1. The morphology and innervation of the midgut (intestine) in the horseshoe crab, Limulus polyphemus was investigated. The organization of this tissue was examined with routine histology. Radioimmunoassay, immunohistochemistry and high performance liquid chromatography were employed to detect, localize and identify peptidergic innervation of the midgut. The actions of synthetic and native
proctolin-like and
FMRFamide-like
peptides were compared on the isolated midgut preparation. 2. Levels of
proctolin and
FMRFamide were determined in extracts of Limulus midgut tissue using radioimmunoassay. High levels of
proctolin-like immunoreactivity (69.5 +/- 11.3 ng/g) were detected, while levels of
FMRFamide-like immunoreactivity (0.8 +/- 0.2 ng/g) were less.
Proctolin levels were equally distributed, while the levels of
FMRFamide-like immunoreactivity exhibited an anterior bias. 3.
Proctolin- and
FMRFamide-like immunoreactivities in the Limulus midgut were localized with immunohistochemistry.
Proctolin- and
FMRFamide-immunoreactive elements were detected in intestinal nerve branches and individual fibers running along the surface of the midgut in whole-mount preparations. In sectioned tissue, staining for these
peptides was observed throughout the midgut, typically associated with muscle bands and fibers. Only a few immunoreactive cell bodies were observed. 4.
Proctolin, and several
FMRFamide-like
peptides produced distinct and opposing actions on the isolated Limulus midgut preparation.
Proctolin elicited
contracture and rhythmic contractions of this tissue, while
FMRFamide and N-terminally extended analogs of
FLRFamide relaxed gut tension.
FMRFamide-like
peptides partially reversed the excitatory actions of
proctolin. 5.
Proctolin- and
FMRFamide-like
peptides in Limulus midgut extracts were partially characterized with high performance liquid chromatography. One peak of
proctolin-like activity was detected on a linear gradient of 18 to 31.5%
acetonitrile. The native
proctolin-like
peptide produced excitatory actions on the isolated midgut preparation which were indistinguishable from those produced by synthetic
proctolin. Several peaks of
FMRFamide-like bioactivity (Busycon radula protractor muscle assay) were detected with a linear gradient of 5 to 30%
acetonitrile. Fractions from two distinct peaks produced
FMRFamide-like inhibitory effects on the isolated Limulus midgut preparation. These findings suggest a role for
proctolin-like and
FMRFamide-like
peptides as regulators of intestinal motility in Limulus.