The subcommissural organ (SCO) of the soft-shelled turtle was studied with the electron microscope.
In the SCO cells, two types of secretory substances are distinguishable; one is represented by secretory vacuoles containing a fine flocculent material which is closely associated with the cisternae of the granular endoplasmic reticulum, and the other by granules with homogeneously dense or finely granular contents which are surrounded by a limiting membrane and believed to be derived from the Golgi apparatus. Both types of secretions are scattered throughout the cytoplasm and are abundant especially in the apical region near the ventricle as well as in the basal processes abutting on capillaries.
In the hypendyma large spaces connecting with the perivascular ones are often observed. Their interior is filled with fine flocculent substances similar to the secretions of the SCO cells. In the endothelial cells lining the capillary lumen also, there are vacuoles frequently containing similar dense or flocculent substances.
The findings of the perivascular spaces and the endothelial cells described are suggestive of a possible basal secretion from the SCO cells of the soft-shelled turtle.

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The modified KUSHIDA and FUJITA method (1967) for in situ PTA staining was applied to the kidney, heart, spleen and brain of the normal adult albino rat. In general, intracellular membranes such as plasma membranes, sarcolemma, membranes constituting junctional areas (synapses, macula adherens, macula occludens etc.), mitochondrial membranes, limiting membranes of lysosomes, membranes of the endoplasmic reticulum including the nuclear envelope were in the negative image, while the mitochondrial matrix, myofilaments, substances in the Z line, the matrix of lysosomes, and nucleoproteins such as ribosomes, chromatin and perichromatin substance were in the positive image. Substances in the cisternae of the endoplasmic reticulum and the basement membrane were also stained moderately positive. Structural similarities observed among the erythrocytic subsurface PTA-positive zone, the subpostsynaptolemmal PTA-positive zone and the macula adherens (grouped as “subplasmalemmal PTA-positive zone”) were suggested in relation to their possible role in ionic movement.
The electron lucent layer of membranes seems to correspond to the classical osmiophilic layer. The membrane showed, however, a triple-layered unit membrane-like structure following poststaining with uranyl acetate and/or lead citrate. The PTA-negative electron lucent leaflet of the membrane may be composed of substances with non-polar groups or with a very low concentration of polar groups. Some doubts on the ROBERTSON's unit membrane theory based on the DAVSON-DANIELLI model were expressed.
It can be concluded in the present study that ultracytochemistry sine osmium tetroxide is promising for some specific purposes in electron microscopy.

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On the basis of the fact that orcein-stained elastic fibers are not decolorized in an intense alkaline medium, a new technique was introduced by which tissue segments of the artery, prior to teasing in caustic potash, were stained with orcein solution containing a surface active agent. The fenestrated membrane could be more readily detected in many of the human arteries such as coronary, basilar, radial, and anterior tibial arteries. Some pictures were presented which suggest variations of the openings in the membrane according to the site of artery and the age of the persons examined.

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The locus coeruleus of the rat was studied with the Golgi method and electron microscope in an attempt to demonstrate morphological substrates for the functional significance of these monoamine neurons.
The locus coeruleus is mainly composed of densely packed medium-sized, spindle-shaped nerve cells with frequent soma spines, while in the rostral part of the nucleus the medium-sized cells are less densely located and mingled with small oval cells. Most of the axons originating from the medium-sized nerve cells take a rostro-ventral course usually giving off an initial collateral. It is interesting that some of the axons demonstrate a curious recurrent way or run caudalwards to the medulla oblongata after a short rostro-ventral course. Afferent fibers are composed of thick and fine fibers, which make complicated neuropil around the nurons. The thick afferents enter the nucleus from a ventral to dorsal direction and take an undulating tortuous course among the neurons, revealing axonal swelling and giving off many terminal boutons.
The cytoplasm of the medium-sized nerve cells demonstrates marked aggregates of rough surfaced ER and prominent development of Golgi complexes, and contains a large number of lysosomal dense bodies. Large granular vesicles as seen in the terminal bags are only small in number in the cytoplasm. Cytoplasmic protrusions are characteristic for the medium-sized nerve cells, being moderate in number, varying in shape and length (0.4-1.7μ) and constituting mostly the postsynaptic sites. Axo-somatic synapses are frequent which make contact with axon terminals similar to axo-soma spine synapses. The nuropil contains different kinds of axo-dendritic and axo-dendritic spine synapses. Occasionally one can observe thickened preterminal axons or axon terminals contacting several dendrites or dendritic spines. Astroglial cells fill up the interneuronal spaces and demonstrate large oval mitochondria with homogeneous content and occasional lipofuscin granules.

