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  • 1
    ISSN: 1432-0878
    Keywords: Key words: Myenteric plexus ; Smooth muscle ; Organotypic culture ; Ultrastructure ; Intestine ; small ; Guinea-pig
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. External muscle and myenteric plexus from the small intestine of adult guinea-pigs were maintained in vitro for 3 or 6 days. Myenteric neurons and smooth muscle cells from such organotypic cultures were examined at the electron-microscopic level. An intact basal lamina was found around the myenteric ganglia and internodal strands. Neuronal membranes, nuclei and subcellular organelles appeared to be well preserved in cultured tissues and ribosomes were abundant. Dogiel type-II neurons were distinguishable by their elongated electron-dense mitochondria, numerous lysosomes and high densities of ribosomes. Vesiculated nerve profiles contained combinations of differently shaped vesicles. Synaptic membrane specializations were found between vesiculated nerve profiles and nerve processes and cell bodies. The majority of nerve fibres were well preserved in the myenteric ganglia, in internodal strands and in bundles running between circular muscle cells. No detectable changes were found in the ultrastructure of the somata and processes of glial cells. Longitudinal and circular muscle cells from cultured tissue had clearly defined membranes with some close associations with neighbouring muscle cells. Caveolae occurred in rows that ran parallel to the long axis of the muscle cells. These results indicate that the ultrastructural features of enteric neurons and smooth muscle of the guinea-pig small intestine are well preserved in organotypic culture.
    Type of Medium: Electronic Resource
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  • 2
    ISSN: 1432-0878
    Keywords: Key words: Organotypic culture ; Myenteric plexus ; Retrograde transport ; Intestine ; small ; Guinea-pig
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. The projections of myenteric neurons within the myenteric plexus of the guinea-pig small intestine were established using retrograde tracing in organotypic culture. Three days after applying the fluorescent dye DiI to a single internodal strand in the myenteric plexus, 500–1000 nerve cell bodies were labelled. Of these, 77% were located oral to the application site, 15% were located anally and 7% were located within 1 mm of this site. Three major morphological types of neurons could be distinguished. Dogiel type I neurons had lamellar dendrites and single axons, Dogiel type II neurons had large smooth cell bodies and several long processes, and filamentous neurons had smooth ovoid cell bodies, single axons and several filamentous dendrites. Dogiel type I, II and filamentous neurons accounted for 54.6%, 38% and 7.4% of all filled cells, respectively. Labelled nerve cell bodies were present up to 13 mm aboral to the DiI application site; all neurons more than 2 mm aboral had Dogiel type I features. On the oral side, Dogiel type I neurons were found up to 110 mm, Dogiel type II neurons up to 100 mm and filamentous neurons up to 80 mm. Neurons with 2 mm oral or aboral to the DiI application site were located up to 7 mm circumferentially and were mainly Dogiel type II cells. This work revealed remarkable polarity within the myenteric plexus, with a significant prevalence of myenteric neurons projecting anally for longer distances than those projecting orally. These long pathways are probably involved in the coordination of intestinal motility.
    Type of Medium: Electronic Resource
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  • 3
    ISSN: 1432-0878
    Keywords: Key words: 5-hydroxytryptamine ; Myenteric neurones ; Retrograde tracing ; Immunohistochemistry ; Chemical coding ; Morphology ; Guinea-pig
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract  Retrograde tracing, combined with immunohistochemistry, was used to study the projections of 5-hydroxytryptamine (5-HT)-accumulating neurones within the ileum of the guinea-pig, with confocal microscopy being used to characterise further their morphology. Two classes of neurones in the myenteric plexus, capable of taking up 5-HT or analogues, were distinguished. One class had Dogiel type I morphology with lamellar dendrites, was located on the edge or in the middle of ganglia and lacked immunoreactivity for somatostatin (SOM). The other class had smooth ovoid cell bodies with multiple filamentous dendrites and a single axon and represented a subset of the SOM-immunoreactive interneurones in the myenteric plexus. Varicosities immunoreactive for 5-HT alone, 5-HT/SOM or SOM alone were present in the myenteric ganglia. Both classes of 5-HT-accumulating neurones had long aboral projections within the myenteric plexus (up to 100 mm long) and to the submucous plexus and probably function as descending interneurones.
