Comparative Analysis of Neuropeptides in Homologous Interneurons and Prohormone Annotation in Nudipleuran Sea Slugs

Abstract

Despite substantial research on neuronal circuits in nudipleuran gastropods, few peptides have been implicated in nudipleuran behavior. In this study, we expanded the understanding of peptides in this clade, using three species with well-studied nervous systems, Hermissenda crassicornis, Melibe leonina, and Pleurobranchaea californica. For each species, we performed sequence homology analysis of de novo transcriptome predictions to identify homologs to 34 of 36 prohormones previously characterized in the gastropods Aplysia californica and Lymnaea stagnalis. We then used single-cell mass spectrometry to characterize peptide profiles in homologous feeding interneurons: the multifunctional ventral white cell (VWC) in P. californica and the small cardioactive peptide B large buccal (SLB) cells in H. crassicornis and M. leonina. The neurons produced overlapping, but not identical, peptide profiles. The H. crassicornis SLB cells expressed peptides from homologs to the FMRFamide (FMRFa), small cardioactive peptide (SCP), LFRFamide (LFRFa), and feeding circuit activating peptides prohormones. The M. leonina SLB cells expressed peptides from homologs to the FMRFa, SCP, LFRFa, and MIP-related peptides prohormones. The VWC, previously shown to express peptides from the FMRFa and QNFLa (a homolog of A. californica pedal peptide 4) prohormones, was shown to also contain SCP peptides. Thus, each neuron expressed peptides from the FMRFa and SCP families, the H. crassicornis and M. leonina SLB cells expressed peptides from the LFRFa family, and each neuron contained peptides from a prohormone not found in the others. These data suggest each neuron performs complex co-transmission, which potentially facilitates a multifunctional role in feeding. Additionally, the unique feeding characteristics of each species may relate, in part, to differences in the peptide profiles of these neurons. These data add chemical insight to enhance our understanding of the neuronal basis of behavior in nudipleurans and other gastropods.

Egg-laying hormone expression in identified neurons across developmental stages and reproductive states of the nudibranchBerghia stephanieae

“…Although conserved across molluscs including gastropods and bivalves (De Oliveira et al, 2019), there has been no indication of ELH in the nudipleuran gastropods despite recent multi-species transcriptomics studies (Lee et al, 2021). However, there is evidence for the existence of an ELH-like hormone.…”

In the sea slug Melibe leonina the posterior nerves communicate stomach distention to inhibit feeding and modify oral hood movements

“…However, unlike most heterobranchs Melibe lacks a buccal mass and complicated chewing mechanics, leading to an exceptionally simple (∼30 neurons) buccal ganglion ( Trimarchi and Watson, 1992 ). Finally, it is an organism about which we have transcriptomic ( Cook et al, 2018 ) and peptidomic ( Lee et al, 2021 ) information, and whose daily rhythms have been thoroughly documented ( Newcomb et al, 2014 ).…”