The specific binding sites of eyestalk- and pericardial organ-crustacean hyperglycaemic hormones (CHHs) in multiple tissues of the blue crab,Callinectes sapidus

Abstract

Crustacean hyperglycaemic hormone from the pericardial organ (PO-CHH) is a CHH-related neuropeptide but its function and target tissues are not known in crustaceans. To investigate this issue, we employed radiolabelled ligand binding and cGMP assays, using eyestalk-CHH (ES-CHH) as a reference neuropeptide. The membranes were prepared from various tissues of Callinectes sapidus: hepatopancreas, hindgut, midgut, gills, heart, abdominal muscles and scaphognathites. Like ES-CHH, recombinant PO-CHH (rPO-CHH) specifically bound to the membranes of scaphognathites=abdominal muscles>midgut>gills> heart>hindgut and hepatopancreas (list order corresponds to the number of binding sites). The specific binding sites of (125)I-ES-CHH in hepatopancreas and gills were saturable and displaceable. The abdominal muscle membrane binding sites were specific and saturable to both CHHs. These binding sites were displaced by homologous neuropeptides, but poorly displaced by the heterologous counterpart. As for the second messenger, the expected increment (3- to >20-fold) in the amount of cGMP produced by ES-CHH was noted in most tissues tested except midgut. Recombinant PO-CHH increased cGMP production 1.5- to 4-fold in scaphognathites, heart, midgut, hindgut and abdominal muscles. The results obtained from the binding study suggest that PO-CHH also has multiple target tissues of which abdominal muscles and scaphognathites are the primary ones. The differences in the primary amino acid sequences of PO-CHH and ES-CHH, particularly in the C-terminal region and in the amidation at C-terminus, may contribute to the truncated responses of hyperglycaemia, cGMP stimulation and binding affinity.