Distinct properties of atrial natriuretic factor receptor subpopulations in epithelial and fibroblast cell lines

We have characterized two atrial natriuretic factor (ANF) receptor subtypes, designated ANF-R1 and ANF-R2, in two established cell lines that express exclusively one receptor subtype. The ANF-R1 receptor is selectively expressed by the kidney epithelial cell line LLC-PK1. It is a 130-kDa protein that has a much higher affinity for the biologically active forms of ANF than for its metabolites. The binding of ANF to this subtype is potentiated by amiloride and by divalent cations. The activation of the ANF-R1 receptor leads to an accumulation of cyclic GMP that is only partially inhibited by methylene blue. The ANF-R2 receptor, which is expressed selectively by the fibroblast cell line NIH-3T3, is a 130-kDa protein composed of two disulfide-linked subunits of 64-kDa. Activation of this subtype by saturating concentrations of ANF does not appear to elicit cyclic GMP production. However, supraphysiological concentrations of ANF induce a nonsaturable accumulation of cyclic GMP with an apparent ED50 in the high micromolar range. In contrast to the ANF-R1 subtype, the stimulation of cyclic GMP production is completely abolished by methylene blue. This subtype recognizes the active forms of ANF as well as its metabolites, and the binding is insensitive to amiloride and is decreased by divalent cations. These two cell lines can serve as models for studying the differential regulatory properties of ANF-R1 and ANF-R2 subtypes. In addition, we have also characterized the two ANF receptor subtypes in rat kidney glomeruli, where they show the same structure and pharmacological characteristics as in the two model cell lines.