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Biomedical subjects

D W Halton

Publications and source records attributed to D W Halton.

At least 73 records · Page 4Linked to original sources
PubMed1995

Isolation and preliminary biological characterization of KPNFIRFamide, a novel FMRFamide-related peptide from the free-living nematode, Panagrellus redivivus.

A novel FMRFamide-related heptapeptide, Lys-Pro-Asn-Phe-Ile-Arg-Phe-NH2 (KPNFIRFamide), was isolated and characterized from acid ethanol extracts of the free-living nematode, Panagrellus redivivus. Whole-worm extracts contained > or = 9 pmol KPNFIRFamide/g wet weight. A synthetic replicate of this peptide induced a rapid relaxation of tone and inhibited spontaneous contractility in isolated innervated and denervated body-wall muscle strips of the parasitic nematode, Ascaris suum. KPNFIRFamide (0.1 nM) induced measurable relaxations in 50% of the muscle preparations examined. Concentrations > or = 0.3 nM induced relaxation in 100% of muscle preparations examined. The relaxation was short-lived at concentrations of peptide > or = 1 microM and displayed a profile typical of receptor desensitization. These data suggest the occurrence of a closely related peptide in A. suum and add further evidence to the concept of primary structural conservation of FaRPs within the nematodes.

Amino Acid Sequence↗
PubMed1995

Comparative analyses of the neuropeptide F (NPF)- and FMRFamide-related peptide (FaRP)-immunoreactivities in Fasciola hepatica and Schistosoma spp.

Immunochemical techniques were used to determine the distribution, chemical characteristics and relative abundance of immunoreactivity (IR) to two native platyhelminth neuropeptides, neuropeptide F (NPF) (Moniezia expansa) and the FMRFamide-related peptide (FaRP), GNFFRFamide, in the trematodes, Fasciola hepatica and Schistosoma mansoni; the larger S. margrebowiei was used in the chemical analysis. Extensive immunostaining for the two peptides was demonstrated throughout the nervous systems of both F. hepatica and S. mansoni, with strong IR also in the innervation of muscular structures, including those associated with the egg-forming apparatus. The patterns of immunostaining were similar to those previously described for the vertebrate neuropeptide Y superfamily of peptides and for FMRFamide. Ultrastructurally, gold labelling of NPF- and GNFFRFamide-IRs was localized exclusively to the contents of secretory vesicles in the axons and somatic cytoplasm of neurones. Double-labelling experiments showed an apparent homogeneity of antigenic sites, in all probability due to the demonstrated cross-reactivity of the FaRP antiserum with NPF. Radioimmunoassay of acid-ethanol extracts of the worms detected 8.3 pmol/g and 4.7 pmol/g equivalents of NPF- and FMRFamide-IRs, respectively, for F. hepatica, and corresponding values of 4.9 pmol/g and 4.3 pmol/g equivalents for S. margrebowiei. Gel-permeation chromatography resolved IR to both peptides in discrete peaks and these eluted in similar positions to synthetic NPF (M. expansa) and GNFFRFamide, respectively.

Amino Acid Sequence↗
PubMed1995

GABA in the nervous system of parasitic flatworms.

In an immunocytochemical study, using an antiserum and a monoclonal antibody specific for the amino acid, gamma-aminobutyric acid (GABA), GABA-like immunoreactivity (GLIR) has been demonstrated for the first time in parasitic flatworms. In Moniezia expansa (Cestoda), GLIR was seen in nerve nets which were closely associated with the body wall musculature and in the longitudinal nerve cords. In the liver fluke Fasciola hepatica (Trematoda), the GLIR occurred in the longitudinal nerve cords and lateral nerves in the posterior half of the worm. GLIR was also detected in subtegumental fibres in F. hepatica. The presence of GABA was verified, using high-pressure liquid chromatography coupled with fluorescence detection. The concentration of GABA (mean +/- S.D.) in M. expansa anterior region was 124.8 +/- 15.3 picomole/mg wet weight, while in F. hepatica it was 16.8 +/- 4.9 picomole/mg. Since several insecticides and antinematodal drugs are thought to interfere with GABA-receptors, the findings indicate that GABAergic neurotransmission may be a potential target for chemotherapy in flatworms too.

