The airway mucosa is known to be densely supplied with several types of peptidergic nerve fibers. The distribution of the nerve fibers in a large extent of the mucosa, however, has been difficult to visualize by observation of conventional thin sections. In the present study, whole mount preparations of the rat tracheal mucosa were designed to demonstrate calcitonin generelated peptide (CGRP)-containing nerves which are most predominant among peptidergic nerves in the trachea. The mucosal sheet of the trachea was separated from the cartilaginous base by means of dispase, a protease, to be processed to immunohistochemistry for CGRP.
In the cartilaginous portion, thick nerve bundles immunoreactive for CGRP ran transversely between the cartilage rings, sending terminal branches toward the epithelium. In the membranous portion, immuno-reactive nerve bundles were thinner than those in the cartilaginous portion and ran longitudinally. A very dense nerve plexus of CGRP-immunoreactive fibers was demonstrated to extend in the basal part of the epithelium; every epithelial cell appeared to contact more than one nerve fiber. Immunohistochemistry of the whole mount preparations also clearly demonstrated the distribution and entire shape of broncho-pulmonary paraneurons scattered in the epithelium.
Ultrastructurally, the CGRP-immunoreactive intraepithelial nerves were found to lack myelin and Schwann sheaths, and to run through the bases of the epithelial cells; they were most frequently surrounded by the cytoplasmic processes of the epithelial cells.
The present study demonstrates that immunohistochemistry of whole mount preparations is a useful tool to morphologically analyze neuronal and paraneuronal distribution throughout the tracheal epithelium.

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The extracellular matrix produced by mono-layer and tridimensional cultures of fibroblasts was investigated using histochemical and ultrastructural methods.
In monolayer cultures, collagen and proteoglycans produced by fibroblasts could not be organized into morphologically recognizable structures. Tridimensional fibroblast cultures produced a well organized matrix with periodic, parallel ordered collagen fibrils of 50nm diameter, criss-crossed by alcianophylic segments 6-10nm thick in diameter and 100-300nm in length, parallel to each other, perpendicular to the collagen fibrils and spaced 67nm from each other. Some alcianophylic segments lay perpendicular to the above described ones, with maximum lengths of 65-70nm. Alcianophylic segments are the ultrastructural evidence of structural proteoglycans. These observations suggest that the culture conditions influence the collagen and proteoglycans secretion, so that the final organization of the matrix results quite different.

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CGRP, substance P (SP), VIP and somatostatin were demonstrated in the internal gills of larval bullfrogs using the immunohistochemical peroxidase-antiperoxidase method. Three groups of immunoreactive fibers were distinguished by their distribution area. The first group includes CGRP, SP and VIP immunoreactive fibers running in the connective tissue of the gill tufts. These fibers were associated with the capillaries. Some CGRP fibers were distributed similarly to SP fibers, and the density of VIP immunoreactive fibers was low. The second group includes CGRP, SP, VIP and somatostatin fibers distributed in the connective tissue surrounding the ceratobranchialia. The third group includes CGRP, SP and somatostatin fibers located in the branchial muscle. Their terminals appeared to be neuromuscular junctions. No immunoreactivity of leuor met-enkephalins was found in the internal gills. From these findings, the first group of fibers is presumed to be involved in modulating ion transport of the internal gills. The second group of fibers except for the somatostatin fibers is thought to be continuations of the first group of fibers. The third group of fibers may possibly be involved in the modulation of transmissions at the neuromuscular junction. It is unclear whether somatostatin terminals are also involved in this.