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Die Oberflächendeckschicht des Ziliarepithels eines Kaninchens wurde elektronenmikroskopisch-histochemisch untersucht. In Bezug auf den Blockierungseffekt für die Eisenbindungsreaktion lassen sich die verschiedenen Invertseifen in folgender Reihe anordnen: Cetylpyridiniumchlorid (CPC)>Cetyldimethylbenzylammoniumchlorid (CDAC), Cetyltrimethylammoniumbromid (CTAB), Acrinol>Benzetoniumchlorid (BC). Die Eisenbindungsvermögen des Plattenepithels (vorderer Abschnitt der Ziliarfortsätze) sind durch 3×10^-3M CPC beinahe vollständig blockiert, während für die Blockierung der positiven Reaktion des Zylinderepithels (hinterer Abschnitt) 5××10^-3M CPC erforderlich ist. Auf Grund dieser Ergebnisse wurden die Kohlenhydratkomponenten der polysaccharidreichen Oberflächendeckschicht diskutiert, und die beschriebenen Blockierungsverfahren für die feine histochemische Analyse solcher sauren Mucosubstanzen als geeignet vorgeschlagen.

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Results of a previous experiment in which the stomachs of rats were immersed in 33% alcohol and the gastric surface epithelial cells were separated so that productin of FUJIE in the surface epithelial cells might be transfused into the alcohol showed that a factor promoting production of secretory granules in the gastric chief cells (viz. productin-FUJIE) was contained in the alcohol. The presence of this factor was proved by the cytological observations of the chief cells following injection of the above mentioned alcohol into unfed rats.
Chemical color testing for reaction of amino acid or protein showed the presence of a phenol group and an imidazol group in the sub-nuclear portion of the gastric surface epithelial cells where productin vacuoles are seen.
Analysis by amino acid analyser showed that the alcohol after immersion of the tissue contained 18 kinds of amino acids and 5 kinds of related compounds.
In this study, pure chemicals corresponding to these extracted substances were injected into unfed rats and the secretory activity of the gastric chief cells was studied morphologically in an attempt to identify a substance producing the same effects as productin.
Arginine, histidine, lysine and tyrosine actively promoted the production of secretory granules in the chief cells while asparagine and serine promoted the vacuolization of the granules. The possibility of these substances as the components of productin was suggested in connection with the histochemical evidence (FUJIE et al. 1957) on the occurrence of the phenol group (tyrosine) and the imidazol group (histidine) in the surface epithelial cells of the stomach.

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The ultimobranchial bodies of two species of elasmobranchs-smooth dog fish, Mustelus manazo BLEEKER and sting ray, Dasyatis akajei MÜLLER and HENLE-were observed with the electron microscope.
1. In the ultimobranchial bodies of smooth dog fish many vesicles are scattered in abundant connective tissue, while in those of sting rays vesicles and cell cords are closely packed.
2. Two types of cells, barrel cells and finger cells, are found in the vesicles in both species. The barrel cells are more endocrine in nature, and the finger cells are more exocrine or supporting. In sting rays the barrel cells are more prevalent in number than the finger cells and the cell cords are exclusively composed of the barrel cells.
3. The barrel cells are round to oval and do not directly limit the vesicle cavity. Many secretory granules are stored in the cytoplasm, and are often in direct contact with the lateral or basal plasma membranes without changing their size and density. The secretory granules appear to be formed in the Golgi zone. Occasionally, filament bundles appear around the nucleus where mitochondria, secretory granules and rough-surfaced endoplasmic reticulums are entangled. The secretory granules are larger and denser in sting rays than in smooth dog fish.
4. The finger cells have elongated cell bodies, well developed interdigitations and desmosomes and surround the barrel cells. They limit the vesicle cavity with free apical surfaces from which short microvilli project to the lumen.

