Neuropeptide F is the most abundant neuropeptide in parasitic flatworms and is analogous to vertebrate neuropeptide Y. This paper examines the effects of neuropeptide F on tetrathyridia of the cestode Mesocestoides vogae and provides preliminary data on the signalling mechanisms employed. Neuropeptide F (>/=10 microM) had profound excitatory effects on larval motility in vitro. The effects were insensitive to high concentrations (1 mM) of the anaesthetic procaine hydrochloride suggesting extraneuronal sites of action. Neuropeptide F activity was not significantly blocked by a FMRFamide-related peptide analog (GNFFRdFamide) that was found to inhibit GNFFRFamide-induced excitation indicating the occurrence of distinct neuropeptide F and FMRFamide-related peptide receptors. Larval treatment with guanosine 5'-O-(2-thiodiphosphate) trilithium salt prior to the addition of neuropeptide F completely abolished the excitatory effects indicating the involvement of G-proteins and a G-protein coupled receptor in neuropeptide F activity. Addition of guanosine 5'-O-(2-thiodiphosphate) following neuropeptide F had limited inhibitory effects consistent with the activation of a signalling cascade by the neuropeptide. With respect to Ca(2+) involvement in neuropeptide F-induced excitation of M. vogae larvae, the L-type Ca(2+)-channel blockers verapamil and nifedipine both abolished neuropeptide F activity as did high Mg(+) concentrations and drugs which blocked sarcoplasmic reticulum Ca(2+)-activated Ca(2+)-channels (ryanodine) and sarcoplasmic reticulum Ca(2+) pumps (cyclopiazonic acid). Therefore, both extracellular and intracellular Ca(2+) is important for neuropeptide F excitation in M. vogae. With respect to second messengers, the protein kinase C inhibitor chelerythrine chloride and the adenylate cyclase inhibitor MDL-2330A both abolished neuropeptide F-induced excitation. The involvement of a signalling pathway that involves protein kinase C was further supported by the fact that phorbol-12-myristate-13-acetate, known to directly activate protein kinase C, had direct excitatory effects on larval motility. Although neuropeptide F is structurally analogous to neuropeptide Y, its mode-of-action in flatworms appears quite distinct from the common signalling mechanism seen in vertebrates.