The copepod crustacean Calanus finmarchicus plays a critical role in the ecology of the Gulf of Maine and other regions of the North Atlantic. To increase our understanding of the physiology of this species, a normalized, whole organism cDNA library was constructed, and expressed sequence tags (ESTs) of the clones were generated. Among these ESTs was one with homology to known cDNAs encoding prepro-A-type allatostatins (A-type ASTs), a well-known family of arthropod peptides that regulate juvenile hormone production in insects. Sequence analysis of the clone from which the EST was generated, with subsequent translation of its open reading frame, showed it to encode five novel A-type ASTs, whose mature structures were predicted to be APYGFGIamide, pE/EPYGFGIamide, ALYGFGIamide, pE/EPYNFGIamide, and pQ/QPYNFGVamide. Each of the peptides is present as a single copy within the prepro-hormone with the exception of APYGFGIamide, which is present in three copies. Surprisingly, the organization of the Calanus prepro-A-type AST, specifically the number of encoded A-type peptides, is more similar to those of insects than it is to the known decapod crustacean prepro-hormones. Moreover, the Calanus A-type ASTs possess isoleucine or valine residues at their carboxy (C)-termini rather than leucine, which is present in most other family members. Wholemount immunohistochemistry suggests that six pairs of somata produce the native Calanus A-type ASTs: five in the protocerebrum and one in the suboesophageal region. To the best of our knowledge, our report is the first characterization of a neuropeptidergic system in a copepod, the first identification of A-type ASTs from a non-decapod crustacean, the first report of crustacean A-type ASTs possessing isoleucine C-terminal residues, and the first report from any species of an A-type peptide possessing a valine C-terminal residue.