Ultramicroscopic study of the developmental change of the xanthophore in the frog, Rana Japonica with special reference to pterinosomes

Structural change in pterinosomes and the behavior of cytoplasmic inclusions in the process of xanthophore differentiation were studied electron microscopically using Rana japonica. At the early stage of xanthophore development, type I pterinosomes had clear limiting membranes and were empty or amorphous within. The nucleus and cytoplasm were characterized by a well-developed nucleolus, mitochondria and Golgi bodies, and a large number of polysomes. At the middle stage, type II pterinosomes had indistinct limiting membranes and a few lamellae. Lipid droplets appeared almost concurrently with glycogen particles in the cytoplasm. At the later stage, type III pterinosomes had concentrically arranged lamellae, lacking clear limiting membranes. Thus, the successive transformation from types I to III was concluded. Adult xanthophores contained types I to III pterinosomes in each cell. A different mechanism is suggested of the differentiation of pterinosomes between the larval and the adult xanthophores.     

The Erythrophore in the Larval and Adult Dorsal Skin of the Brown Frog,Rana ornativentris: Its Differentiation,Migration, and Pigmentary Organelle Formation

To determine whether or not the erythrophore originates from xanthophores in the dorsal skin of the brown frog, Rana ornativentris, we morphologically examined the differentiation and migration of the two chromatophore types and their pigmentary organelle formation. At an early tadpole stage, three kinds of chromatophores, xanthophores, iridophores, and melanophores, appeared in the subdermis, whereas the erythrophore did so just before the foreleg protrusion stage. By the middle of metamorphosis, most chromatophores other than erythrophores had migrated to the subepidermal space. Erythrophores, which appeared late in the subdermis, proliferated actively there during metamorphosis and finished moving into the subepidermal space by the completion of metamorphosis. Carotenoid vesicles and pterinosomes within the erythrophores and xanthophores showed several significant differences in structure. In xanthophores, carotenoid vesicles were abundant throughout life, whereas those in erythrophores decreased in number with the growth of the frogs. The fibrous materials contained in the pterinosomes were initially scattered but soon formed a concentric lamellar structure. In erythrophores, the lamellar structure began to form at the periphery of the organelles but at the center in xanthophores. In addition, the pterinosomes of erythrophores were uniform in size throughout development, while those of xanthophores showed a tendency to become smaller after metamorphosis. The pterinosomes of xanthophores were significantly larger than those of erythrophores. These findings suggest that an erythrophore is not a transformed xanthophore, although they resemble each other closely in many respects.     

Pigmentary system of the adult alpine salamander Salamandra atra aurorae (Trevisan, 1982)

The pigmentary system of skin from adult specimens of the amphibian urodele Salamandra atra aurorae was investigated by light microscope, electron microscope, and biochemical studies. Yellow (dorsum and head) and black (flank and belly) skin was tested. Three chromatophore types are present in yellow skin: xanthophores, iridophores, and melanophores. Xanthophores are located in the epidermis whereas iridophores and melanophores are found in the dermis. Xanthophores contain types I, II, and III pterinosomes. Some pterinosomes are very electron-dense. Black skin has a single type of chromatophore: the melanophores. Some melanophores are located in the epidermis. In contrast to the dermal melanophores, these present, in addition to typical melanosomes, organelles with different morphology and vesicles having a limiting membrane and containing little amorphous material. Both skin types present some pteridines and flavins, though they are qualitatively and quantitatively more abundant in yellow skin extracts.     

Pigment cell differentiation in the fire-bellied toad,Bombina orientalis. I. Structural,chemical, and physical aspects of the adult pigment pattern

Wild-collected adults of Bombina orientalis are bright green dorsally and red to red-orange ventrally. As a prelude to an analysis of the differentiation of pigment cells in developing B. orientalis, we describe structural and chemical aspects of the fully differentiated pigment pattern of the “normal” adult. Structurally, differences between dorsal green and ventral red skin are summarized as follows: (1) Dorsal green skin contains a “typical” dermal chromatophore unit comprised of melanophores, iridophores, and xanthophores. Red skin contains predominantly carotenoid-containing xanthophores (erythrophores), and skin from black spot areas contains only melanophores. (2) In ventral red skin, there is also a thin layer of deep-lying iridophores that presumably are not involved in the observed color pattern. (3) Xanthophores of red and green skin are morphologically distinguishable from each other. Dorsal skin xanthophores contain both pterinosomes and carotenoid vesicles; ventral skin xanthophores contain only carotenoid vesicles. Carotenoid vesicles in dorsal xanthophores are much larger but less electron dense than comparable structures in ventral xanthophores. The presence of carotenes in ventral skin accounts for the bright red-orange color of the belly of this frog. Similar pigments are also present in green skin, but in smaller quantities and in conjunction with both colored (yellow) and colorless pteridines. From spectral data obtained for xanthophore pigments and structural data obtained from the size and arrangement of reflecting platelets in the iridophore layer, we attempt to explain the phenomenon of observed green color in B. orientalis.     

