A detailed analysis of the difference in the localization and the immunoreactivity for various surface markers among folliculo-stellate cells, macrophages, and dendritic cells was performed using immunohistochemistry and electron microscopy of the rat pituitary gland. The folliculo-stellate cells were selectively labeled by an antiserum against SIOO protein. The majority of dendritic cells were immunoreactive for the MHC class II (Ia) antigen (OX6) and/or the dendritic cell antibodies (OX62). The main population of macrophages was positive for the macrophage antibodies (ED1, ED2, and/or OX42). The cellular density of adenohypophyseal macrophages was significantly lower than that of folliculo-stellate cells and of dendritic cells. All the neurohypophyseal microglial cells were labeled with OX42, while the mAb OX6 labeled a small population of cells different from the cells identified by OX42 in the neurohypophysis. Double-immunoperoxidase staining for ED1 and OX6 revealed that positively stained cells could be classified into ED1⁺OX6^-, ED1⁺OX6⁺, and ED1^-OX6⁺ cells. Double staining with OX62 and OX6 mAbs showed that about 60% of the OX6⁺ cells were also immunolabeled with OX62 in the anterior lobe: OX62 detects a subpopulation of dendritic cells but does not recognize macrophage populations. Furthermore, double staining for S1OO and OX6 resulted in no S1O0⁺ OX6⁺ cells. At the electron-microscopic level, reaction products for OX6 were confirmed in the cell membrane and labeled cells were distinguished from macrophages and folliculo-stellate cells by distinctive short, broad cytoplasmic processes and the rare presence of cytoplasmic organelles. Such cytological characteristics of the OX6-positive cells in the pituitary gland are similar to dendritic cells. Our results suggest that resident dendritic cells and folliculo-stellate cells are two different main components of interstitial cells in the pituitary gland.

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We investigated the binding of four types of lectin to follicle-associated epithelium overlying the nasal-associated lymphoid tissue (NALT) of rats in order to identify M-cell specific surface markers and to determine the influence of lectin administration to NALT on the uptake of a particulate antigen. The NALT tissues were incubated with a panel of four types of lectin conjugated to horseradish peroxidase (HRP). Ulex europaeus-1 (UEA-1) and Dolichos biflorus (DBA) lectin stained the surface of M-cells and goblet cells. Uniform staining by Triticum vulgaris (WGA) was detected in the M-cells, ciliated cells and goblet cells. In contrast, staining of Griffonia simplicifolia I isolectin-B₄ (GS I-B₄) was almost exclusively M-cell specific. The administration of M-cell specific lectin (GS I-B₄) to NALT suppressed the uptake of baker’s yeast particles administered later, whereas the non-specific one (UEA-1) had no influence on the uptake. These results indicate that GS I-B₄ is a useful marker for the identification of rat NALT M-cells and that such a specific expression of surface glycoconjugates by M-cells may permit the targeting of vaccines and drugs to the antigen sampling sites of the nose. It also appears possible to block the uptake of pathogens by an administration of M-cell specific lectin to NALT.

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The present study demonstrated the precise spatial relationship between meshes in the perineuronal proteoglycan network and the terminal boutons of synaptically associated axons. Sections from the rat cerebellum were stained with cationic colloidal iron (pH 1.0-1.5), and successively immunostained with anti-calbindin-D-28K monoclonal antibody. Cationic iron stained sulfated proteoglyeans around the nerve cell of the medial cerebellar nucleus, whereas the anti-calbindin antibody labeled the Purkinje cells including their axons terminating on large neurons in the cerebellar nucleus. It was found that each synaptic bouton fits into a mesh of the perineuronal network. The individual meshes appeared to be divided by partitions faintly stained with the colloidal iron. Electron microscopy of cationic colloidal iron-stained ultrathin sections revealed that the synaptic boutons were separated from each other by the proteoglycan matrix and that each of them was further divided into two or more contact areas of presynaptic membrane by the same matrix. This suggests that individual synapses are protected against the effects of adjacent synaptic transmission, and that each of them may be subdivided by this manner of partitioning. like pads of a cat’s paw.

