Internalization of atrial natriuretic factor by AtT‐20 corticotropin‐secreting cells

Because extended exposure of AtT‐20 corticotropin‐secreting cells to atrial natriuretic factor (ANF) results in a desensitization of ANF‐induced cGMP synthesis, we sought to establish whether pretreatment of AtT‐20 cells with the atrial peptide also led to an internalization process. In fact, by coupling an ultrastructural approach to cryoultramicrotomy, ANF‐immunoreactivity was detected at both the plasma membrane level and at intracellular sites in AtT‐20 cells. Internalization was observed within 5 min at which time labelling was observed in the plasma membrane level, in vacuole‐like structures in close proximity to the plasma membrane, in cytoplasmic matrix and sometimes in mitochondria. After 30 min exposure Golgi apparatus, mitochondria and nuclear euchromatin were also labelled. Following 1‐4 hr, labelling in other cell compartments, e.g. lysosomal, was increased, while it was reduced in plasma membranes and vacuole‐like structures. Secretory granules and endoplasmic reticulum were not labelled throughout the time course. Extraction of a intracellular [125I] ANF from AtT‐20 cells following 4 hr incubation suggested that about 90% of the peptide was intact. The data suggest that internalization of ANF may serve to terminate the biological response associated with ANF receptor activation; subcellular distribution of internalized, intact ANF suggests that the peptide may have other, as yet unidentified, intracellular actions.