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Periodic acid-silver methenamine was used to stain the middle piece of the atypical spermatozoa of Cipangopaludina malleata REEVE at the level of electron microscopy. It was found that the PAS-positive layer of the middle piece consists of glycoprotein which seems to have originated from the DNA molecules. Glycoprotein was revealed neither on the plasma membrane nor on the limiting membrane of any of the bodies constituting the middle piece nor in the tail flagella.

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1. A method of electron microscope autoradiography was established using nuclear research emulsions manufactured in japan: Sakura NR-H1 and NR-H2. Several biological applications were performed, and simultaneously the improvement in resolution was studied.
2. The aim of electron microscope autoradiography is to reveal the exact localization of certain substances at the macromolecular level. In order to attain this object the establishment of an extremely fine grain development method is indispensable.
3. Although the limit of resolution in electron microscope autoradiography is supposed to be the diameter of silver halide grains in emulsion, it may be improved by considering the fact that the contact area between the silver grain and the section surface is the minimum unit of resolution.
4. Thiamine, labelled with ³H, was made insoluble and precipitated with chloroplatinic acid at fixation. Developed silver grains completely overlap the B₁-PtCl₆ precipitates.
5. The minimum resolution of the electron microscope authoradiogrphy was determined histologically to be within 100Å. The following hypothesis is proposed to explain this high resolution although the introduction of extremely fine grain development is also considered one of the factors.
Some of latent images are formed at the contact point (surface) between the polyhedral silver halide grain and the section surface. The latent image seems to be formed by the impact of β-particles from directly below, and since it is in contact with the section it remains in place even after development and gelatin removal. This latent image finally becomes a developed silver grain in the electron microscope autoradiogram.

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Histological studies were made of thymus grafts in dd-mice. Whole thymuses were removed from neonatal female mice, and grafted to the mother animals in the subcapsular space of the kidney. Thymus grafts rapidly regenerated, grew and then involuted in almost the same way as the normal thymus. Immediately after grafting, thymus grafts showed an extensively necrotic central area surrounded by a narrow peripheral zone of surviving cortical tissue. By 8 days after grafting, grafts regenerated from the peripheral zone and restored a structure essentially identical to the normal thymus. Most histological features of thymus grafts were similar to those of the normal thymus of corresponding age, although the cortex and medulla were irregularly distributed in grafts. In addition, many thymus grafts contained lymphoid nodules with germinal centers within the medulla. The mechanism by which lymphoid nodules appeared in thymus grafts is unknown, but is discussed in some detail in light of the recent information available.