MORPHOLOGICAL AND BIOCHEMICAL CHARACTERIZATION OF GOLDFISH ERYTHROPHORES AND THEIR PTERINOSOMES

The fine structure of integumental erythrophores and the intracellular location of pteridine and carotenoid pigments in adult goldfish, Carassius auratus, were studied by means of cytochemistry, paper and thin-layer chromatography, ionophoresis, density-gradient centrifugal fractionation, and electron microscopy. The ultrastructure of erythrophores is characterized by large numbers of somewhat ellipsoidal pigment granules and a well-developed system of tubules which resembles endoplasmic reticulum. The combined morphological and biochemical approaches show that pteridine pigments of erythrophores are located characteristically in pigment granules and are the primary yellow pigments of these organelles. Accordingly, this organelle is considered to be the "pterinosome" which was originally found in swordtail erythrophores. Major pteridines obtainable from goldfish pterinosomes are sepiapterin, 7-hydroxybiopterin, isoxanthopterin, and 6-carboxyisoxanthopterin. Density-gradient fractions indicate that carotenoids are mostly associated with the endoplasmic reticulum. Both tyrosinase and possibly a tyrosinase inhibitor containing sulfhydryl groups are present in the pterinosome. The possible existence of a tyrosinase inhibitor is suggested by the marked increase of tyrosinase activity upon the addition of iodoacetamide or p-chloromercuribenzoic acid. In the light of their fine structure, pigmentary composition, and enzymatic properties, the erythrophores and pterinosomes are discussed with respect to their probable functions and their relationship to melanophores.     

Origin of melanosome structures and cytochemical localizations of tyrosinase activity in differentiating epidermal melanocytes of newborn mouse skin

The mode of differentiation of epidermal melanocytes was studied by ultrastructural cytochemistry in the skin of newborn mice of strain C57BL/10J. From observations of epidermal melanoblasts and melanocytes, stage I melanosomes, including both unit membranes and inner matrices, appear to be formed from Golgi vacuoles or rough endoplasmic reticulum (RER). Stage I melanosomes were positive to ammoniacal silver-nitrate reaction in the melanoblasts of 1-day-old mice. All stages of melanosomes were similarly positive in the differentiating melanocytes of 2-day-old mice. However, Golgi apparatus, RER, and vesicles were negative. Therefore, it is conceivable that structural proteins, originated from Golgi vacuoles or RER, are developed into specialized proteins and are detected by this reaction in stage I melanosomes. Stage I melanosomes were dopa-negative in the melanoblasts. Stage I and II melanosomes were similarly negative in the differentiating melanocytes. Thus, the melanoblasts are thought to begin production of stage I melanosomes prior to the onset of tyrosinase activity. In the differentiating melanocytes, dopa-melanin depositions were observed in stage III and IV melanosomes, trans Golgi saccules, and small vesicles derived from these saccules, but not in RER. These vesicles were in contact with, or fused to, melanosomes. These findings suggest that tyrosinase may be transferred by Golgi vesicles into stage I and II melanosomes originating from Golgi vacuoles or RER.     

Analysis of xanthophore and pterinosome biogenesis in zebrafish using methylene blue and pteridine autofluorescence

We have identified two simple methods to analyse xanthophore and pterinosome biogenesis in zebrafish. The first uses methylene blue (methylthionium chloride), a redox dye which specifically labels xanthophores and pterinosomes, while the second uses autofluorescence to detect pteridine levels; these methods may be used to detect the number, location and shape of xanthophores and pterinosomes. These assays were applied to two zebrafish mutants--brie and yobo--and revealed that both mutants have pterinosome biogenesis and pteridine synthesis defects. Additionally, using capillary electrophoresis, we provide evidence that sepiapterin is responsible for the yellow colour and blue-light induced fluorescence in zebrafish embryos.     