    Type of Medium: Electronic Resource
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  • 4
    ISSN: 1432-0878
    Keywords: Key words Neural tracing ; Biotinamide ; Cholinergic neurons ; Histochemistry ; Visceral sensory neurons ; Sympathetic neurons ; Guinea-pig
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract A novel technique for rapid anterograde labelling of cut axons in vitro was used to visualise the peripheral branches of mesenteric nerve trunks supplying the guinea-pig small intestine. Biotinamide, dissolved in an artificial intracellular solution, was applied to the cut ends of the mesenteric nerves and the tissue was maintained in organ culture overnight. Labelled nerve fibres were visualised by fluorescein isothiocyanate (FITC)-conjugated streptavidin. Intense staining of nerve fibres and terminal varicosities in the ganglia and internodal strands of the myenteric plexus was achieved up to 15 mm from the application site. Filled fibres formed baskets around some myenteric nerve cell bodies, suggesting target-specific neurotransmission. When combined with multiple-labelling immunohistochemistry for tyrosine hydroxylase (TH), calcitonin gene-related protein (CGRP) or choline acetyltransferase (ChAT), most anterogradely labelled nerve fibres, and many pericellular baskets, were found to be TH immunoreactive, indicating their postganglionic sympathetic origin. Double-labelling immunohistochemistry revealed that the postganglionic sympathetic pericellular baskets preferentially surrounded 5-hydroxytryptamine (5-HT)-handling myenteric neurons. Some biotinamide-filled fibres were CGRP immunoreactive, and are likely to originate from spinal sensory neurons. We describe for the first time many pericellular baskets labelled from the mesenteric nerves which were ChAT immunoreactive. Retrogradely filled intestinofugal nerve cell bodies were also observed, all of which had a single axon arising from a small nerve cell body with short filamentous or lamellar dendrites. Many of these cells were ChAT immunoreactive. This in vitro technique is effective in identifying the fine arrangement of nerve terminals arising from nerve trunks in the periphery.
    Type of Medium: Electronic Resource
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  • 5
    ISSN: 1432-0878
    Keywords: Calretinin ; Enteric nervous system ; Calcium binding protein ; Small intestine ; Cholinergic neurons ; Myenteric plexus ; Guinea-pig
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary Immunoreactivity for calretinin, a calcium-binding protein, was studied in neurones in the guinea-pig small intestine. 26±1% of myenteric neurones and 12±3% of submucous neurones were immunoreactive for calretinin. All calretinin-immunoreactive neurones were also immunoreactive for choline acetyltransferase and hence are likely to be cholinergic. In the myenteric plexus, two subtypes of Dogiel type-I calretinin-immunoreactive neurones could be distinguished from their projections and neurochemical coding. Some calretinin-immunoreactive myenteric neurones had short projections to the tertiary plexus, and hence are likely to be cholinergic motor neurones to the longitudinal muscle. Some of these cells were also immunoreactive for substance P. The remaining myenteric neurones, immunoreactive for calretinin, enkephalin, neurofilament protein triplet and substance P, are likely to be orad-projecting, cholinergic interneurones. Calretinin immunoreactivity was also found in cholinergic neurones in the submucosa, which project to the submucosal vasculature and mucosal glands, and which are likely to mediate vasodilation. Thus, calretinin immunoreactivity in the guinea-pig small intestine is confined to three functional classes of cholinergic neurones. It is possible, for the first time, to distinguish these classes of cells from other enteric neurones.
    Type of Medium: Electronic Resource
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  • 6
    ISSN: 1432-0878
    Keywords: Submucous plexus ; Mucosa ; Muscularis mucosae ; Intestine, small ; Retrograde transport ; Dil ; Guinea-pig
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary Double-labelling immunohistochemistry and retrograde transport of the carbocyanine dye, DiI, were used to establish the pathways of submucous neurons to the mucosa of the guinea-pig small intestine. Following the application of DiI to a villus, DiI-labelled nerve cell bodies were found in the submucous plexus up to 8.3 mm circumferentially and 3.8 mm longitudinally. The size of each of the four characterised classes of submucous neurons was determined and their distributions and projections mapped. Cells characterised by vasoactive intestinal polypeptide immunoreactivity accounted for 52% of DiI-labelled cells and had the longest projections. Cells characterised by neuropeptide Y (19%) or by calretinin immunoreactivity (13% of all DiI-labelled neurons) had relatively short projections and cells with substance P immunoreactivity (20%) had intermediate lengths of projection. When DiI was applied directly to the submucous plexus, filled neurons of all classes had significantly shorter projections, indicating that they must run for considerable distances in other pathways to the mucosa, probably via the non-ganglionated plexus. On average, each villus is innervated by at least 70 submucous neurons. From quantitative estimates there are 9 submucous neurons per villus. Thus, each submucous neuron is likely to supply about 8 villi. This demonstrates a high degree of convergence and divergence in the innervation of the mucosa.
    Type of Medium: Electronic Resource
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  • 7
    ISSN: 1432-0878
    Keywords: Myenteric plexus ; Enteric nervous system ; Intestine, small ; Ultrastructure ; Innervation, of intestinal muscle ; Guinea-pig
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract The tertiary component of the myenteric plexus consists of interlacing fine nerve fibre bundles that run between its principal ganglia and connecting nerve strands. It was revealed by zinc iodide-osmium impregnation and substance P immunohistochemistry at the light-microscope level. The plexus was situated against the inner face of the longitudinal muscle and was present along the length of the small intestine at a density that did not vary markedly from proximal to distal. Nerve bundles did not appear to be present in the longitudinal muscle as judged by light microscopy, although numberous fibre bundles were encountered within the circular muscle layer. At the ultrastructural level, nerve fibre bundles of the tertiary plexus were found in grooves formed by the innermost layer of longitudinal smooth muscle cells. In the distal parts of the small intestine, some of these nerve fibre bundles occasionally penetrated the longitudinal muscle coat. Vesiculated profiles in nerve fibre bundles of the tertiary plexus contained variable proportions of small clear and large granular vesicles; they often approached to within 50–200 nm of the longitudinal smooth muscle cells. Fibroblast-like cells lay between strands of the tertiary plexus and the circular muscle but were never intercalated between nerve fibre varicosities and the longitudinal muscle. These anatomical relationships are consistent with the tertiary plexus being the major site of neurotransmission to the longitudinal muscle of the guinea-pig small intestine.
