Sustained exposure of GH4C1 rat pituitary cells to phorbol esters or the hypothalamic tripeptide thyrotropin-releasing hormone decreases the number of phorbol ester binding sites without changing affinity (down modulation; Jaken, S., Tashjian, A. H., Jr., and Blumberg, P. M. Cancer Res., 41: 2175-2181, 1981). In untreated control cultures, the concentration of receptors in lysates was not significantly different from the concentration in intact cells. In contrast, in down-modulated cultures, the concentration of receptors in lysates was greater than the concentration in the corresponding intact cells and instead was equal to the concentration in lysates of control cultures. Therefore, the processes of both homologous (phorbol ester-induced) and heterologous (thyrotropin-releasing hormone-induced) phorbol ester receptor down modulation involves the generation of a cryptic receptor state that is revealed upon cell lysis. Furthermore, comparison of receptor properties in cells and lysates revealed that the affinity of the receptor for phorbol dibutyrate was decreased from a Kd of 11.1 +/- 0.6 nM (S.D.) in intact cells to 40 +/- 10 nM in cell lysates. Binding of the receptor for the structurally related diterpene, mezerein, fit a one-component model in intact cells, but was better fit to a two-component model in cell lysates. That is, the apparently homogeneous phorbol 12,13-dibutyrate receptor population in cell lysates appeared heterologous with respect to mezerein binding. This is not due to a preferential recovery of a subset of receptors in the lysates, because no substantial loss of receptor number was associated with cell lysis. Instead, these results suggest that the affinity and ligand specificity of the phorbol ester receptor may depend on its cellular environment.