Temporal and cellular requirements for Fms signaling during zebrafish adult pigment pattern development

Ectothermic vertebrates exhibit a diverse array of adult pigment patterns. A common element of these patterns is alternating dark and light stripes each comprising different classes of neural crest-derived pigment cells. In the zebrafish, Danio rerio, alternating horizontal stripes of black melanophores and yellow xanthophores are a prominent feature of the adult pigment pattern. In fms mutant zebrafish, however, xanthophores fail to develop and melanophore stripes are severely disrupted. fms encodes a type III receptor tyrosine kinase expressed by xanthophores and their precursors and is the closest known homologue of kit, which has long been studied for roles in pigment pattern development in amniotes. In this study we assess the cellular and temporal requirements for Fms activity in promoting adult pigment pattern development. By transplanting cells between fms mutants and either wild-type or nacre mutant zebrafish, we show that fms acts autonomously to the xanthophore lineage in promoting the striped arrangement of adult melanophores. To identify critical periods for fms activity, we isolated temperature sensitive alleles of fms and performed reciprocal temperature shift experiments at a range of stages from embryo to adult. These analyses demonstrate that Fms is essential for maintaining cells of the xanthophore lineage as well as maintaining the organization of melanophore stripes throughout development. Finally, we show that restoring Fms activity even at late larval stages allows essentially complete recovery of xanthophores and the development of a normal melanophore stripe pattern. Our findings suggest that fms is not required for establishing a population of precursor cells during embryogenesis but is required for recruiting pigment cell precursors to xanthophore fates, with concomitant effects on melanophore organization.     

GOLGI APPARATUS, GERL, AND LYSOSOMES OF NEURONS IN RAT DORSAL ROOT GANGLIA, STUDIED BY THICK SECTION AND THIN SECTION CYTOCHEMISTRY

New insights into the ultrastructure and phosphatase localizations of Golgi apparatus and GERL, and into the probable origin of lysosomes in the neurons of fetal dorsal root ganglia and the small neurons of adult ganglia have come from studying thick (0.5–1.0 µ) as well as thin (up to 500 A) sections by conventional electron microscopy. Tilting the thick specimens, by a goniometer stage, has helped to increase our understanding of the three-dimensional aspects of the Golgi apparatus and GERL. One Golgi element, situated at the inner aspect of the Golgi stack, displays thiamine pyrophosphatase and nucleoside diphosphatase activities. This element exhibits regular geometric arrays (hexagons) of interconnected tubules without evidence of a flattened portion (saccule or cisterna). In contrast, GERL shows acid phosphatase activity and possesses small cisternal portions and anastomosing tubules. Lysosomes appear to bud from GERL. Osmium deposits, following prolonged osmication, are found in the outer Golgi element. Serial 0.5-µ and thin sections of thiamine pyrophosphatase-incubated material demonstrate that, in the neurons studied, the Golgi apparatus is a continuous network coursing through the cytoplasm. Serial thick sections of acid phosphatase-incubated tissue suggest that GERL is also a continuous structure throughout the cytoplasm. Tubules of smooth endoplasmic reticulum, possibly part of GERL, extend into the polygonal compartments of the inner Golgi element. The possible physiological significance of a polygonal arrangement of a phosphatase-rich Golgi element in proximity to smooth ER is considered. A tentative diagram of the Golgi stack and associated endoplasmic reticulum in these neurons has been drawn.     

Immunofluorescence evidence for cytoskeletal rearrangement accompanying pigment redistribution in goldfish xanthophores

