We examined adenosine 5'-triphosphate (ATP), pertussis toxin (PT) and phorbol myristate acetate (PMA), a protein kinase C (PKC) activator, modulation of atrial natriuretic peptide (ANP)-stimulated cell-membrane guanylate cyclase (ANP-s-GC) activity and ANP stimulation of whole-cell cGMP accumulation (ANP-s-cGMP) in an ANP-receptor-transduction cell model, the human renal cell line (SK-NEP-1). Acute and long-term effects of PMA on PKC isotype activity are different: Acute (20-min) PMA activation of PKC inhibits ANP-s-cGMP and ANP-s-GC; whereas, long-term (36-h) PMA treatment inhibits slightly less by only partially down-regulating PKC activity, the type-III PKC isotype being 36-h resistant. Long-term 10(-7)M PMA treatment of cells neither affected membrane basal GC activity nor ANP-s-GC activity but partially inhibited ATP enhancement of ANP-s-GC. This partial inhibition was completely reversed by the PKC inhibitor H7 and a PKC inhibitory antibody but only partially reversed by the antibody to the catalytic domain of PKC type III. The EC50 for ATP and its non-phosphorylating analog ATP gamma S in the presence of acute PMA inhibition of ANP-s-cGMP was similar (approximately 10(-9)). This enhancement of PMA inhibition was two orders of magnitude more sensitive (EC50 10(-7)M) than inhibition of ANP-s-cGMP that we previously reported for acute PMA treatment of whole SK-NEP-1 cells. The three- to four-fold ATP enhancement of cell membrane ANP-s-GC was not blocked by 12-hour preincubation of cells with 150 ng/mL PT but was completely blocked if 2-x-10(-7)M PMA was then added for 20 minutes, indicating that acute activation of PKC by PMA does not require a functional "G-type" protein. Acute PMA inhibition of ANP-s-cGMP was reversed by permeabilizing SK-NEP-1 cells to a specific PKC inhibitory peptide, further confirming that PMA inhibition was mediated through PKC activation. These data demonstrated that ANP-s-GC and ANP-s-cGMP were modified through non-phosphorylating interactions with ATP, by multiple PMA activatable PKC isoforms, and that neither were affected by PT-sensitive guanine-nucleotide-binding (G)-protein(s).