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The numbers of neurons of the ganglion cell layer (GCL) and their distribution in the retina of an Australian lizard Ctenophorus nuchalis were investigated. Retinal wholemounts and sections were prepared for light microscopic and optic nerves for electron microscopic study. Counts of cell numbers in the GCL from wholemounts varied from 200, 000 to 380, 000. Neurons in the GCL were non-uniformly distributed, forming a high cell density streak along the naso-temporal axis of the retina. Neurons of the GCL formed 2 to 9 layers in the visual streak and a single layer in the rest of the retina. The number of neurons of the GCL in this area was estimated at about 2, 100, 000. Althouth the visual streak represented only 16% of the total retinal surface area, it contained about 90% of all neurons of the GCL. Optic axon counts yielded 147, 000 myelinated and 2, 643, 000 unmyelinated fibres. The estimated optic fibre number of 2, 790, 000 was 18.2% less than the total number of neurons counted from sections in the GCL of the same eye. The unexpected high number of neurons in the area of the visual streak indicates that cell numbers obtained only from wholemount preparations may vastly underestimate the total neuron numbers in the GCL of the lizard retina.

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To examine the ultimate fate of hypertrophic chondrocytes, Meckel's cartilage bars from 18-day-old mouse embryos were transplanted into isogenic mouse spleen for 3, 7, 14 and 21 days and observed at the light and electron microscopic levels.
The midportions of these Meckel's cartilage bars were used as explants; they were characterized by many hypertrophic chondrocytes containing euchromatic round nuclei, a large amount of glycogen particles, and some vacuoles. Grafted cartilage adapted well to the splenic tissue, showing intense metachromasia around the territorial matrix. Ultrastructural observations indicated that the number of large vacuoles and glycogen aggregates in the hypertrophic cells became markedly reduced with grafting time, whereas the Golgi apparatus and rough endoplasmic reticulum were welldeveloped. Needle-like crystals showing initial apatite deposition appeared in association with matrix vesicles; these proliferated as time elapsed after transplantation.
On day 14 after transplantation, cells displaying such various structural features as pyknotic nuclei, large vacuoles, and cytoplasmic shrinkage were noted in addition to intact hypertrophic chondrocytes. Following resorption of the calcified cartilage by multinucleated giant cells, many osteoblasts appeared along the border of the calcified matrix. Some remaining hypertrophic cells in the calcified matrix had transformed into osteocyte-like cells.
On day 21, the resorbed area of the calcified cartilage was invaded by many blood vessels. Hypertrophic chondrocytes, now exposed from cellular lacunae, and the osteocyte-like cells in the calcified matrix displayed involutional changes.
The present study showed that, although the hypertrophic chondrocytes in Meckel's cartilage essentially underwent regressive changes, they retained the ability to stimulate endochondral ossification within the microenvironment of the spleen. In addition, some of these cells were transformed into osteocyte-like cells.

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Histological and histochemical observations of more than 150 extracted human deciduous teeth revealed that, prior to shedding, odontoclastic resorption as a rule takes place at the pulpal surface of coronal dentin. We also found that this phenomenon occurs in all kinds of deciduous teeth. The process of this internal resorption of coronal dentin of deciduous teeth clearly showed time-related histological changes. During the time the roots were actively being resorbed, the pulpal tissue retained its normal structure. However, when root resorption neared completion, inflammatory cells started to gradually infiltrate into the pulp, and odontoblasts began to degenerate. After that, multinucleate odontoclasts appeared, and resorption proceeded from the predentin to the dentin. The odontoclastic activity was initially detected only on the pulpal surface at the bottom areas of the crown. It gradually spread towards the pulpal horn regions along the wall of the pulp chamber. However, this internal resorption of coronal dentin did not continue until the teeth were finally shed. After the elimination of resorption, the resorbed dentin surface was repaired by a cementumlike deposition or covered with fibrous connective tissue.

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Splenic white pulp of the Aleutian skate (Bathyraja aleutica), an elasmobranch, was investigated using light and transmission electron microscopy. The major cellular constituent was plasma cells, of the typical Marshalko type, characterized by well developed rough-endoplasmic reticulum and a Golgi complex. The morphology of the rough-endoplasmic reticulum was variable, being lamellar in some cells and spherical in others. Plasma cells with distended cisternae of rough-endoplasmic reticulum and cells with Russel bodies were often observed. Only a small number of lymphocytes were encountered. These findings indicate that the splenic white pulp is the major site for immunoglobulin production in this fish, thus confirming our previous immunocytochemical observation. Globules presumably containing immunoglobulin were found consistently associated with the Golgi complex. The secretion mechanism of immunoglobulin by plasma cells is discussed in connection with the globules.

