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  • 1
    ISSN: 1432-1106
    Keywords: Transplantation ; Serotonin neurons ; Hypothalamus ; Electron microscopy ; Rat
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract We have previously reported that a cell suspension from the rostral part of the embryonic raphe grafted to the basal hypothalamus of 5,7-dihydroxytryptamine-denervated rats produced incomplete serotonin (5-HT) re-innervation of the suprachiasmatic nucleus (SCN) as opposed to hyper-innervation of the supraoptic nucleus (SON). We took advantage of this experimental model to investigate whether the graft-derived, 5-HT fibres retained normal ultrastructural features, and, particularly, a normal density of synaptic junctions, irrespective of the extent of target re-innervation. The intrinsic features of immunostained, graft-derived 5-HT axonal varicosities in both the SCN (ventral portion) and the SON were essentially similar to those exhibited by the respective endogenous innervation. Analysis of well-preserved varicosities in uninterrupted series of thin sections allowed us to evaluate directly the proportions of junctional to non-junctional 5-HT varicosities in both regions. Synaptic incidences were also remarkably conserved after grafting (45.5% in the SCN versus 38.5% in the SON; 48% and 38% in normal rats, respectively). Synapses were primarily reestablished on dendritic shafts, which also were identified as the major post-synaptic targets of the normal 5-HT innervations. We noted, however, a tendency toward increased numbers of symmetrical versus asymmetrical synapses in both the SCN and SON of grafted rats. Thus, irrespective of whether hypo-or hyper-innervation patterns developed post-grafting, the transplanted 5-HT neurons essentially retained normal ultrastructural features in their target territories, with a normal incidence of synaptic junctions. The data provide further support to the hypothesis that the innervation territory is the major determinant of the frequency with which ingrowing 5-HT fibres make synaptic junctions.
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  • 2
    ISSN: 1460-9568
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Medicine
    Notes: Survival and development of fetal serotonin (5-HT) neurons grafted to various brain areas in adult mammals have been suggested to be under host influences. The aim of this study was to determine whether the suprachiasmatic nucleus of the hypothalamus (SCN), a region receiving a 5-HT input which is one of the densest and the most heavily synaptic in the brain, can actually support the development of transplanted 5-HT neurons. The time course and extent of 5-HT reinnervation were therefore investigated with 5-HT immunocytochemistry in adult rats subjected to intraventricular injection of 5,7-dihydroxytryptamine and subsequent grafting of fetal cell suspension of mesencephalic raphe neurons. The ultrastructural features of the newly formed 5-HT terminal plexa were also examined. Serotonin reinnervation of the SCN remained partial up to 4 months post-transplantation, with no apparent predilection of the reinnervating fibres for any particular portion of the nucleus, thus differing from the normal 5-HT innervation of the SCN both quantitatively and qualitatively. This was in sharp contrast to the 5-HT hyperinnervation observed in neighbouring areas such as the supraoptic nucleus, a structure normally provided with only few 5-HT fibres, and the ventral wall of the third ventricle. The graft-derived 5-HT-axons, however, displayed ultrastructural features that did not appear different from those of their normal counterparts; in particular they re-established defined synaptic contacts with the host population. These results may indicate that the mature SCN specifically lacks a trophic factor necessary for the ingrowth of graft-derived 5-HT fibres, or that it represents an inhibitory environment for such an ingrowth. The limited ability of regrowing 5-HT axons to restore a normal density of 5-HT innervation could also be related to the fact that these neurons normally establish a relatively high number of synaptic contacts in the target region.
    Type of Medium: Electronic Resource
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  • 3
    ISSN: 1476-4687
    Source: Nature Archives 1869 - 2009
    Topics: Biology , Chemistry and Pharmacology , Medicine , Natural Sciences in General , Physics
    Notes: [Auszug] All experiments were carried out in adult Sprague-Dawley rats (200-400 g), pretreated with a monoamine oxidase inhibitor (-phenylisopropylhydrazine, 5 mg per kg intraperitoneally, 18 and 2 h earlier) and anaesthetised with sodium pentobarbital. 100-200 µCi of 3,4-dihydroxy ...
