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  • 1
    ISSN: 1432-1955
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract The reproductive system of the monogenean gill parasite,Diclidophora merlangi, was examined for the presence of cholinergic, serotoninergic and peptidergic innervation using cytochemical and immunocytochemical techniques. Cholinesterase activity and 5-hydroxytryptamine immunoreactivity (5-HT-IR) were confined to neural elements of the male reproductive system, being evident in the innervation of the cirrus, whereas only 5-HT was present in nerves and somata of the elongate seminal vesicle. Peptidergic innervation was localised to both the male and female reproductive systems of the worm. Within the female reproductive apparatus pancreatic polypeptide, peptide tyrosine tyrosine, neuropeptide Y, substance P, neurokinin A, eledoisin, FMRFamide and gastrin/cholecystokinin immunoreactive fibres and somata were observed in the oviduct, vitelline reservoir and ovovitelline duct. Intense peptide immunoreactivity was identified in fibres in the wall of the ootype and in a surrounding population (〉100) of somata that were situated beyond Mehlis' gland cells and all of which were connected to the ootype wall by fine cytoplasmic connectives. The strategic location of this peptidergic cell population infers its involvement in the egg-forming sequence in this platyhelminth parasite.
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  • 2
    ISSN: 1432-1955
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract Adult and 3-week-old juvenileFasciola hepatica were examined for the presence of the cytoskeletal protein actin. Techniques of direct fluorescence using fluorescein isothiocyanate (FITC)-phalloidin and of indirect immunofluorescence using a monoclonal anti-actin antibody (MAA) demonstrated actin in the testes, sub-tegumental and gut musculature, tegumental cell bodies and tegumental spines. In contrast, polyclonal anti-actin antibody (PAA) revealed immunostaining only in the vitellaria. Effective removal of the tegument with 1% (w/v) sodium dodecyl sulphate (SDS) was confirmed by scanning electron microscopy (SEM), and this enabled immunoblotting of whole fluke and tegumental fractions with and without spines. Whole fluke fractions produced three bands corresponding to molecules exhibiting relative molecular weights of 43, 28 and 15 kDa, respectively, whereas the tegumental fraction with spines revealed a single band corresponding to 15 kDa in size. The fraction without spines displayed no bands. The present study localised actin in a number of different tissue types within the liver fluke. Using MAA, three forms of actin have been identified in the whole fluke and a single one in the tegumental spines.
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  • 3
    ISSN: 1432-1955
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract Antisera to a highly conserved region of chromogranin A (sequence KELTAE) and to a hexapeptide (sequence KGQELE) adjacent to the putative C-terminus of pancreastatin, a peptide whose sequence is found within the chromogranin A molecule, have been used to examine the localisation of immunoreactivity (IR) to these peptides inAscaris suum. IR to both peptides was found in the nerve rings and nerve cords. In addition, KGQELE-IR was also observed in the pharyngeal neurones and in a network of fibres on the surface of the female gonoduct. The staining was specific in that it could be abolished by preincubation of the antisera with the appropriate antigen. The two antisera appeared to be staining different subsets of neurones, suggesting that (at least) two peptides were being recognised. The widespread distribution of IR to both peptides throughout the nervous system of the parasite suggests that the peptides carrying the epitopes recognised by the antisera are of fundamental importance to the functioning of the parasite's nervous system.
