Allatostatin-C signaling in the crab <i>Carcinus maenas</i> is implicated in the ecdysis program

The allatostatin (AST) family of neuropeptides are widespread in arthropods. The multitude of structures and pleiotropic actions reflect the tremendous morphological, physiological and behavioral diversity of the phylum. Regarding the AST-C (with C-terminal PISCF motif) peptides, crustaceans commonly express three (AST-C, AST-CC and AST-CCC) that have likely arisen by gene duplication. However, we know little regarding their physiologically relevant actions. Here, we functionally characterize the cognate receptor for AST-C and AST-CC, determine tissue expression, and comprehensively examine the localization of AST mRNA and peptide. We also measured peptide release, circulating titers and performed bioassays to investigate possible roles. AST-C and AST-CC activate a single receptor (AST-CRd), but this, and other candidate receptors, were not activated by AST-CCC. Whole-mount in situ hybridization and hybridization chain reaction fluorescent in situ hybridization complemented neuropeptide immunolocalization strategies and revealed extensive expression of AST-Cs in the central nervous system. AST-C or AST-CCC expressing neurons were found in the cerebral ganglia, but AST-CC expression was never observed. Of note, we infer that AST-C and AST-CC are co-expressed in every neuron expressing crustacean cardioactive peptide (CCAP) and bursicon (BURS); all four peptides are released from the pericardial organs during a brief period coinciding with completion of emergence. In contrast to other studies, none of the AST-C peptides exhibited any effect on ecdysteroid synthesis or cardiac activity. However, expression of the AST-C receptor on hemocytes suggests a tantalizing glimpse of possible functions in immune modulation following ecdysis, at a time when crustaceans are vulnerable to pathogens.