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  • 4
    ISSN: 1573-7381
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Summary Cellular relationships between serotonin (5-HT) axon terminals and neurons containing vasoactive intestinal peptide (VIP) were characterized by combined radioautography and immunocytochemistry in rat suprachiasmatic nucleus (SCN). Light microscopic immunoradioautographs showed significant overlap between (3H)5-HT uptake sites and VIP-immunoreactive elements in the ventral half of the SCN. Of the 255 (3H)5-HT-labelled axonal profiles detected in a systematic electron microscopic survey of single thin sections from this area, 75 (30%) were directly apposed to VIP-immunoreactive nerve cell bodies and/or dendrites. Radioautographically labelled 5-HT varicosities often showed well-differentiated, symmetrical or asymmetrical synaptic junctions, 60% of which were established on VIP-immunoreactive nerve cell bodies or dendrites. In a separate sampling of 198 (3H)5-HT-labelled terminals seen in apposition with VIP-immunoreactive elements, 50 showed a junctional complex at the site of contact. Postsynaptic immunoreactive elements were mostly dendrites but also included nerve cell bodies. Despite the methodological limitations inherent to the present double labelling approach, these data strongly support the view that VIP neurons are prime synaptic targets for 5-HT afferents in the SCN. VIP/5-HT interactions are thus likely to play an important functional role in this nucleus and may in particular subserve the 5-HT mediated regulation of certain circadian rhythms, including that of pituitary hormone secretion.
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  • 5
    ISSN: 1573-7381
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Summary Serotonin (5-HT) and γ-aminobutyric acid (GABA) nerve endings were identified in the rat suprachiasmatic nucleus (SCN) by combined [3H]5-HT uptake radioautography and glutamate decarboxylase (GAD) immunocytochemistry at the electron microscope level. In areas of good overlap between radioautographic and immunocytochemical labellings, there were no axonal varicosities exhibiting both labellings, indicating that 5-HT and GABA are not co-localized in the SCN. The systematic survey in these areas of all profiles that had accumulated [3H]5-HT and of all GAD-immunoreactive varicosities allowed the analysis of 247 of the former and 896, i.e. an almost four-fold greater number, of the latter. This seems concordant with the view that GABA endings would be the most numerous of all classes of nerve terminals so far identified in the SCN. More than 22% of the [3H]5-HT labelled profiles showed the membrane specialization typically associated with synap'tic junctions. Thereby, it was possible to evaluate that about 45% of the 5-HT terminals actually form a synapse in the SCN. Some 37% of the GAD-positive varicose profiles which could be formally interpreted also showed well differentiated synaptic contacts, suggesting that the GABAergic innervation of the SCN could be entirely junctional. Whereas 5-HT terminals usually innervated only one dendritic or somatic element, a convergence of several GABAergic terminals onto the same postsynaptic target also receiving a 5-HT input was frequently observed. Of all the [3H]5-HT labelled varicose profiles scanned, as much as 41% were directly apposed to at least one GAD-immunoreactive profile, indicating that these 5-HT/GABA axonal interfaces could well represent privileged sites of interactions between the two transmitters. Taken together, these data could be of potential value in determining the neurochemical mechanisms subserving cellular integration of rhythmic signals in the SCN.
    Type of Medium: Electronic Resource
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  • 6
    ISSN: 1573-7381
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Summary Light- and electron-microscopic immunocytochemistry was used to investigate grafts of foetal hypothalamic tissue implanted close to the site of the suprachiasmatic nucleus in adult rats with bilateral surgical ablation of this nucleus. The transplants contained vasoactive intestinal peptide and vasopressin cell clusters, which have previously been shown to characterize functional suprachiasmatic nucleus grafts. Vasoactive intestinal peptide and vasopressin neurons presented synaptic features that have not been described in the native suprachiasmatic nucleus. More specifically, their terminals within the graft were involved in ‘double’ synapses with separate unlabelled dendrites. Moreover, in dually stained sections, an unexpected synaptic investment of vasoactive intestinal peptide neurons by vasopressin endings was detected, which revealed reversed vasoactive intestinal peptide/vasopressin interactions compared to those described in the native nucleus. These observations could reflect some immature features of the grafted neurons. Ultrastructural relationships of monoaminergic fibres arising from host and/or intragraft neurons were also examined. Within the engrafted suprachiasmatic nucleus, tyrosine hydroxylase-labelled fibres, which probably belonged to cografted dopaminergic neurons, showed normal patterns of distribution and synaptic connections, with no preferential relationships with vasoactive intestinal peptide or vasopressin neurons. Serotoninergic axons arborized within transplants but, in agreement with previous data showing an inhibitory influence of the suprachiasmatic nucleus on ingrowing serotoninergic fibres, they had no predilection for the area corresponding to that nucleus. In spite of their relative scarcity, serotoninergic fibres within the engrafted suprachiasmatic nucleus showed an almost normal synaptic incidence, but synapses were not predominantly shared with the vasoactive intestinal peptide neurons, known to be their major targets in the native nucleus. This may contribute not only to the failure of functional grafts to synchronize with environmental conditions, but also to the inability of transplants to restore hormonal rhythms such as estrous cyclicity.