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With rat fetuses of the 8th to 20th day of gestation and with neonates and postnates of one to seven days of age, the cytogenesis of anterior pituitary cells was investigated light and electron microscopically. The activity of pituitary trophic hormones was evaluated in reference to the histological findings on the developing target organs.
The anterior lobe of rat fetuses, outgrowing from Rathke's pouch on the 15th day of gestation, consists of a single kind of agranular primordial cell. From the 17th day of gestation, they begin to granulate and turn into ambiguous cells containing minute granules, whose developmental orientation is still unknown. From the 17th to 20th day of gestation, the growing granules suggest their transformation into undifferentiated basophils. During late pregnancy the cellular transformation proceeds exclusively along the basophil axis, and undifferentiated thyrotrophs gain β-granules, and the follicles of the thyroid show a gradual development throughout perinatal life. In the late embryonal life, gonadotrophs with their characteristic large granules develop considerably in males. As this is timed by a transitory increase of Leydig cells in the testis, a discharge of gonadotrophin at this period is suggested. The involution of some gonadotrophs after birth is concurrent with that of Leydig cells. The development of gonadotrophs is delayed in female fetuses. No gonadotrophs reappear throughout postnatal life in either sex.
The development of acidophils is retarded to that of the cells along the basophil axis during embryonic life. The ambiguous cells developing along the acidophil axis are generally ellopsoid or elongated in shape, and begin to contain dense granules. After birth, the cellular transformation along the acidophil axis is activated.
Within 5 days of age, numerous young acidophils full of α-granules are produced. On the other hand, the fetal adrenal cortex has been substituted by the permanent cortex by the postnatal third day. The activated acidophil axis thus seems to be related to ACTH secretion.
Pituitary primordial cells in embryonal life are characterized by abundant rosettes and the absence of dense granules. From the late fetal period on, they decrease in number, having been transformed into granulated cells. Some primordial cells are retained without completing their defferentiation into the so-called chromophobes, while the target organs are still developing. Mitosis takes place frequently in primordial cells and ambiguous cells, but not in mature chromophils. The present findings support the theory of two developmental transformation axes of the anterior pituitary cells.

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Observations were made on the pancreatic islets of the snake, Elaphe climacophora, and a teleost, yellow-tail or Seriola quinqueradiata, using alternate thick and thin sections for light and electron microscopy.
Under the light microscope, the snake islets contain A-cells with red-stained secretory granules and B-cells with blue-stained granules. The islets of the yellow-tail consist of A-cells with red granules, B₁-cells with dark blue granules, B₂-cells with light blue granules and C-cells without granules (modified aldehyde thionine-hematoxylin-phloxin B).
Electron microscopy indicates that the A-cells of the snake islets contain both small dense and large, less dense granules. B-cells are divided into two types, one of which contains dense spherical granules whereas the other, less dense crystalline ones. Some crystalline granules show a hexagonal shape and a lamellar structure with a periodicity of about 90Å.
The yellow-tail islets contain five different cell types; A, B₁, B₂, C₁ and C₂ cells. The A-cells contain dense spherical granules. The B₁-cells possess dense crystal granules, some of which are hexagonal in shape like the B-cell granules of the snake. The B₂-cells contain less dense amorphous granules. The C₁-cells usually show empty vacuoles, whereas the C₂-cells contain no secretory granules. Transitional cells occur between B₁, B₂ and C₁ cells and between A and C₂ cells, suggesting that they represent functionally different states of the B and A cells rather than different cell types.
The eruptocrine or Type IV extrusion of crystalline granules of the yellow-tail B-cells was clearly shown.

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The effects of some histamine-liberating agents on rat peritoneal mast cells were examined under the electron microscope and the release of histamine was measured by means of bioassay.
Compound 48/80, when given in a small dose (0.1μg/ml), caused swelling of the granules and widening of the perigranular space only in a small part of the cells. With moderate doses (1.0-5.0μg/ml), perigranular vacuoles developed either individually causing a honeycomb-like cytoplasm or, by confluence, into a single large vacuole leaving a thin cytoplasmic sheet at the periphery of the cell. Degranulation occurred mainly in the cells with the former type of vacuolation. When a very large dose (100μg/ml) of compound 48/80 was given, vacuolation was marked but the granules were neither discharged nor swollen. Toluidine blue (50μg/ml) and sinomenine (500μg/ml) caused cytological changes in the mast cells similar to those under moderate dose treatment with compound 48/80. The effect of anti-rat rabbit serum was characterized by greater swelling of the granules and more conspicuous peripheral vacuolation than in the cases of the histamine liberators. Pretreatment with 10^-4M of 2, 4-dinitrophenol almost completely inhibited any cytological change inducible by compound 48/80.
By comparing the electron micrographs with the grades of histamine release in the corresponding cases measured by bioassay (Table 1), it was proposed that the most reliable morphological criterion of the release of histamine is neither degranulation nor granule swelling, but the occurrence of vacuoles around the granules.