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As reptiles are situated phylogenetically between the amphibians and the mammals, they exhibit considerable variation in the structure of their tongues. The present study, one of a series of studies on reptile tongues, aims to demonstrate the three-dimensional structure of the dorsal lingual surface of a turtle, the Japanese terrapin Clemmys japonica, and to clarify the ultrastructural features of the lingual epithelial cells. In the study lingual papillae were observed by scanning electron microscopy to be widely distributed over the dorsal surface of the tongue. Irregularly shaped (conical, columnar or angular) papillae were located in the anterior and central areas, and ridge-like ones, in the latero-posterior area. Histological examination revealed that the connective tissue penetrated into the core of the papillae, and the epithelium was of a stratified squamous and/or cuboidal type. Under the transmission electron microscope, two types of cells were identified in the intermediate layer of the apical epithelium of the lingual papilla: one type was probably an immature mucous cell, whereas the other was elongated in a baso-apical direction, its cytoplasm containing fine granules. In the surface layer of the apical epithelium, typical mucous cells and cells containing numerous, fine, electron-lucent granules were recognized. Both types of cells possessed microvilli on their free-surfaces. In the lateral epithelium of the lingual papillae, the cytological features from the basal layer to the superficial intermediate layer were essentially the same as in the apical epithelium. However, in the surface layer, mucous cells were significantly larger in number than in the apical epithelium.

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The pericellular plexus of Purkinje cell axons in the cerebellar nuclei was first recorded by Cajal (1911), using the Golgi method. The present study observed the axonal plexus in the rat by scanning electron microscopy after the plexus' detachment from the soma of target neurons by NaOH maceration. The pericellular plexus revealed numerous axons with ellipsoidal and moniliform swellings. They branched and crossed with each other to form, as a whole, a reticulum which enveloped the target neuron instead of multiple isolated boutons. Some axon terminals were separated from each other by thin glial processes, while others lacked such septa, thus being directly juxtaposed. Immunohistochemistry for spot 35 protein, a Purkinje cell-specific protein, detected similar beaded and reticular axons terminating on the neuronal somata, confirming their identification as Purkinje cells. Transmission electron microscopic observation showed that most of the axon terminals in question contained elliptical vesicles, characteristic for Purkinje cells. The vesicles were not accumulated toward small patchy areas of synaptic specialization but disseminated along the entire length of the terminal portion of axons.

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The architecture and surface structures of the primary and secondary osseous spiral laminae in the cochlea of the bat, an animal able to hear high frequency sounds, were examined by scanning electron microscopy to understand the micromechanical adaptations of the bony supportive elements in the inner ear to the specific hearing function. The bat used was Myotis frater kaguyae.
The myotis bat cochlea was seen to consist of a hook and a spiral portion with one and three-quarter turns and was characterized by: 1) a distinct ridge-like projection running spirally along the middle line on the vestibular leaf of the primary osseous spiral lamina; 2) a wide secondary osseous spiral lamina; and 3) a narrow spiral fissure between the primary and secondary osseous spiral laminae. The ridge on the primary osseous spiral lamina was 150μm high in the hook and basal turn, then lowered toward apex, and flattened before the apical end. The surface structures appeared to provide a firm anchorage of the auditory teeth. The secondary osseous spiral lamina, which anchors the fibers of the basilar membrane, was sharply projected and measured 150μm in width in the hook, and then narrowed gradually toward apex to disappear in the helicotrema. The spiral fissure for the basilar membrane was about 40μm in width in the hook and about 120μm in the apical turn.
The findings suggest the presence of a narrow and rigid basilar membrane with a high amount of fibers and a strong bony support to the auditory teeth for scaffolding the tectorial membrane during intense vibration in response to high frequency sounds. The above-mentioned specialized structures of the primary and secondary osseous spiral laminae in the bat cochlea appear to adapt to the micromechanics of high frequency hearing functions.

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