Animals↗
PubMed1995

Localization of Diploptera punctata allatostatin-like immunoreactivity in helminths: an immunocytochemical study.

The nervous systems of helminths are predominantly peptidergic in nature, although it is likely that the full range of regulatory peptides used by these organisms has yet to be elucidated. Attempts to identify novel helminth neuropeptides are being made using immunocytochemistry with antisera raised against peptides isolated originally from insects. One of these antisera was raised against allatostatin III, a peptide isolated originally from the cockroach, Diploptera punctata, and a member of a family of related peptides found in insects. Allatostatin immunoreactivity was found throughout the nervous systems of Mesocestoides corti tetrathyridia, and adult Moniezia expansa, Diclidophora merlangi, Fasciola hepatica, Schistosoma mansoni, Ascaris suum and Panagrellus redivivus. Immunostaining was observed in the nerve cords and anterior ganglia of all the helminths. It was also apparent in the subtegumental nerves and around the reproductive apparatus of the flatworms, in neurones in the pharynx of D. merlangi, F. hepatica, A. suum and P. redivivus, and in fibres innervating the anterior sense organs in the nematodes. Immunostaining in all species was both reproducible and specific in that it could be abolished by pre-absorption of the antiserum with allatostatins I-IV. These results suggest that molecules related to the D. punctata allatostatins are important components in the nervous systems of a number of helminth parasites, and a free-living nematode. Their distribution within the nervous system suggests they function as neurotransmitters/neuromodulators with roles in locomotion, feeding, reproduction and sensory perception.

Animals↗
PubMed1994

Peptides related to the Diploptera punctata allatostatins in nonarthropod invertebrates: an immunocytochemical survey.

The allatostatins are a family of peptides isolated originally from the cockroach, Diploptera punctata. Related peptides have been identified in Periplaneta americana and the blowfly, Calliphora vomitoria. These peptides have been shown to be potent inhibitors of juvenile hormone synthesis in these species. A peptide inhibitor of juvenile hormone biosynthesis has also been isolated from the moth, Manduca sexta; however, this peptide has no structural homology with the D. punctata-type allatostatins. Investigations of the phylogeny of the D. punctata allatostatin peptide family have been started by examining a number of nonarthropod invertebrates for the presence of allatostatin-like molecules using immunocytochemistry with antisera directed against the conserved C-terminal region of this family. Allatostatin-like immunoreactivity (ALIR) was demonstrated in the nervous systems of Hydra oligactis (Hydrozoa), Moniezia expansa (Cestoda), Schistosoma mansoni (Trematoda), Artioposthia triangulata (Turbellaria), Ascaris suum (Nematoda), Lumbricus terrestris (Oligochaeta), Limax pseudoflavus (Gastropoda), and Eledone cirrhosa (Cephalopoda). ALIR could not be demonstrated in Ciona intestinalis (Ascidiacea). These results suggest that molecules related to the allatostatins may play an important role in nervous system function in many invertebrates as well as in insects and that they also have an ancient evolutionary lineage.

Amino Acid Sequence↗
PubMed1994

The corrected primary structure of chicken (avian) pancreatic polypeptide.

Chicken (avian) pancreatic polypeptide was the first member of the pancreatic polypeptide (PP)/neuropeptide Y (NPY) superfamily to be discovered and structurally-characterised. In this 36 amino acid residue, C-terminally amidated peptide, residues 22 and 23 were identified as Asp and Asn, respectively. However, sequencing of chicken PP using modem automated gas-phase sequencing technology has revealed that the original primary structure is incorrect in that residue 22 is Asn and that residue 23 is Asp. After digestion of chicken PP with endoproteinase Asp-N, fragments of chicken PP corresponding in molecular mass to residues 16-22 and 23-36, were unequivocally identified. The corrected primary structure of chicken PP is therefore: Gly-Pro-Ser-Gln-Pro-Thr-Tyr-Pro-Gly-Asp-Asp- Ala-Pro-Val-Glu-Asp-Leu-Ile-Arg- Phe-Tyr-Asn-Asp-Leu-Gln-Gln-Tyr-Leu-Asn-Val-Val-Thr-Arg-His-Arg-Tyr-NH2.