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The first data based on immunolabeling techniques of goat fetus adenohypophysis show that the structure and ultrastructure of growth hormone (GH)-, prolactin (PRL)-, and GH- plus PRL-secreting cells (SMT cells) in fetuses aged 100 days differ from those in the adult. Both cell number and cell size are smaller in the fetus, and the percentage of dark cells decreases with development. The data do not support the hypothesis that SMT cells represent the common origin of GH- and PRL-cells.

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We investigated the muscular structure and innervation of the gastroduodenal junction in the guinea pig. In the gastroduodenal junction, the innermost layer of the circular muscle contained numerous nerve fibers and terminals. Since this nerve network continued onto the deep muscular plexus (DMP) of the duodenum, we surmised that the numerous nerve fibers in the gastroduodenal junction were specialized DMP in the most proximal part of the duodenum. The innermost layer containing many nerve fibers was about 1000 μm in length and 100 μm in thickness in the proximal duodenum. This layer contained numerous connective tissue fibers composed of collagen and elastic fibers. Five to 30 smooth muscle cells lay in contact with each other and were surrounded by fine connective tissue. The nerve fibers in the proximal duodenum contained nerve terminals immunoreactive for choline acetyltransferase, dynorphin, enkephalin, galanin, gastrin-releasing peptide, nitric oxide synthase, substance P, and vasoactive intestinal polypeptide. Adrenergic fibers which contained tyrosine hydroxylase immunoreactivity were rare in the proximal duodenum. In the innermost layer of the proximal duodenum, there were numerous c-Kit immunopositive cells that were in contact with nerve terminals. This study allowed us to clarify the specific architecture of the most proximal portion of the duodenum. The functional sigrrificance of the proximal duodenum in relation to the electrical connection and neural cooperation of the musculature between the antrum and the duodenum is also discussed.

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In order to understand the fine structure and distribution of the interstitial glandular cells (IGCs) and associated elements in the human fetal ovary, we studied human fetal ovaries at 16 weeks post fertilization (p. f.) by transmission electron microscopy.
  Semithin sections revealed voluminous typical IGCs usually grouped in clusters, located in the interstitium among the ovigerous cords. Isolated primordial follicles were seen in the cords located close to the interstitium in which IGCs were present. Besides the main ultrastructural characteristics of steroid secreting cells, the IGCs showed lipofuscin granules and stacks of annulate lamellae in their cytoplasm. Fibrocytes, macrophages and mast cells were detected close to the IGCs. In particular, the fibrocytes were located around the IGCs, with which they occasionally formed focal cell contacts. Fibrocytes issued numerous long projections, which, together with collagen fibers, surrounded the clusters of IGCs and small vessels (mainly capillaries), often extending into the intercellular spaces among IGCs.
  These data indicated that, already at the initiation of folliculogenesis, the IGCs are present numerously in a close association with the ovigerous cords. The morphological aspects of IGCs were comparable to that of fetal testis interstitial (Leydig) cells and hilar cells in adult ovary, and suggest that fetal IGCs may be source of adult ovary hilar cells. In addition, we have here demonstrated for the first time that IGCs are associated with stromal cells whose distribution seems to support IGCs microtopography. Fetal ovarian fibrocytes revealed a structural arrangement similar to that of the “compartmentalizing cells” previously described in the adult testis. Macrophages and mast cells presumably have a role as local modulators of steroid synthesis. Mast cells may also affect fibrocyte organization and vascular permeability. We thus suggest that IGCs and associated cells may form a glandular unit in the human fetal ovary similar to that in the adult testis, and this structure is likely involved in early steroid secretion during gonadal differentiation.

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The presence and distribution of nicotinamide dinucleotide phosphate diaphorase (NADPH-d)-containing neurons have been studied by means of NADPH-d histochemistry in different regions of the adult cow gut, from the esophagus to the rectum.
  NADPH-d and nitric oxide synthase (NOS) were constantly recognized to be colocalized in the same neuron. The colocalization of vasoactive iutestinal polypeptide (VIP) and galanin in such nitrergic neurons was also studied by means of combined histochemical and immunofluorescence techniques. NADPH-d-positive neurons were present along the myenteric plexus of the entire gut, and in the submucous plexus from the abomasum to the rectum. Notably, they formed two types of nerve networks in the submucous connective tissue of the jejunum-ileum. NADPH-d-positive innervation of the muscle layers occurred throughout the tract, and sometimes a clear correspondence was noted between the number of reactive fibres and the thickness of the muscle. Nitrergic fibres also occurred in the mucosa and often were in relation to glands and blood vessels. The nitrergic neurons varied in size, shape, and intensity of staining, and often their terminals were seen to surround unstained perikarya. Various types of neurons were recognized on the basis of their chemical content; one of them contained galanin, VIP and NOS simultaneously.
  The present results suggest that the nitrergic neurons of the bovine gastrointestinal tract play roles presumably for controlling the motility of the gut and the conduction of interneuronal imprlses.