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The specificity in the detection of tissue histamine was examined in three previously known histochemical methods: 1) prolonged immersion of tissue in Reinecke salt solution (SCHAUER and WERLE, 1959), 2) diazotized parabromoaniline staining or fluorescence microscopy after formaldehyde vapor-fixation (LAGUNOFF, PHILLIPS and BENDITT, 1961), 3) fluorescence microscopy after o-phthalaldehyde staining (JUHLIN and SHELLEY, 1968). Besides the normal mast cells of the rat, mouse, hamster and guinea pig, their changes caused by a histamine liberator, Polymyxin B were observed. The occurrence of histamine in the strange-shaped mast cells in the triton brain (INOUE, AKITA and SHIMAI, 1968) was also examined.
1. The o-phthalaldehyde staining applied to the fresh specimens was found to be the best of the three methods examined for the demonstration of histamine in the mast cells.
2. However, as some cells other than the mast cells, e. g. the serous cells of the salivary gland, the small blood vessel walls and part of the epithelium of the convoluted tubules of the kidney of the rat, and red and white blood cells and the small blood vessels of the mouse, hamster and guinea pig, emit simultaneously a bright yellow fluorescence, the presence and non-presence of histamine should be dicided carefully by taking into consideration the species of animals examined, the histochemical procedures used and the possible variety in the manner of the binding of histamine in the cytoplasm.
3. The process of histamine release could be revealed by o-phthalaldehyde staining applied to the mast cells of the hamster which was injected with Polymyxin B.
4. Histamine was not demonstrated in the mast cells in the mesentery and brain of the Triturus pyrrhogaster (Boie) and in the mesentery of the Rana catesbiana (Shaw).

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As a part of the phylogenetic studies of the thyroid gland, the anuran thyroid was studied by electron microscopy, electron microscopic autoradiography with ¹²⁵I and by histochemistry for acid phosphatase activity.
The fine structure of the anuran thyroid cell is almost similar to that of mammalia and fish. The rough endoplasmic reticulum is well developed in the basal part of the cell. Subapical small vesicles which have been considered to be derived from the Golgi apparatus and to contain secretory materials to the follicle lumen are also seen as in mammalia and fish. Characteristic are relatively well developed tonofilaments, numerous and large intracellular colloid droplets, and the occurrence of filamentous structures in some of the droplets. These findings are similar to those in some species of teleost and chimaeroid fish. About two thirds of the large droplets, most of the small dense granules and some parts of the Golgi apparatus show an acid phosphatase activity. The small dense granules which are sometimes in contact with the large colloid droplets are considered to be primary lysosomes derived from the Golgi apparatus. The large droplets likely correspond to reabsorbed colloid to be hydrolyzed by lysosomes. This presumed pattern of cell function in the amphibian thyroid is almost the same to that in mammals.
The capillary endothelium shows numerous pores as that of the higher vertebrates and some species of fish.
The present findings indicate that the iodination of the thyroglobulin takes place chiefly in the follicle lumen and in the apical cell membrane region.

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The islet of Langerhans of Carassius carassius longsdorfii (TEMMINCK et SCHLEGEL) has been observed by light and electron microscopy.
1. The islet tissue of the Carassius is concentrated into several spherical bodies embedded in the exocrine pancreas or in the mesentery. They are surrounded by a distinct connective tissue capsule, and richly supplied by blood capillaries.
2. By light microscopy, A, B and D cells are distinguished according to their staining reactions. The cells deserving the name of “clear” or “agranular” cells under the light microscope can be identified by electron microscopy as D cells containing no or few secretory granules. B cells form a network of cell cords throughout the islets, in the meshes of which A, D and “clear” cells are distributed. The population ratio of A:B:D cells comes to 25:50:25 (of which 10 are the light microscopic “clear cells”).
3. Argyrophil islet cells are demonstrated in the silver impregnated specimens and correspond in shape, distribution and number to the D cells.
4. Under the electron microscope the three islet cell types are distinguished by the specific ultrastructural features of their secretory granules. The D cells of the Carassius are further characterized by the abundant occurrence of the vesicles and tubules of the agranular endoplasmic reticulum.
5. The α-granules of the Carassius (200-400mμ in diameter) consist of a membranous sac encasing a homogeneous spherical core of high electron density with a slight space between the two being characteristic. The β-granules of the Carassius (200-400mμ in diameter) closely resemble those of the carp (TITLBACH, 1966), showing a membranous sac and a spherical core which is composed of a homogeneous substance and needle- or bar-shaped crystalloids; the electron lucent space between the sac and the core is inconspicuous. In the δ-granules of the Carassius (200-400mμ in diameter), a round membrane sac is closely attached to a finely granular or flocculent core of low electron density. A few smaller and denser granules may occur among the ordinary δ-granules and near the Golgi complex; they are regarded as immature α-granules.
6. In the Carassius, the formation of secretory granules in the Golgi complex is evidenced in the three islet cell types. The release of secretory granules by means of emiocytosis is confirmed in A and D cells but not in B cells. Findings suggestive of the granule release by intracytoplasmic dissolution are obtained in all three islet cell types but especially conspicuously in B cells.