Amino Acid Sequence↗
PubMed1994

KSAYMRFamide: a novel FMRFamide-related heptapeptide from the free-living nematode, Panagrellus redivivus, which is myoactive in the parasitic nematode, Ascaris suum.

In nematodes, FMRFamide-related peptides (FaRPs) have been structurally characterised from the parasite, Ascaris suum, and from two free-living species, Panagrellus redivivus and Caenorhabditis elegans. While both FaRPs isolated from P. redivivus (PF1 and PF2) have been identified in C. elegans, the two heptapeptides isolated from A. suum (AF1 and AF2) have until recently been considered unique to this parasitic species. We have recently isolated AF2 from P. redivivus and, during this study, an additional novel heptapeptide amide, Lys-Ser-Ala-Tyr-Met-Arg-Phe amide (KSAYMRFamide), was structurally characterised. A synthetic replicate of this peptide induced a rapid concentration-dependent muscle tension increase in an isolated. A. suum somatic muscle preparation, with a threshold of approximately 0.1 microM. These data suggest that the complement of FaRPs in parasitic and free-living nematodes may not be as radically different as preliminary studies would suggest, and that the absence of AF1, AF2 and KSAYMRFamide on the C.elegans FMRFamide-related peptide gene (flp-1) may imply the presence of at least two different FaRP genes in nematodes.

Amino Acid Sequence↗
PubMed1994

RYIRFamide: a turbellarian FMRFamide-related peptide (FaRP).

FMRFamide was isolated originally from neural-tissue extracts of a bivalve mollusc, since when either authentic FMRFamide or a series of structurally-related peptides have been isolated from representative arthropods, annelids and many additional molluscs. However, to date no information exists as to the definitive presence and primary structure of a FaRP in a free-living flatworm. Here, we report the isolation and primary structure of a FaRP from the free-living turbellarian, Artioposthia triangulata, a species from which NPF has been previously structurally-characterised. Unlike molluscs and insects, in which several FaRPs are expressed, only a single member of this family was detected in this turbellarian. The primary structure of this turbellarian FaRP was established as Arg-Tyr-Ile-Arg-Phe-NH2 (RYIRFamide) and the molecular mass as 752.7 Da. These data have established unequivocally that FaRPs occur in the nervous systems of the most phylogenetically-ancient invertebrates which display bilaterally-symmetrical neuronal plans and that authentic FMRFamide is probably not the original member of the family in molecular evolutionary terms.

Amino Acid Sequence↗
PubMed1994

Grillotia erinaceus (Cestoda, Trypanorhyncha): localization of neuroactive substances in the plerocercoid, using confocal and electron-microscopic immunocytochemistry.

Indirect immunocytochemistry, in conjunction with confocal scanning laser microscopy and electron-microscopic immunogold labeling, has been used to localize neuropeptide and 5-hydroxytryptamine (5-HT) immunoreactivities (IRs) in the plerocercoid (scolex and surrounding blastocyst) of the trypanorhynch tapeworm, Grillotia erinaceus. Antisera directed to two native cestode neuropeptides, neuropeptide F and the FMRFamide-related peptide, GNFFRFamide, were used to demonstrate the presence of a well-developed and extensive peptide-immunoreactive nervous system of central and peripheral elements in the juvenile scolex. Neuronal connectivity exists between the scolex and the surrounding blastocyst, in which there is a rich innervation of varicose fibers displaying peptide IR. Ultrastructurally, gold labeling of the peptide IR was found exclusively over the contents of dense secretory vesicles in the axons and somatic cytoplasm of neurons. Double-labeling experiments demonstrated an apparent colocalization of peptide IR, although the results of antigen preadsorption procedures indicated substantial cross-reactivity of the two antisera. A separate and well-differentiated 5-HT-immunoreactive nervous system, with a similar anatomical arrangement as the peptide-immunoreactive nervous system, is present in both the scolex and blastocyst.

Animals↗
PubMed1994

Isolation and structural characterisation of herring gull (Larus argentatus) pancreatic polypeptide.