Immunofluorescence and phase-contrast microscopic studies of goldfish xanthophores with aggregated or dispersed pigment show two unusual features. First, immunofluorescence studies with anti-actin show punctate structures instead of filaments. These punctate structures are unique for the xanthophores and are absent from both goldfish dermal non-pigment cells and a dedifferentiated cell line (GEM-81) derived from a goldfish xanthophore tumor. Comparison of immunofluorescence and phase-contrast microscopic images with electron microscopic images of thin sections and of Triton-insoluble cytoskeletons show that these punctate structures represent pterinosomes with radiating F-actin. The high local concentration of actin around the pterinosomes results in strong localized fluorescence such that, when the images have proper brightness for these structures, individual actin filaments elsewhere in the cell are too weak in their fluorescence to be visible in the micrographs. Second, whereas immunofluorescence images with anti-tubulin show typical patterns in xanthophores with either aggregated or dispersed pigment, namely, filaments radiating out from the microtubule organizing center, immunofluorescence images with anti-actin or with anti-intermediate filament proteins show different patterns in xanthophores with aggregated versus dispersed pigment. In cells with dispersed pigment, the punctate structures seen with anti-actin are relatively evenly distributed in the cytoplasm, and intermediate filaments appear usually as a dense perinuclear band and long filaments elsewhere in the cytoplasm. In cells with aggregated pigment, both intermediate filaments and pterinosomes with associated actin are largely excluded from the space occupied by the pigment aggregate, and the band of intermediate filaments surrounds not only the nucleus but also the pigment aggregate. The patterns of distribution of the different cytoskeleton components, together with previous results from this laboratory, indicate that formation of the pigment aggregate depends at least in part on the interaction between pigment organelles and microtubules. The possibility that intermediate filaments may play a role in the formation/stabilization of the pigment aggregate is discussed.     

Immunoelectron microscopic localization of tyrosinase in the mouse melanocyte

In the melanocyte, tyrosinase is known as the dey enzyme for melanin formation. Purified tyrosinase protein was prepared that was capable of oxidizing tyrosine. The localization of tyrosinase antigen in the melanocyte was studied using antiserum against tyrosinase. DOPA (L-3,4-dihydroxyphenylalanine)-reaction product and tyrosinase antigen were found on the same organelles i.e., premelanosomes, melanosomes, GERL, and Golgi vesicles. This result seems to suggest that it is cytochemically appropriate to use DOPA as the substrate of tyrosinase. It appeared that tyrosinase antigen was present as granule-like structures inside GERL cisterna and associated with its membrane.     

ON THE POSSIBLE FUNCTION OF COATED VESICLES IN MELANOGENESIS OF THE REGENERATING FOWL FEATHER

How tyrosinase becomes associated with the premelanosomes was investigated by histochemical demonstration of tyrosinase activity by the use of dihydroxyphenylalanine (DOPA) in melanocytes of regenerating fowl feathers. The reaction product of DOPA was localized in the anastomosing membrane tubules associated with the concave side of some dictyosomes of the Golgi apparatus and in coated vesicles most of which were in connection with the dictyosomes. No reaction product was found in early premelanosomes. In premelanosomes, the reaction product of DOPA appears first in vesicles approximately 400 A in diameter which are surrounded by a matrix with a characteristic periodicity. These observations seem to allow the speculation that the coated vesicles function in the transport of tyrosinase, and that the premelanosomes are formed in a process which is not necessarily dependent on the Golgi apparatus as was assumed earlier.     

Formation of pterinosomes and carotenoid granules in xanthophores of the teleost Oryzias latipes as revealed by the rapid-freezing and freeze-substitution method

Summary The rapid-freezing and freeze-substitution method was applied for the ultrastructural study of the dermal chromatophores of a teleost, Oryzias latipes. The method was found to be suitable for preserving fragile membranous structures within melanophores and xanthophores. In addition, relatively high electron density in overall profile indicates that the procedure is effective in reducing the extraction of cytoplasmic ground substances that inevitably occurs during the process of conventional chemical fixation and the following dehydration. The improved ultrastructural images clearly show that the pterinosomes, the characteristic pigmentary organelles of xanthophores, are formed through several distinct developmental stages starting from the loose congregations of vesicles derived from the Golgi complex. The earlier stages of development are similar to those found in melanosome formation. Whereas carotenoid pigments in xanthophores in conventional aldehyde-osmium-fixed materials are found to be electrondense membrane-free particles, they are identified as membrane-bounded organelles in the present study. The envelope of these carotenoid vesicles does not exhibit a typical trilaminar structure but appears to be an extremely thin membrane. Carotenoid vesicles are, in most cases, in direct contact with the outer surface of tubular endoplasmic reticulum.     

THE GOLGI ZONE OF THE RAT SPERMATID AND ITS ROLE IN THE FORMATION OF CYTOPLASMIC VESICLES

Electron microscopic study of the Golgi zone of the rat spermatids revealed two main types of vesicular structures: flat vesicles generally accumulated in stacks and spheroidal vesicles of various sizes. The Golgi zone of young spermatids showed a predominance of flat vesicles and small spherical vesicles, while the "residual" Golgi zone of the maturing spermatids displayed an increased number of large spherical vesicles and thus became increasingly vacuolated in appearance. The images presented were suggestive of a transformation of the flat vesicles into spherical vesicles by fragmentation and dilatation and also of a passage of the spherical vesicles from the Golgi zone to the surrounding cytoplasm.     

