Bovine chromaffin cells contain high affinity NPY binding sites coupled through a pertussis toxin-sensitive G protein to inhibition of cAMP accumulation. NPY alone does not alter [3H]inositol phosphate formation from [3H]phosphoinositides in these cells. Increasing NPY concentrations, in the presence of ATP (300 microM), produced a dose-dependent enhancement in [3H]-inositol phosphate formation, EC50 = 3.2 nM. Inclusion of the selective NPY-Y1 receptor antagonist BW1229 (1 microM) produced a marked decrease in NPY potency (EC50 = 3.3 microM). The Y1 receptor agonist, [Leu31, Pro34]-NPY, was equally effective with NPY, whereas NPY18-36, a Y2 receptor agonist, was much less effective. Inclusion of NPY with ATP also produced an enhancement in the release of intracellular Ca2+. The ability of NPY to enhance both [3H]inositol phosphate formation and the release of intracellular Ca2+ was pertussis toxin-insensitive. NPY action on bovine chromaffin cell receptor(s) appears to facilitated by different G proteins: one which can inhibit cAMP accumulation via a pertussis toxin-sensitive process and another which can enhance ATP activation of the inositol phosphate signaling pathway by a pertussis toxin-insensitive process.