Pancreatic polypeptide (PP) has been isolated from pancreatic extracts of the herring gull (Larus argentatus) using a radioimmunoassay employing an antiserum (PP 221), generated to the conserved C-terminal hexapeptide amide of mammalian PP. Gel permeation and reverse-phase HPLC fractionation of crude pancreatic extracts resolved a single molecular form of gull PP in each case. Purification of gull PP to homogeneity indicated that the mammalian PP antiserum employed was only 4% cross-reactive with this peptide. When radioimmunoassay data were corrected for this, the herring gull pancreas was found to contain 5 nmol PP/g wet wt. The molecular mass of gull PP was determined as 4237 +/- 2 Da by 252Cf plasma desorption mass spectroscopy (PDMS). Gas-phase sequencing established unequivocally the entire primary structure of a 36-amino-acid residue peptide as Gly-Pro-Val-Gln-Pro-Thr-Tyr-Pro-Gly-Asp-Asp-Ala-Pro-Val-Glu-Asp-Leu-Val- Arg-Phe - Try-Asn-Asp-Leu-Gln-Gln-Tyr-Leu-Asn-Val-Val-Thr-Arg-His-Arg-Tyr. The computed molecular mass of this peptide (4235.6 Da) was in close agreement with that derived by PDMS. The primary structure of herring gull PP differs from that of the chicken in four residues at positions 3 (Val/Ser), 18 (Val/IIe), 22 (Asn/Asp), and 23 (Asp/Asn). Avian PP thus appears to be a highly conserved regulatory peptide.

Amino Acid Sequence↗
PubMed1994

Serotonin and neuropeptide immunoreactivities in the intramolluscan stages of three marine trematode parasites.

Using an indirect immunofluorescence technique interfaced with confocal scanning laser microscopy, whole-mount preparations of three genera of marine trematode larvae, Cryptocotyle lingua, Cercaria emasculans and Himasthla 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 of C. lingua. 5-HT IR was confined to the peripheral nervous systems of the cercariae of C. emasculans and the rediae of C. 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 of C. 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 cestode Moniezia 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.

Amino Acid Sequence↗
PubMed1994

Ultrastructural localisation of FMRFamide- and pancreatic polypeptide-immunoreactivities within the central nervous system of the liver fluke, Fasciola hepatica (Trematoda, Digenea).

A post-embedding immunogold technique was used to examine the subcellular distribution of immunoreactivities to the invertebrate peptide, FMR-Famide, and to vertebrate pancreatic polypeptide (PP) within the central nervous system of the trematode, Fasciola hepatica. Gold labeling of peptide was localised exclusively over both dense-cored and ellipsoidal electron-dense vesicles (with a homogeneous matrix) present within nerve cell bodies, small and 'giant' nerve processes of the neuropile in the cerebral ganglia and transverse commissure, as well as in the main longitudinal nerve cords. Double labeling demonstrated an apparent co-localisation of FMRFamide and PP immunoreactivities in the same dense-cored vesicles, although populations of ellipsoidal electron-dense vesicles that labeled solely for FMRFamide were also evident. Antigen pre-absorption studies indicated little, if any, cross-reactivity of the two antisera.

Animals↗
PubMed1994

5-hydroxytryptamine-immunoreactivity in the monogenean parasite, Entobdella soleae.

5-HT-immunoreactivity in Entobdella soleae was found to be extensive throughout both the central and peripheral nervous systems, with the strongest staining occurring in the innervation of the forebody, most notably in the paired cerebral ganglia, pharynx and adhesive pads. In the reproductive system, staining was evident throughout the numerous cell bodies and fibres innervating the musculature of the egg-assembly apparatus. The haptor contained an extensive array of serotoninergic fibres derived from the main longitudinal cords; this array was associated with the haptoral muscles and sclerites, and possibly with the ventral sensory papillae.

Animals↗
PubMed1994

Immunocytochemical localization of neuropeptide F-immunoreactivity in the circumoesophageal ganglia of the gastropod mollusc, Helix aspersa using electron microscopy.

