THE CHOLINERGIC, SEROTONINERGIC AND PEPTIDERGIC COMPONENTS OF THE NERVOUS-SYSTEM OF MONIEZIA-EXPANSA (CESTODA, CYCLOPHYLLIDEA)

The central (CNS) and peripheral (PNS) nervous systems of the cyclophyllidean tapeworm, Moniezia expansa, were examined for the presence of cholinergic, serotoninergic and peptidergic elements using enzyme cytochemical and immunocytochemical techniques in conjunction with light and confocal scanning laser microscopy. Cholinesterase activity and 5-hydroxytryptamine- and regulatory peptide-immunoreactivities (IRs) were localized to the nerve fibres and cell bodies of all of the major neuronal components in the CNS of the worm, including the cerebral ganglia and connecting commissure, the 10 longitudinal nerve cords and associated transverse ring commissures. Although each of the 3 systems appeared well developed and comprised a significant portion of the nervous system, the serotoninergic constituent was the most highly developed, consisting of a vast array of nerve fibres and cell bodies distributed throughout the strobila of the worm. A close association of cholinesterase reactivity and peptide-IRs was evident throughout the CNS, indicating the possible co-localization of acetylcholine and neuropeptides. Within the PNS, cholinergic activity and serotoninergic- and peptidergic-IRs occurred in the subtegumental network of nerve fibres and somatic musculature. Although all 3 neurochemical elements were present in the acetabula, they were found in different nerve fibres; only cholinergic and peptidergic cell bodies were found. The common genital opening, vagina and ootype regions of the reproductive system displayed a rich innervation of all 3 types of neuronal populations. Within the peptidergic system, immunostaining with antisera raised to the C-terminus of the neuropeptide Y superfamily of peptides and the invertebrate peptides, neuropeptide F (M. expansa) and FMRFamide was the most prevalent. Limited positive-IR for substance P and neurokinin A were also recorded in the CNS of the worm.

NEUROPEPTIDE-F (MONIEZIA-EXPANSA) - LOCALIZATION AND CHARACTERIZATION USING SPECIFIC ANTISERA

Immunocytochemical techniques used in conjunction with confocal scanning laser microscopy (CSLM) and electron microscopy have been used to demonstrate, for the first time, the distribution of the parasitic platyhelminth neuropeptide, neuropeptide F (NPF) in the cestode, Moniezia expansa. Antisera were raised to intact NPF(1-39) and to the C-terminal decapeptide of NPF(30-39). These antisera were characterized and validated for use in both immunocytochemistry and radioimmunoassay (RIA). NPF immunoreactivity (IR) was detected using both antisera throughout all of the major components of the central and peripheral nervous systems of the worm. The pattern of NPF-IR was found to mirror the IR obtained using a C-terminally directed pancreatic polypeptide (PP) antiserum and FMRFamide antisera; blocking studies using these antisera revealed that FMRFamide and PP antisera cross-react with NPF(M. expansa). RIA of acid-alcohol extracts of the worm measured 114 ng/g using the C-terminal NPF antiserum and 56 ng/g using the whole-molecule-directed antiserum. While the C-terminally-directed NPF antiserum cross-reacts with NPF-related peptides from other invertebrates, the whole-molecule-directed NPF antiserum is specific for NPF(M. expansa). The C-terminal NPF antiserum has potential for use in the identification and purification of NPF analogues from other platyhelminth parasites.

NEUROPEPTIDE F-IMMUNOREACTIVITY IN THE MONOGENEAN PARASITE DICLIDOPHORA-MERLANGI

The localisation and distribution of neuropeptide F (NPF)-immunoreactivity (IR) in the monogenean fish-gill parasite, Diclidophora merlangi, have been investigated by whole-mount immunocytochemistry interfaced with confocal scanning laser microscopy and, at the ultrastructural level, by indirect immunogold labeling. Using antisera directed to intact synthetic NPF (Moniezia expansa, residues 1-39) or to the C-terminal decapeptide (residues 30-39) of synthetic NPF (M. expansa), immunostaining was found throughout the central (CNS) and peripheral nervous systems (PNS), including the innervation of the reproductive system. Immunoreactivity was found to be more intense using the antiserum to the C-terminal decapeptide fragment of NPF. At the subcellular level, gold labeling of NPF-IR was found exclusively over the contents of dense-cored vesicles that occupied nerve axons of both the CNS and the PNS. The distribution pattern of immunostaining for NPF mirrored exactly that previously documented for the vertebrate pancreatic polypeptide (PP) family of peptides and for FMRFamide. This finding and the results of preabsorption experiments strongly suggest that NPF is the predominant native neuropeptide in D. merlangi and that it accounts for most of the immunostaining previously obtained with PP and FMRFamide antisera.