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  • 7
    ISSN: 1432-0878
    Keywords: Suprachiasmatic nucleus ; Catecholamines ; Dopamine ; Electron microscopy ; Vasoactive intestinal peptide ; Vasopressin ; Rat (Sprague Dawley)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract Catecholaminergic fibers in the suprachiasmatic nucleus of adult rats were investigated by use of light- and electron-microscopic immunocytochemistry. The suprachiasmatic nucleus receives a modest density of tyrosine hydroxylase-containing axons, homogeneously distributed in the nucleus and forming varicosities throughout its entire rostro-caudal extension. Immunolabeling with antibodies against dopamine showed that this catecholamine input comprises a dopaminergic component. Many tyrosine hydroxylase-positive cells were localized at the immediate periphery of the suprachiasmatic nucleus. With electron-microscopic examination, dendrites of these neurons were found within the limits of the nucleus as well as at a border zone between the suprachiasmatic nucleus proper and the optic tract where they received unlabeled synapses, providing a morphological support for a possible role of dopaminergic neurons in the integration and/or transfer of light-related signals. More than 91% of catecholaminergic axonal varicosities were found to establish morphologically defined synapses with dendrites. To investigate whether these synapses might be shared with neurons of one or both of the two main peptidergic populations of the nucleus, namely vasoactive intestinal peptide- and vasopressin-containing neurons, we carried out doublelabeling experments combining immunoperoxidase and immunogold-silver labeling. Results showed only a few cases of direct association of the catecholaminergic terminals with these peptidergic categories. In both types of dually stained sections, catecholaminergic synapses were preferentially made with unlabeled dendrites. The homogeneous distribution of tyrosine hydroxylase-immunoreactive fibers in the suprachiasmatic nucleus could therefore reflect a lack of significant catecholaminergic innervation of both vasoactive intestinal peptide- and vasopressin-synthesizing neurons.
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  • 8
    ISSN: 1432-0878
    Keywords: Key words: Suprachiasmatic nucleus ; Catecholamines ; Dopamine ; Electron microscopy ; Vasoactive intestinal peptide ; Vasopressin ; Rat (Sprague Dawley)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. Catecholaminergic fibers in the suprachiasmatic nucleus of adult rats were investigated by use of light- and electron-microscopic immunocytochemistry. The suprachiasmatic nucleus receives a modest density of tyrosine hydroxylase-containing axons, homogeneously distributed in the nucleus and forming varicosities throughout its entire rostro-caudal extension. Immunolabeling with antibodies against dopamine showed that this catecholamine input comprises a dopaminergic component. Many tyrosine hydroxylase-positive cells were localized at the immediate periphery of the suprachiasmatic nucleus. With electron-microscopic examination, dendrites of these neurons were found within the limits of the nucleus as well as at a border zone between the suprachiasmatic nucleus proper and the optic tract where they received unlabeled synapses, providing a morphological support for a possible role of dopaminergic neurons in the integration and/or transfer of light-related signals. More than 91% of catecholaminergic axonal varicosities were found to establish morphologically defined synapses with dendrites. To investigate whether these synapses might be shared with neurons of one or both of the two main peptidergic populations of the nucleus, namely vasoactive intestinal peptide- and vasopressin-containing neurons, we carried out double-labeling experiments combining immunoperoxidase and immunogold-silver labeling. Results showed only a few cases of direct association of the catecholaminergic termin als with these peptidergic categories. In both types of dually stained sections, catecholaminergic synapses were preferentially made with unlabeled dendrites. The homogeneous distribution of tyrosine hydroxylase-immunoreactive fibers in the suprachiasmatic nucleus could therefore reflect a lack of significant catecholaminergic innervation of both vasoactive intestinal peptide- and vasopressin-synthesizing neurons.
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  • 9
    ISSN: 0741-0581
    Keywords: Radioautography ; Immunocytochemistry ; Monoamines ; Peptides, GABA ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Natural Sciences in General
    Notes: Radioautography and immunocytochemistry may be combined at the light and electron microscopic levels for simultaneously localizing uptake sites for exogenous transmitter molecules [such as (3H)monoamines or (3H)amino acids] and endogenous transmitter-related antigens (classical transmitters and their synthesizing enzymes as well as neuropeptides) in the central nervous system. Silver grain accumulations indicative of transmitter uptake sites are readily distinguishable from immunocytochemical labels of the peroxidase-antiperoxidase (PAP), avitin-biotin, or colloidal gold methods. The combination of uptake radioautography and immunocytochemistry may be applied to the investigation of (1) the chemical identity of (3H) transmitter-accumulating elements, (2) the coexistence of different neurotransmitters within the same neurons, and (3) the cellular basis of interactions between certain neurotransmitters, in particular monoamines, GABA, and neuropeptides. This article describes and evaluates the method and reviews the available experimental data derived from its application.
    Additional Material: 7 Ill.
    Type of Medium: Electronic Resource
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