The mechanism by which serotonin (5-HT3) receptors mediate a rise in cyclic-GMP level was investigated in a neuronal cell line. Inhibitors of phospholipase A2 (mepacrine) and of lipoxygenase (eicosatetraynoic acid or nordihydroguaiaretic acid) suppressed the action of serotonin. On the other hand, inhibition by hemoglobin indicates a role for nitric oxide which could be in part responsible for the cyclic-GMP effect as an intercellular stimulant. The suppression of the serotonin effect by the arginine analogues N omega-methyl-L-arginine and canavanine is consistent with the notion that nitric oxide could be released from arginine. The serotonin-induced rise of cyclic-GMP level depends on the presence of extracellular Ca2+ with half-maximal stimulation at 0.3 mM Ca2+. The serotonin-stimulated rise of cyclic GMP was inhibited by (a) addition of inorganic blockers of Ca2(+)-permeable channels (La3+, half-maximal inhibitory concentration (IC50) 0.04 mM; Mn2+, IC50, 0.4 mM; Co2+, IC50, 0.9 mM; Ni2+, IC50, 1.2 mM) and (b) of organic blockers (diltiazem: IC50, 6 microM, methoxyverapamil: IC50, 3 microM and (c) intracellular application of the Ca2+ chelator bis-(O-aminophenoxy)-ethane-N,N,N',N'-tetraacetic acid (IC50, 2 microM). Thus, two pathways for the activation of soluble guanylate cyclase by serotonin are possible: (a) via lipoxygenase products of arachidonic acid and/or (b) via nitric oxide or a related nitroso compound. Serotonin mediates a rise of cytosolic Ca2+ activity due to entry of extracellular Ca2+. It still has to be investigated which step depends on a rise of cytosolic Ca2+ activity that appears to be a prerequisite for activation of guanylate cyclase.