Characterization, localization and temporal expression of crustacean hyperglycemic hormone (CHH) in the behaviorally rhythmic peracarid crustaceans, Eurydice pulchra (Leach) and Talitrus saltator (Montagu)

Crustacean hyperglycemic hormone (CHH) has been extensively studied in decapod crustaceans where it is known to exert pleiotropic effects, including regulation of blood glucose levels. Hyperglycemia in decapods seems to be temporally gated to coincide with periods of activity, under circadian clock control. Here, we used gene cloning, in situ hybridization and immunohistochemistry to describe the characterization and localization of CHH in two peracarid crustaceans, Eurydice pulchra and Talitrus saltator. We also exploited the robust behavioral rhythmicity of these species to test the hypothesis that CHH mRNA expression would resonate with their circatidal (12.4 h) and circadian (24 h) behavioral phenotypes. We show that both species express a single CHH transcript in the cerebral ganglia, encoding peptides featuring all expected, conserved characteristics of other CHHs. E. pulchra preproCHH is an amidated 73 amino acid peptide N-terminally flanked by a short, 18 amino acid precursor related peptide (CPRP) whilst the T. saltator prohormone is also amidated but 72 amino acids in length and has a 56 residue CPRP. The localization of both was mapped by immunohistochemistry to the protocerebrum with axon tracts leading to the sinus gland and into the tritocerebrum, with striking similarities to terrestrial isopod species. We substantiated the cellular position of CHH immunoreactive cells by in situ hybridization. Although both species showed robust activity rhythms, neither exhibited rhythmic transcriptional activity indicating that CHH transcription is not likely to be under clock control. These data make a contribution to the inventory of CHHs that is currently lacking for non-decapod species.