Identification and functional characterization of relaxin-type and pedal peptide/orcokinin-type neuropeptides in the starfish Asterias rubens

Abstract

Neuropeptides are neuronal signaling molecules that regulate many physiological and behavioural processes in vertebrates and invertebrates. Investigation of neuropeptide signaling in echinoderms (e.g. starfish) can provide insights into the evolution of neuropeptide systems because as deuterostomian invertebrates they occupy an “intermediate” phylogenetic position between vertebrates and protostomian invertebrates. Recent analysis of neural transcriptome data from the starfish Asterias rubens has identified 40 transcripts encoding neuropeptide precursors. Here the expression and function of neuropeptides derived from four of these precursors was investigated: relaxin-like gonad-stimulating peptide precursor (AruRGPP), relaxin-like peptide precursor 2 (AruRLPP2), pedal peptide-like neuropeptide precursors 1 and 2 (ArPPLNP1 and ArPPLNP2). AruRGP induces spawning of ovarian fragments from A. rubens. Analysis of the expression of AruRGPP in A. rubens using mRNA in situ hybridization revealed expression by cells in the radial nerve cords, circumoral nerve ring and tube feet. Furthermore, a band of AruRGPP-expressing cells was also identified in the body wall epithelium lining the cavity that surrounds the sensory terminal tentacle and optic cushion at the tips of the arms. Discovery of these cells is important because they are candidate physiological mediators for hormonal control of starfish spawning in response to environmental cues. Interestingly, AruRLPP2 is also expressed in the same region of the arm tip as AruRGPP but the physiological role(s) of AruRLP2 is not yet known. Analysis of the expression of ArPPLNP1 and ArPPLNP2 using mRNA in situ hybridization revealed a widespread pattern of expression in A. rubens. Furthermore, immunohistochemical localization of peptides derived from these precursors revealed immunostaining in neuronal processes innervating muscles. Consistent with this pattern of expression, peptides derived from ArPPLNP1 and ArPPLNP2 act as muscle relaxants in starfish. Interestingly, this contrasts with previous findings from protostomian invertebrates, where pedal peptide/orcokinin-type neuropeptides act as muscle contractants.