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The effect of a loss of occlusal stimuli upon the distribution and structure of the periodontal mechanoreceptors of the rat mandibular molar was examined after extracting opposing molars.
  The hypofunctional periodontal ligament narrowed significantly two weeks after tooth extraction, associated with an altered morphology of the Ruffini endings that showed typical dendritic profiles in normal controls. At four weeks and later periods after extraction, the Ruffini endings-including those without light microscopic changes - demonstrated unusual ultrastructural features such as the eccentric localization of mitochondria along the axonal membrane and loss of other cell organelles, unusual elongation of axonal microprojections, or a deep invagination of the Schwann sheath into the axoplasm. Immunoreactivity for the growth-associated protein-43 (GAP-43) in the Ruffini endings was restricted to the Schwann element in both the normal and hypofunctional periodontal ligament, but the reaction was weaker and even negligible in some cases in the latter ligament.
  The present results suggest that occlusal stimuli are essential for maintaining the structural integrity of the periodontal ligament, including that of periodontal mechanoreceptors. A decreased immunoreactivity for GAP-43 in the Schwann sheaths supports the notion of a possible functional alteration in the Ruffini endings that showed no structural abnormality.

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The expression of heat shock protein (Hsp) 25 during odontogenesis in the dental pulp and enamel organ of rat incisors was investigated by immunocytochemistry and confocal microscopy. In the process of dentin formation, immature odontoblasts first exhibited Hsp 25-immunoreactivity, and increased in immunointensity with the advance of their differentiation. In the dental pulp, in contrast, intense immunoreaction in the mesenchymal cells became weak or negative in parallel with the progress of cell differentiation. The immunoreaction for Hsp 25 in the enamel organ revealed a characteristic stage-related alteration during amelogenesis. In secretory ameloblasts, the immunoreaction for Hsp 25 was found throughout their cell bodies, intense reactivity being located near the proximal and distal terminal webs. At the maturation stage, ruffle-ended ameloblasts (RA) consistently showed Hsp 25-immunoreactivity throughout the cell bodies, whereas smooth-ended ameloblasts (SA) lacking a ruffled border were weak in immunoreaction at the distal cytoplasm. Other cellular elements of the enamel organ were negative. The subcellular localization of Hsp 25-immunoreactivity in this study appeared essentially identical to that of actin filaments as demonstrated by confocal microscopy using rhodamine-labeled phalloidin. These inununocytochemical data suggest that the Hsp 25 molecule is involved in reinforcement of the cell layer following cell movement during odontogenesis and in the formation and maintenance of the ruffled border of RA.

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A new “orcein-picroindigocarmine staining”, a colour combination of orcein, indigo carmine, and picric acid, was developed for histological applications. The new technique was tested on different human tissues.
  Colours ranging from red to brown, yellow, green and blue were observed in paraffine sections of tissues stained by this method. Nuclear structures in all tissues were stained dark brown to dark blue. Squamous epithelium was stained light brown with varying shades of blue in upper horny layers, whereas the ciliated epithelium was tinged blue grey. When connective tissue was stained, collagen fibrils appeared strongly blue next to elastic fibres, which took on a rust brown tinge; cellular components were all coloured brown. The matrix of hyaline cartilage was stained in different shades of blue, with the chondrocytes rust brown. Sections of bone components appeared dark blue to dark green. Skeletal muscle cells were coloured yellow and green with blue collagenous septa.
  The new staining is useful for distinguishing connective tissue components such as elastic fibres and collagen fibrils. It also demonstrates chondrocytes in favourable contrast to the cartilage matrix.
The technique produces aesthetic staining colouring that could supplement histological investigations and provide an alternative to other staining materials.

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