Characterization of binding of the annelidan myoactive peptides, GGNG peptides, to tissues of the earthworm, Eisenia foetida

The GGNG peptides are myoactive peptides which we have isolated from several species of annelids. Two types of peptides have been isolated as GGNG peptides: earthworm excitatory peptides (EEP) and leech excitatory peptide (LEP). The EEP and LEP are highly homologous with each other but act specifically on earthworms and leeches, respectively. To approach the mechanism of the specific action, we attempted to characterize the receptors for EEP in the earthworm Eisenia foetida, from which EEP have been isolated, by a radioreceptor assay. To prepare a radiolabeled ligand, we first coupled EEP to Bolton-Hunter reagent and then labeled it with ¹²⁵I ([¹²⁵I] BH-EEP). Binding of the [¹²⁵I] BH-EEP to the membrane preparation of the earthworm tissue (crop-gizzard) was reversible, saturable, and specific with a Kd of 4.9 ± 1.2 nM and a B(max) of 15.9 ± 2.0 fmol/mg wet tissue. For the earthworm membrane preparation, EEP showed a much greater potency than LEP in displacing [¹²⁵I] BH-EEP, suggesting that the earthworm tissues contain EEP-specific receptors. This result would account for the specific biological activity of EEP and LEP on the earthworm and leech tissues, respectively. The binding capacity was high in the anterior part of digestive tract including the esophagus, crop and gizzard, and the nephridia. From these results, together with the observed myotropic activity of EEP on gut tissues, it is assumed that EEP is physiologically involved in regulation of the gut motility in the earthworm E. foetida.