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  • 4
    ISSN: 1432-1955
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract The distribution of actin filaments in the spermatogenic cells ofFasciola hepatica was determined using a fluorescent derivative of phalloidin. Actin was localised primarily in the region of separation of a secondary spermatogonium from a primary spermatogonium, in the inner faces at the centre of four-cell clusters of tertiary spermatogonia and in the cytophore region of spermatocyte and spermatid rosettes. The effect of the microfilament inhibitor cytochalasin B (100 μg/ml) on the ultrastructure of the spermatogenic cells was determined in vitro by transmission electron microscopy using tissue-slice material. Cytochalasin B treatment led to the formation of bi- and multinucleate cells, whose frequency increased with progressively longer incubation periods. Few typical rosettes of spermatocyte and spermatid cells were evident from 6 h onwards, being replaced by syncytial masses of cells. Spermatozoon formation became abnormal in the longer treatment periods, the spermatozoa containing variable numbers of axonemes and an altered distribution of cortical microtubules. Multiple axonemes were observed in the cytoplasm of spermatid cells. The results are discussed in relation to the established role of actin in the cytokinesis phase of cell division and to the effects of cytochalasin B on other tissues and organ systems within the fluke.
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  • 5
    ISSN: 1432-1955
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract The localisation and distribution of neuropeptides in the peripheral nervous system of the pig roundwormAscaris suum have been determined by an indirect immunofluoresence technique in conjunction with confocal microscopy. Of the 31 antisera tested, immunostaining was obtained only with antisera to peptide YY (PYY), pancreatic polypeptide (PP) and FMRFamide. Immunostaining for PYY and FMRFamide was evident in the amphidial and papillary ganglia associated with the anterior nerve ring and in the nerves from these ganglia that terminated in sensory receptors within the buccal lips of the parasite. The only peptide immunoreactivity (IR) observed in the reproductive system of either sex was that evident in the nerve supply to the distal region of the vagina in the female worm. It took the form of a well-developed plexus of parallel nerve fibres, cross-connectives and looped commissures. The nerve net diminished in the more proximal region of the vagina. PP-IR was less intense than that for PYY and FMRFamide and was more restricted in distribution, being confined to a small number of nerve fibres in the nerve supply to the vagina; it did not occur in the nerves supplying the anterior sensory receptors. The possible roles of neuropeptides in the sensory and reproductive biology of nematodes are discussed.
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  • 6
    ISSN: 1432-1955
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract Using an indirect immunofluorescence technique interfaced with confocal scanning laser microscopy, whole-mount preparations of three general of marine trematode larvae,Cryptocotyle lingua, Cercaria emasculans andHimasthla leptosoma, were screened for 5-hydroxytryptamine (5-HT) and selected neuropeptide immunoreactivities (IRs). IRs for pancreatic polypeptide (PP), peptide YY (PYY) and FMRFamide were found in the central nervous systems of the three species of cercariae, immunostaining the paired ganglia and central commissure and the longitudinal nerve cords, with slight differences in both distribution and intensity of IRs being observed for the different antisera used. PP, PYY and FMRFamide IRs were evident in both central and peripheral components of the nervous system in the rediae ofC. lingua. 5-HT IR was confined to the peripheral nervous systems of the cercariae ofC. emasculans and the rediae ofC. lingua, appearing in the form of a network of immunoreactive fibres and associated large cell bodies. A moderate substance P IR was observed in the nervous system of the cercariae ofC. lingua. The patterns of immunostaining described were compared with those obtained using antiserum directed to the C-terminal decapeptide amide of neuropeptide F (NPF), a native parasitic peptide from the cestodeMoniezia expansa. Results demonstrated that serotoninergic and peptidergic components were present in the nervous systems of all of the trematode larvae studied and that some, if not all, of the IR for PP, PYY and FMRFamide was due to the presence of a trematode NPF homologue.