Neuropeptide F (NPF)-immunoreactivity has been localized by electron microscopy in the central nervous system of the gastropod mollusc, Helix aspersa using a region-specific antiserum in conjunction with post-embedding immunogold labelling. The specific antiserum employed, designated NPF3, was raised to a synthetic N-terminal fragment of H. aspersa NPF. Gold-labelling revealed that NPF-immunoreactivity was found almost exclusively over the contents of dense-cored secretory vesicles within nerve axons in both cerebral and subesophageal ganglia. Double-labelling of NPF and FMRFamide demonstrated no apparent co-localization in the paired cerebral ganglia. Instead, immunoreactivities of the two neuropeptides were found to have distinct locations within the ganglia. These findings suggest that NPF may be an important neuroregulator in H. aspersa and that in the cerebral ganglia it may be anatomically-distinct from FMRFamide and related peptides.

Amino Acid Sequence↗
PubMed1994

Ultrastructural localization of pancreatic polypeptide- and FMRFamide immunoreactivities within the central nervous system of the nematode, Ascaris suum (Nematoda: Ascaroidea).

A post-embedding immunogold technique has been used to examine the subcellular distribution of immunoreactivities to vertebrate pancreatic polypeptide (PP) and to the invertebrate peptide, FMRFamide within the central nervous system (CNS) of the nematode, Ascaris suum. Gold labelling of peptide was localized exclusively over dense-cored vesicles within nerve cell bodies, nerve axons and nerve terminals of the main ganglia and nerve cords in the CNS. Double-labelling of peptides demonstrated an apparent co-localization of PP and FMRFamide immunoreactivities in the same dense-cored vesicles, although populations of dense-cored vesicles that labelled solely for FMRFamide were also evident. Antigen preabsorption studies indicated little or no cross-reactivity between the two antisera.

Animals↗
PubMed1994

Identification and characterization of the cysteine and serine proteinases of the trematode, Haplometra cylindracea and determination of their haemoglobinase activity.

The excreted/secreted proteinases of Haplometra cylindracea maintained in vitro, were found to hydrolyse the fluorogenic substrates, Z-ArgArg-NHMec and Z-PheArg-NHMec. This activity was shown to be typically that of cysteine proteinases, as turn-over of both substrates could be blocked by pre-incubation with peptidyl diazomethyl ketones. The biotinylated affinity reagent, biotin-Phe Ala-DMK, used in combination with Z-PheTyr(OBut)-DMK, was employed for the labelling and characterization of these cysteine proteinase activities. Three cathepsin B-like species were detected, with molecular weights of 48, 22-23 and 14 kDa, together with a cathepsin L-like enzyme, with a molecular weight of 55 kDa. The proteinases were also found to have hydrolytic activity towards the substrate, Z-GlyGlyArg-NHMec, which could be blocked by pre-incubation with either of the serine proteinase-selective reagents, Z-Argp(OPh)2 or biotin-Lysp(OPh)2, showing the activity to be trypsin-like. Using the biotinylated affinity label to characterize the trypsin-like enzymes revealed two molecular species with molecular weights of 20 and 24 kDa.

Affinity Labels↗
PubMed1994

The FMRFamide-like neuropeptide AF2 (Ascaris suum) is present in the free-living nematode, Panagrellus redivivus (Nematoda, Rhabditida).

Available primary structural information suggests that the FMRFamide-related peptides (FaRPs) from parasitic and free-living nematodes are different, and that free-living forms may not represent appropriate models for the study of the neurochemistry of parasitic forms in the laboratory. However, here we report the isolation and unequivocal identification of AF2 (originally isolated from the parasite, Ascaris suum) from acidified alcoholic extracts of the free-living species, Panagrellus redivivus. While reverse-phase HPLC analysis of extracts revealed FMRFamide-immunoreactivity to be highly heterogeneous, AF2 was the predominant FMRFamide-immunoreactive peptide present (at least 26 pmol/g wet weight of worms). This peptide was also the major immunoreactant identified by an antiserum raised to the conserved C-terminal hexapeptide amide of mammalian pancreatic polypeptide (PP), which has been used previously to isolate neuropeptide F (NPF). These observations were confirmed by radioimmunoassay and chromatographic fractionation of an acidified alcoholic extract of A. suum heads. The FMRFamide-related peptides present in a nematode extract may be highly dependent on the extraction medium employed, and these data would suggest that this complement of neuropeptides may not be as different between parasitic and free-living nematodes as initial studies have suggested. Finally, all of the evidence suggests that NPF is not present in nematodes and that the PP-immunoreactant previously demonstrated immunochemically is probably AF2.

Amino Acid Sequence↗