Salivary electrolyte secretion is under the control of the autonomic nervous system. In this paper we report that HSY, an epithelial cell line derived from the acinar-intercalated duct region of the human parotid gland, responds to muscarinic-cholinergic (generation of Ca2+ signal) and beta-adrenergic (generation of cAMP signal), but not to alpha-adrenergic (lack of Ca2+ signal), receptor stimulation. The muscarinic response was studied in detail. Carbachol (10(-4) M, muscarinic agonist) or A23187 (5 microM, calcium ionophore) stimulation of HSY cells increases both 86Rb (K+) influx and efflux, resulting in no change in net equilibrium 86Rb content. Atropine (10(-5) M, muscarinic antagonist) blocks both the carbachol-generated Ca2+ signal and carbachol-stimulated 86Rb fluxes, but has no effect on either the A23187-generated Ca2+ signal or A23187-stimulated 86Rb fluxes. Carbachol- and A23187-stimulated 86Rb fluxes are substantially inhibited by two K+ channel blockers, quinine (0.3 mM) and scorpion venom containing charybdotoxin (33 micrograms/ml). The inhibition of these stimulated fluxes by another K+ channel blocker, tetraethylammonium chloride (5 mM), is less pronounced. Protein kinase C (PKC) seems to be involved in the regulation of the 86Rb fluxes as 10(-7) M PMA (phorbol ester, phorbol-12-myristate-13-acetate) substantially inhibits the muscarinic-stimulated 86Rb efflux and influx. Because this concentration of PMA totally inhibits the carbachol-generated Ca2+ signal and only 80% of the muscarinic-stimulated 86Rb influx, it seems that a portion of the carbachol-stimulated 86Rb flux (i.e. that portion not inhibited by PMA) might occur independently of the Ca2+ signal. PMA fails to inhibit the A23187-stimulated 86Rb fluxes, however, suggesting that PKC regulates Ca(2+)-sensitive K+ channel activity by regulating the Ca2+ signal, and not steps distal to this event. 4-alpha-Phorbol-12,13-didecanoate, a phorbol ester which fails to activate PKC, fails to inhibit either the carbachol-stimulated increase in intracellular free Ca2+, or carbachol-stimulated 86Rb fluxes.