Xanthophore migration from the dermis to the epidermis and dermal remodeling during Salamandra salamandra salamandra (L.) larval development

During larval development of Salamandra salamandra salamandra chromatophores organize to form the definitive pigment pattern constituted by a black background with yellow patches that are characterized by epidermal xanthophores and dermal iridophores. Simultaneously the dermis undergoes remodeling from the larval stage to that typical of the adult. In the present study we ultrastucturally and immunocytochemically examined skin fragments of S. s. salamandra larvae and juveniles in order to investigate the modalities of xanthophore migration and differentiation in the context of dermal remodeling from the larval to adult stage. Semithin and thin sections showed that the dermis in newly born larvae consists of a compact connective tissue (basement lamella), to which fibroblasts and xanthophores adhere, and of a loose deep collagen layer. As larval development proceeds, fibroblasts and xanthophores invade the basement lamella, skin glands develop and the adult dermis forms. At metamorphosis, xanthophores reach the epidermis crossing through the basal lamina. We examined immunocytochemically the expression of signal molecules, such as fibronectin, vitronectin, beta1-integrin, chondroitin sulfate, E-cadherin, N-cadherin and plasminogen activator, which are known to be involved in regulating morphogenetic events. Their role in dermal remodeling and in pigment pattern formation is discussed.     

Relationship between the golgi apparatus, gerl, and secretory granules in acinar cells of the rat exorbital lacrimal gland

The method of secretory granuleformation in the acinar cells of the rat exorbital lacrimal gland was studied by electron microscope morphological and cytochemical techniques. Immature secretory granules at the inner face of the Golgi apparatus were frequently attached to a narrow cisternal structure similar to GERL as described in neurons by Novikoff et al. (Novikoff, P. M., A. B. Novikoff, N. Quintana, and J.-J. Hauw. 1971. J. Cell Bio. 50:859-886). In the lacrimal gland. GERL was located adjacent to the inner Golgi saccule, or separated from it by a variable distance. Portions of GERL were often closely paralleled by modified cisternae of rough endoplasmic reticulum (RER), which lacked ribosomes on the surface adjacent to GERL. Diaminobenzidine reaction product of the secretory enzyme peroxidase was localized in the cisternae of the nuclear envelope, RER, peripheral Golgi vesicles, Golgi saccules, and immature and mature secretory granules. GERL was usually free of peroxidase reaction product or contained only a small amount. Thiamine pyrophosphatase reaction product was present in two to four inner Golgi saccules; occasionally, the innermost saccule was dilated and fenestrated, and contained less reaction product than the next adjacent saccule. Acid phosphatase (AcPase) reaction product was present in GERL, immature granules, and, rarely, in the innermost saccule, but not in the rest of the Golgi saccules. Thick sections of AcPase preparations viewed at 100 kV revealed that GERL consisted of cisternal, and fenestrated or tublular portions. The immature granules were attached to GERL by multiple connections to the tublular portions. These results suggest that, in the rat exorbital lacrimal gland, the Golgi saccules participate in the transport of secretory proteins, and that GERL is involved in the formation of secretory granules.     