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The eccrine sweat glands in the foot pads of adult mice of the C57BL strain fixed either with osmium tetroxide or with a mixture of glutaraldehyde and osmium tetroxide were observed with the electron microscope.
The secretory portion of the eccrine gland consists of glandular cells and myoepithelial cells. After simple osmium fixation, the glandular cells show a rather uniform appearance, but the fixation with glutaraldehyde and osmium may reveal strong variations in electron density of the nucleus as well as the cytoplasm. Thus, the specimens after the latter fixation show glandular cells with three different grades of density, i.e., the dark, intermediate and clear cells.
Microvilli of the dark cells are the shortest, those of the intermediate cell are intermediary and those of the clear cell are the longest, suggesting that the dark cell may be brought about through shrinkage of the cell. It is highly probable that the occurrence of dark and clear cells may be the result of fixation artefacts or of some pathological imbalance in water content from cell to cell.
It is well known that the secretory vacuoles of the human and monkey eccrine sweat glands can be demonstrated as dark granules after fixation with glutaraldehyde and osmium. Those of the mouse gland, however, are always clear not only after simple osmium fixation but also after double or mixed fixation with glutaraldehyde and osmium. This implies that the substance in the secretory vacuoles of the mouse eccrine gland is chemically different to some extent from that of the human and monkey glands.
The intercellular canaliculi which can be found in the human, monkey and cat eccrine glands and are lined exclusively with clear cells are not found in the mouse and rat glands. Therefore, it may be said that the rodent glands are primitive in structure.
The agranular endoplasmic reticulum is first found in the animal eccrine glands in this study, and the agranular reticulum is intermingled with secretory vacuoles, though in the human gland the agranular reticulum appears exclusively in the basal clear cells while secretory vacuoles are prominent in the superficial dark cells. The fact that both structures occur in one and the same cell in the mouse gland may indicate that the mouse eccrine gland is undifferentiated.
The above mentioned facts all suggest that the secretory cells of the rodent eccrine glands are essentially monotypical.
The ultrastructure of the intradermal and intraepidermal sweat ducts was also described and the keratinization process of the duct cells was discussed.

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Human duodenal mucosa obtained in three surgical operations was examined under the electron microscope with the following results:
1. In all the three cases five different types of endocrine cells occurred in the basal parts of the crypts and of the duodenal gland ducts: 1) enterochromaffin (EC) cells with dense, polymorphous granules, 2) type 2 cells with large-sized round granules, 3) type 3 cells characterized by small round granules and an infranuclear fibrous substance, 4) type 4 cells containing similar granules and characterized by a rounded shape with a constricted neck and an apical disk provided with very long microvilli, and 5) intestinal D cells with granules morphologically identical with those in the pancreatic D cells.
2. All these types of cells seemed to face the lumen of the crypts of ducts with a narrowed apical process covered with microvilli. Discussion was made on the possible significance of this structure for the perception of some information from the lumen such as the changes in pH.

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