ISJ 12: 173-175, 2015 ISJ 12: 173-175, 2015 ISSN 1824-307X LETTER TO EDITOR On the existence of possible pituitary adenylate cyclase-activating polypeptide adenylate type 1 receptor in earthworms L Molnár1, E Pollák1, I Somogyi1, P Engelmann2 1Department of Comparative Anatomy and Developmental Biology, Faculty of Sciences, University of Pécs, Ifjúság u. 6, H-7624 Pécs, Hungary 2Department of Immunology and Biotechnology, Clinical Center, University of Pécs, Szigeti u. 12, H-7643 Pécs, Hungary Accepted May 29, 2015 To the Editor Neuropeptides, expressed by both invertebrate and vertebrate nervous systems (Grimmelikhuijzen and Hauser, 2012) play critical roles in both synaptic and non-synaptic signalling mechanisms regulating the functions of nervous, endocrine and immune systems (Nässel and Larhammar, 2013). In contrast to vertebrates the invertebrate nervous system contains high number of neurosecretory cells that produce and elaborate several neurohormones (most of them are peptides) that act on peripheral tissues e.g., body wall muscles, alimentary canal, excretory and genital organs (Taghert and Nitabach, 2012). However, some of the neuropeptides are neuromodulators in the central nervous system mediating the firing mechanism and synaptic transmission of neurons (van den Pol, 2012). While broad range of neuropeptides is known from some invertebrates like planarians, roundworms, insects or molluscs (De Haes et al., 2014), but less is known from the annelids, especially from earthworms. Most of the neuropeptides, like proctolin, FMRFamide-like peptides, neuropeptide Y (Lengvari et al., 2001), substance P and ACTH-like peptide (Aros et al., 1980), believed to be regulator molecules in earthworms had only been identified by immunocytochemical stainings. There are only a few neuropeptides, e.g., CAPA-peptides (Herbert et al., 2009) and pituitary adenylate cyclase activating polypeptide (PACAP)-like peptides (Somogyvári- Vígh et al., 2000) which expression, pattern and function have investigated by various experimental protocols in details. PACAP is a highly conserved member of the VIP/secretin/glucagon peptide family found in neuronal elements of both the CNS and several peripheral tissues of vertebrates. It acts as a pleiotropic neuropeptide via three heptahelical G- protein-linked receptors, one PACAP-specific (PAC1) ___________________________________________________________________________ Corresponding author: Peter Engelmann Department of Immunology and Biotechnology Clinical Center, University of Pécs Pécs, H-7643, Szigeti u. 12, Hungary E-mail: engelmann.peter@pte.hu receptor and two receptors that are shared with VIP (VPAC1 and VPAC2). PACAP has been implicated in a variety of central nervous functions, including hypophysiotropic function, learning and memory formation, psychomotor function, and the response to nerve injury. Furthermore, it is a potent antiapoptotic and neurotrophic substance (Vaudry et al., 2009). By means of immunohistochemistry PACAP- like activity was identified in the central and peripheral nervous system of some earthworm species, namely Eisenia fetida, Lumbricus terrestris and L. polyphemus (Reglödi et al., 2000). Two distinct forms of PACAP-like molecules (consist from 27 or 38 amino acids) were found in the CNS of L. polyphemus of which the smaller molecule proved to be the predominant form (Somogyvári- Vígh et al., 2000). Applying whole mount preparations to immunohistochemical observations Molnár and colleagues (2006) showed that each PACAP-like molecules occurred in distinct neuron populations of both L. terrestris and E. fetida and concluded that these molecules might mediate distinct physiological processes in earthworms. The biological activity of the purified HPLC fractions of both PACAP27 and PACAP38-like peptides isolated from the earthworm CNS was investigated in in vitro experimental conditions. Both PACAP-like molecule fractions from earthworms have activated adenylate cyclase and possessed sequence similarities to mammalian PACAP isoforms (Somogyvári-Vígh et al., 2000). Occurrence, distribution and cellular transport of the specific PAC1-receptor in neural structures of the ventral nerve cord ganglia in E. fetida was investigated by light and electron microscopic immunocytochemistry. The experimental results revealed that PAC1-receptor-like immunoreactivity localized both on intracellular membranes (endoplasmic reticulum cisternae) and special domains of plasma membrane (synaptic membrane) of neurons as well. It suggests a compound transport and presentation process of the receptor molecule on the cell surface which supposes its role in synaptic transmission (Molnár et al., 2008). 173   mailto:engelmann.peter@pte.hu A possible role of PACAP-like peptides in mediation of both embryogenesis (Boros et al., 2008) and posterior regeneration of E. fetida (Várhalmi et al., 2008) was suggested based on the quantitative analysis of the expression of PACAP- like peptides applying one of the most sensitive immunoserological methods, the radioimmunoassay. During both the embryonic development and segment regeneration the overexpression of PACAP-like proteins was detected and a characteristic antero-posterior gradient found in regenerating ventral nerve cord ganglia suggesting that they had a strong influence on cell cycle and tissue differentiation in earthworms (Boros et al., 2008, Várhalmi et al., 2008). The expression of PAC1-receptor was investigated by immunocytochemical and immunoserological methods (Western blot and Far Western blot) during the embryonic development of E. fetida and showed its existence in germinal layers and developing ventral nerve cord ganglia (Boros et al., 2010). According to the PACAP and PAC1/VPAC receptor localization studies in vertebrates it was clear that their expression is not restricted only to neuroendocrine and separate vegetative organs (Vaudry et al., 2009). We observed PAC1-like immunoreactivity in coelomocytes by means of immunocytochemistry and Western blot. Among coelomocytes the amoebocyte subgroup expressed more abundantly this receptor, but whether this immunocyte is only a responder and/or a producer of the neuropeptide was not clear. To answer this, first immunoserological experiments (RIA) were performed from coelomocytes of regenerating earthworms and proved the PACAP-like immunoreactivity in this immune cell compartment as well (Somogyi et al., 2009). These immunological cross-reaction based results should be further supported by molecular biological approaches to isolate and characterize the mRNA transcripts for PACAP and PAC1 receptor from earthworms. Recent molecular evidence from other phyla (Pirger et al., 2010; Lugo et al., 2013) and our findings strengthen the notion for evolutionary conservation of this neuropeptide family across animal kingdom. 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