Effect of colchicine on the Golgi complex of rat pancreatic acinar cells

Colchicine administered to adult rats at a dosage of 0.5 mg/100 g of body weight effected a disorganization of the Golgi apparatus in pancreatic acinar cells. The results obtained after various periods of treatment (10 min to 6 h) showed (a) changes in all components of the Golgi complex, and (b) occurrence of large vacuoles that predominated in cytoplasmic areas outside the Golgi region. The alterations in Golgi stacks concerned elements of the proximal and distal side: (a) accumulation of transport vesicles, (b) formation of small, polymorphic secretion granules, and (c) alterations in the cytochemical localization of enzymes and reaction product after osmification. Transport vesicles accumulated and accompanied short, dilated cisternae, which lack mostly the reaction products of thiamine pyrophosphatase, inosine diphosphatase, and acid phosphatase, and osmium deposits after prolonged osmification. After 4 to 6 h of treatment, accumulated transport vesicles occupied extensive cellular areas; stacked cisternae were not demonstrable in these regions. The changes on the distal Golgi side included GERL elements: condensing vacuoles were diminished; they were substituted by small, polymorphic zymogen granules, which appeared to be formed by distal Golgi cisternae and by rigid lamellae. Unusually extended coated regions covered condensing vacuoles, rigid lamellae, and polymorphic secretion granules. A cytochemical distinction between Golgi components and GERL was possible neither in controls nor after colchicine treatment. The cytochemical alterations in Golgi components were demonstrable 20-30 min following administration of colchicine; at 45 min, initial morphological changes--augmentation of transport vesicles and formation of polymorphic zymogen granules--became apparent. 20 min after administration of colchicine, conspicuous groups of large vacuoles occurred. They were located mostly in distinct fields between cisternae of the endoplasmic reticulum, and were accompanied by small osmium--reactive vesicles. Stacked cisternae were not demonstrable in these fields. Vacuoles and vesicles were devoid of reaction products of thiamine pyrophosphatase, inosine diphosphatase, and acid phosphatase. The results provide evidence that formation of stacked Golgi cisternae is impaired after colchicine treatment. The colchicine--induced disintegration of the Golgi complex suggests a regulatory function of microtubules in the organization of the Golgi apparatus.     

FUNCTIONS OF COATED VESICLES DURING PROTEIN ABSORPTION IN THE RAT VAS DEFERENS

The role of coated vesicles during the absorption of horseradish peroxidase was investigated in the epithelium of the rat vas deferens by electron microscopy and cytochemistry. Peroxidase was introduced into the vas lumen in vivo. Tissue was excised at selected intervals, fixed in formaldehyde-glutaraldehyde, sectioned without freezing, incubated in Karnovsky's medium, postfixed in OsO₄, and processed for electron microscopy. Some controls and peroxidase-perfused specimens were incubated with TPP,¹ GP, and CMP. Attention was focused on the Golgi complex, apical multivesicular bodies, and two populations of coated vesicles; large (> 1000 A) ones concentrated in the apical cytoplasm and small (<750 A) ones found primarily in the Golgi region. 10 min after peroxidase injection, the tracer is found adhering to the surface plasmalemma, concentrated in bristle-coated invaginations, and within large coated vesicles. After 20–45 min, it is present in large smooth vesicles, apical multivesicular bodies, and dense bodies. Peroxidase is not seen in small coated vesicles at any interval. Counts of small coated vesicles reveal that during peroxidase absorption they first increase in number in the Golgi region and later, in the apical cytoplasm. In both control and peroxidase-perfused specimens incubated with TPP, reaction product is seen in several Golgi cisternae and in small coated vesicles in the Golgi region. With GP, reaction product is seen in one to two Golgi cisternae, multivesicular bodies, dense bodies, and small coated vesicles present in the Golgi region or near multivesicular bodies. The results demonstrate that (a) this epithelium functions in the absorption of protein from the duct lumen, (b) large coated vesicles serve as heterophagosomes to transport absorbed protein to lysosomes, and (c) some small coated vesicles serve as primary lysosomes to transport hydrolytic enzymes from the Golgi complex to multivesicular bodies.     

Chromaffinity, uranaffinity and argentaffinity of small granule-containing (SGC) cells in rat superior cervical ganglia.

A systemic examination on the small granule-containing (SGC) cells in rat superior cervical ganglia was conducted by conventional and cytochemical electron microscopy including chromaffin, argentaffin and uranaffin reactions. According to the fine structure of dense cored vesicles (DCVs) in the cytoplasm, three types of small granule-containing (SGC) cells were revealed--Type I: 90-160 nm vesicles with cores of moderate or low electron density; Type II: 130-330 nm vesicles, polymorphic with highly electron dense cores; Type III: elongated vesicles (170 nm x 60 nm) with cores of moderate to low electron density. The majority of SGC cells were the Type I cells (78%) and Type II and III cells made up 13% and 9% of SGC cell population, respectively. Cytochemical results demonstrated that only the Type II cells displayed a positive chromaffin reaction and all three types of SGC cells showed argentaffinity and uranaffinity. The present study is the first to demonstrate the argentaffin reaction at ultrastructural level in SGC cells of sympathetic ganglia. Based on the results of the present study we also concluded that (1) the DCVs of Type II SGC cells contained noradrenaline and (2) biogenic amines and nucleotides (ATPs) coexisted in the DCVs of all three types of SGC cells.