clearly stimulated cyclic AMP production. Moreover, the Ka for each hormone on this system was similar to that obtained from the floating pig skin slice system. Receptor-adenylate cyclase unit (coupling) in this assay system is therefore preserved as it occurs in intact tissue or cells. Because of the "leaky" nature of our preparation, phosphorylated compounds such as GTP and its analogue and NaF can penetrate the cell membrane and stimulate cyclic AMP production. In this system refractoriness is still maintained to subsequent stimulation by a receptor activator, and cholera toxin can be shown to dramatically increase the activity of GTP on the GTP binding protein, presumably by preventing GTP hydrolysis.