    Type of Medium: Electronic Resource
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  • 8
    ISSN: 1432-0878
    Keywords: Myenteric plexus ; Smooth muscle ; Organotypic culture ; Ultrastructure ; Intestine, small ; Guinea-pig
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract External muscle and myenteric plexus from the small intestine of adult guinea-pigs were maintained in vitro for 3 or 6 days. Myenteric neurons and smooth muscle cells from such organotypic cultures were examined at the electron-microscopic level. An intact basal lamina was found around the myenteric ganglia and internodal strands. Neuronal membranes, nuclei and subcellular organelles appeared to be well preserved in cultured tissues and ribosomes were abundant. Dogiel type-II neurons were distinguishable by their elongated electron-dense mitochondria, numerous lysosomes and high densities of ribosomes. Vesiculated nerve profiles contained combinations of differently shaped vesicles. Synaptic membrane specializations were found between vesiculated nerve profiles and nerve processes and cell bodies. The majority of nerve fibres were well preserved in the myenteric ganglia, in internodal strands and in bundles running between circular muscle cells. No detectable changes were found in the ultrastructure of the somata and processes of glial cells. Longitudinal and circular muscle cells from cultured tissue had clearly defined membranes with some close associations with neighbouring muscle cells. Caveolae occurred in rows that ran parallel to the long axis of the muscle cells. These results indicate that the ultrastructural features of enteric neurons and smooth muscle of the guinea-pig small intestine are well preserved in organotypic culture.
    Type of Medium: Electronic Resource
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  • 9
    ISSN: 1432-0878
    Keywords: Calcium-binding protein ; Enteric nervous system ; Intestine ; Immunocytochemistry ; Guinea-pig ; Rat ; Man
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary Immunoreactivity for vitamin D-dependent calcium-binding protein (CaBP) has been localized in nerve cell bodies and nerve fibres in the gastrointestinal tracts of guinea-pig, rat and man. CaBP immunoreactivity was found in a high proportion of nerve cell bodies of the myenteric plexus, particularly in the small intestine. It was also found in submucous neurons of the small and large intestines. Immunoreactive nerve fibres were numerous in the myenteric ganglia, and were also common in the submucous ganglia and in the intestinal mucosa. Immunoreactive fibres were rare in the circular and longitudinal muscle coats. In the myenteric ganglia of the guinea-pig small intestine the immunoreactivity is restricted to one class of nerve cell bodies, type-II neurons of Dogiel, which display calcium action potentials in their cell bodies. These neurons were also immunoreactive with antibodies to spot 35 protein, a calcium-binding protein from the cerebellum. From the distribution of their terminals and the electrophysiological properties of these neurons it is suggested they might be sensory neurons, or perhaps interneurons. The discovery of CaBP in restricted sub-groups of enteric neurons may provide an important key for the analysis of their functions.
    Type of Medium: Electronic Resource
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  • 10
    ISSN: 1432-0878
    Keywords: Calcitonin gene-related peptide ; Cholecystokinin ; Choline acetyltransferase ; Neuropeptide Y ; Somatostatin ; Enteric nervous system ; Intestine, small ; Guinea-pig
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary Antisera to neuropeptide Y (NPY) gave an intense immunohistochemical reaction of certain nerve cells in the myenteric and submucous plexuses of the guinea-pig small intestine. Each nerve cell had up to 20 branching, tapering processes that were less than ∼50 μm long and a long process that could be followed for a considerable distance. This morphology corresponds to that of the type-III cells of Dogiel. The long process of each myenteric cell ran through the circular muscle to the submucosa, and in most cases the process could be traced to the mucosa. The submucous nerve cell bodies also had processes that extended to the mucosa. These cell bodies, in both plexuses, also stained with antisera raised against calcitonin generelated peptide (CGRP), cholecystokinin (CCK), choline acetyltransferase (ChAT) and somatostatin (SOM), but did not stain with antibodies against enkephalin, substance P or vasoactive intestinal peptide. Thus, it has been possible for the first time to trace the processes of chemically specified neurons through the layers of the intestinal wall and to show by a direct method that CGRP/CCK/ChAT/NPY/ SOM myenteric and submucous nerves cells provide terminals in the mucosa.
    Type of Medium: Electronic Resource
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