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  • 7
    ISSN: 1432-1955
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract The localisation and distribution of the cholinergic and serotoninergic components of the nervous system in the plerocercoid, adult and free proglottis stages of the tetraphyllidean tapewormTrilocularia acanthiaevulgaris were determined by enzyme histochemical and immunocytochemical techniques. The central nerve ring (CNR) in the scolex contains two lateral ganglia and gives rise to five pairs of longitudinal nerve cords (LNC's; three lateral, two median). The nerve cords run posteriorly throughout the bodies of the plerocercoid and adult worms and the free proglottis. Nerves from the CNR and accessory lateral LNC's pass to the bothridia, where they give rise to extensive nerve plexuses. As the individual proglottides develop along the strobila, a small nerve ring forms at the anterior end of each proglottis; within the nerve ring, distinct bilateral ganglia develop prior to the release of the proglottis. All ten LNC's are present in the free proglottis. The genital atrium and cirrus sac are innervated by cholinergic and serotoninergic elements. The cholinergic nervous system predominates in the CNS within the scolex, whereas there is a larger population of 5-HT-immunoreactive nerve cells associated with the LNC's and segmental ganglia along the strobila and within the free proglottis.
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  • 8
    ISSN: 1432-1955
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract The localization and distribution of seven neuropeptides in the nervous system of the plerocercoid, adult and free proglottis stages of the tetraphyllidean tapewormTrilocularia acanthiaevulgaris have been determined by an indirect immunofluorescence technique. Six of the peptides are vertebrate-derived, namely, pancreatic polypeptide (PP), peptide tyrosine tyrosine (PYY), vasoactive intestinal polypeptide (VIP), peptide histidine isoleucine (PHI), substance P (SP) and somatostatin (SRIF); the seventh is the invertebrate neuropeptide, FMRFamide. This is the first demonstration of VIP and SP immunoreactivity in a castode parasite, and for SRIF this is its first description in any parasitic platyhelminth. Cell bodies and nerve fibres immunoreactive to PP, PYY, VIP, SP and FMRFamide are present throughout the CNS; the distributions of PHI and SRIF were more restricted. In the PNS, nerve fibres immunoreactive to PP occur in the bothridia, whilst in the free proglottis nerve fibres immunoreactive to PYY and VIP innervate the gonads; VIP-immunoreactive nerve elements also supply the reproductive ducts. Extra-neuronal sitings of peptide immunoreactivities were evident for PHI, in association with the excretory system, and for SRIF, in presumed tegumental cell bodies in the free proglottis. The results are discussed in relation to the possible roles of the peptides in the neurophysiology and developmental biology of the worm.
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  • 9
    ISSN: 1432-1955
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract The localistion and distribution of the cholinergic, serotoninergic and peptidergic components of the nervous system of the frog-lung flukeHaplometra cylindracea have been determined by the application of standard enzyme cytochemical and immunocytochemical techniques to cryostat sections and whole-mount preparations. Cholinesterase activity (ChE), as indicative of acetylcholine, has been demonstrated cytochemically in the CNS and PNS; however, the anterior ganglia were notably unreactive. The occurrence of serotonin was examined by an indirect immunofluorescence technique, and immunoreactivity (IR) was demonstrable in small, paired anterior ganglia and in fine nerve fibres associated with the somatic muscle, cirrus and gonopore. The peptidergic protion of the nervous system was investigated using antisera to 17 mammalian regulatory peptides and the invertebrate peptide FMRFamide, and was visualised by both indirect immunofluorescence and confocal scanning laser microscopy. Positive immunostaining occurred with antisera raised against pancreatic polypeptide (PP), peptide tyrosine tyrosine (PYY), substance P (SP), peptide histidine isoleucine (PHI) and FMRFamide. Immunoreactivity to PP, PYY and FMRFamide was widespread throughout the nervous system and was evident in large, paired anterior ganglia, the dorsal commissure, main nerve tracts and the extensive array of small fibres that constitute the PNS. In contrast, the distribution of nerves immunoreactive to SP and PHI was less apparent, with PHI-IR occurring exclusively within the fibrous neuropile of the ganglia and in fibres of the ventral nerve cord. Results are discussed with respect to the distribution of the various neurochemical elements and their roles as putative neurotransmitters and/or regulatory molecules.
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  • 10
    ISSN: 1432-1955
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract 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 inD. merlangi and that it accounts for most of the immunostaining previously obtained with PP and FMRFamide antisera.
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