Genes encoding proteins orthologous to members of the neuropeptide Y (NPY) and NPY receptor family occur in the genomes of Drosophila melanogaster and Anopheles gambiae. For insects, distinct peptides implicated in receptor activation include full-length NPY-like molecules, termed neuropeptide F (NPFs), and shorter peptides, termed short neuropeptide F (sNPFs). Full-length NPFs possessing 36 amino acids are structurally related to NPY, pancreatic polypeptide (PP), and peptide YY (PYY), which occur in vertebrates. Functions for NPY, PP, and PYY include regulation of food intake, circadian rhythms, gut enzyme secretion and motility; NPF is known to affect feeding in D. melanogaster. For genes encoding dipteran sNPFs, putative precursors contain multiple candidate peptides with characteristic C-terminal sequences of either LRLRFamide or RLRWamide. The insect NPF receptors (NPFRs), like their vertebrate counterparts, have predicted features typical of G protein-coupled receptors. Gene structures of the NPFs and their receptors are conserved between these dipterans. Our approach of cloning and stably expressing NPFRs in mammalian cells allows for the identification of cognate ligand-receptor pairs via radioreceptor binding assays. The localization of NPFs and NPFRs in midgut and CNS resembles their vertebrate counterparts and suggests roles for NPFs in feeding and reproduction in Diptera and perhaps other insects.