The dry and wet stigmas ofPrimula obconica: Ultrastructural and cytochemical dimorphisms

Summary The stigmatic papillae of the distylous speciesPrimula obconica are studied by means of cytochemical, light and electron microscopic techniques. The papillae on thrum stigmas are smaller than those on pin stigmas. At the bud stage, secretory vesicles are not a conspicious part of the cytoplasm, although certain signs of secretory activity are present. The young papillae bear the thin, superficial pellicle typical to dry stigmas. Small vesicles are numerous in mature papillae of both morphs, and seem to originate from the ER. A layer of closely packed, osmiophilic globuli is present in the outermost part of mature walls of pin papillae. At sites with cuticle disruption, the globuli seem to migrate outwards to be incorporated with the copious, blistery exudate. Due to this exudate the pin stigma is characterized as wet. Cytochemical tests suggest that the exudate contains mainly lipids, and different carbohydrates and protein are detected. It reacts positively in tests for peroxidase, acid phosphatase, and non-specific esterases. The thrum stigma remains dry at maturity, with a distinct pellicle also reacting positively to the enzyme tests. Only a few, scattered osmiophilic globuli, sized and situated as those in the pin papillae walls, are found in the thrum walls, and they do not form a proper layer.Thus the generally accepted correlation between dry stigmas and the sporophytic kind of self-incompatibility system is not substantiated withinP. obconica, and the possible influence of the dimorphisms to the pollen/stigma interaction is discussed.     

ULTRASTRUCTURAL CHANGES OF CHLOROPLASTS IN ATTACHED AND DETACHED,AGING PRIMARY WHEAT LEAVES

Degradation of chloroplasts is shown in mesophyll cells of primary leaves of wheat. The sequence of ultrastructural changes in chloroplasts of naturally senescing leaves is compared with that of detached, aging leaves. In chloroplasts of naturally senescing leaves, the first indications of aging are the appearance of osmiophilic globuli and reorientation of the thylakoidal system. The membranes of the grana and intergrana lamellae then become distended and later dissociate into distinct vesicles. Concurrent with these membrane changes, osmiophilic globuli increase in size and number, and the stroma breaks down. Finally, the chloroplast envelope ruptures and plastid contents disperse throughout the cell's interior. In chloroplasts of mesophyll cells in detached, aging leaves, initial changes also include appearance of osmiophilic globuli, but later stages of chloroplast degradation are different. The chloroplast envelope ruptures before the lamellae break down. Swelling of grana and intergrana lamellae is not pronounced and, additionally, the thylakoidal system degenerates without forming vesicles or numerous osmiophilic globuli. These differences in the sequence of chloroplast degradation indicate that naturally senescing leaves rather than detached, aging leaves should be used in studies of chloroplast senescence.     

FINE STRUCTURE OF THE ZYGOTE AND EARLY EMBRYO IN QUERCUS GAMBELII

This investigation begins with the late zygote and traces ultrastructural development to the late globular stage of the embryo. Two nucleoli and satellite nucleoli sometimes occur in the zygote nucleus. Mitochondria, dictyosomes, cytoplasmic ribosomes, rough ER, and lipid bodies are numerous in the zygote. Microbodies are occasionally seen. The cell wall becomes well developed before the first division. No plasmodesmata occur in the zygote wall. The basal cell of the proembryo and the suspensor cells of the later embryo have very dense cytoplasm with a high concentration of cytoplasmic ribosomes. The nuclei are very electron opaque. The terminal cell and the cells of the embryo proper have a fine structure similar to that of the zygote. Plastids increase in number, size, starch content, and amount of thylakoid lamellae as the embryo develops. Mitochondria are numerous and appear active at all stages. Dictyosome activity, ribosomal aggregation, and the amount of ER are highest during the late globular stage. Lipid bodies are present up to the early globular stage, then disappear. The inner cell walls of the embryo are thin and have many plasmodesmata. These walls begin to thicken at the late globular stage, and at this time the size of the embryo begins to show an increase over that of the zygote. The results show a corresponding increase in the amount and activity of the metabolic machinery as the development of the embryo progresses. Lipids are probably more important as a nutrient source in the zygote and early embryo; starch becomes more important in the late stages. Absorption of nutrient material into the embryo sac and developing embryo appears to be from the chalazal end.     

Zoosporogenesis in Pseudoperonospora cubensis, the causal agent of cucurbit downy mildew

During sporulation of Pseudoperonospora cubensis on cucumber leaves ( Cucumis saliva ) zoosporangia are formed on the dichotomously branched sporangiophore. The mature zoosporangium has a preformed discharge papilla and the cytoplasm is uncleaved. The zoosporangium wall is decorated and the outer layer of the wall is electron opaque in ultrathin sections. As the zoosporangium is able to survive freezing (- 18°C) for prolonged periods of time (3–4 months) the zoosporangium may serve as the "resting" structure which survives overwintering in Northern latitudes in the absence of oospore formation.
Zoospore cleavage can be synchronized by placing freshly harvested zoosporangia in distilled water. Cleavage of the zoosporangial cytoplasm is by means of the fusion of small vesicles apparently derived from dictyosomes which become highly active after zoosporogenesis is induced.
Vesicles with an osmiophilic electron opaque content are the dominant type of vesicle found in the zoosporangia. The content of these vesicles undergoes dynamic changes during zoosporogenesis and during the late stages of sporogenesis the content becomes finely striated as is typical of these vesicles when observed in the zoospore. On the basis of the results presented here it is suggested that zoosporangium formation and zoosporogenesis in P. cubensis could serve as a model system for assays with obligate oomycetous plant pathogens, also in relation to fungicide mode of action studies.     

Reproduction in seagrasses: pollen wall morphogenesis in Amphibolis antarctica and wall structure in filiform grains

The pre–meiotic anther of the marine angiosperm Amphibolis antarctica contains microsporocytes and sterile cells. The microsporocytes divide conventionally to produce tetrads, but the sterile cells degenerate and contribute to the future pe–riplasmodium. Each tetrad of young microspores is contained within a vesicle defined by a membrane. After release from the tetrad, the microspores increase in length and rapidly become filiform. The microspore nucleus soon divides and partitioning of the cytoplasm delimits the generative cell from the vegetative cell of the binucleate pollen grain. The division and the early pollen growth occurs while the grains are segregated within vesicles in the periplasmodium. These compartments, established at microspore release, remain structurally intact throughout the vacuolate period of pollen development, when pollen wall assembly begins. This process is initiated as particles migrate from the inner face of the vesicle membrane into the lumen of the vesicle and microfibrillar elements form between adjacent particles. The particles and microfibrils form a loose, three–dimensional network. The vesicle membrane then disappears and the binuclate grains become immersed in the tapetal residuum. Additional wall components are now deposited upon the primary fibrillar stratum. Short lamellae, resembling fragments of membrane, frequently associated with electron–opaque globuli, are found intermixed with the surface microfibrils. Apparently, granular material originating in the degenerating periplasmodium may be the precursor of the globuli, and contact with the lamellae brings about an alteration in state. At this stage the pollen wall is resolved as two distinct fibrillar strata and the lamellae and globuli are incorporated as inclusions into the superficial zone of the outer stratum. The mature pollen wall exhibits faint stratification and the presence of the subsurface inclusions is readily demonstrated in germinating grains by section staining with phosphotungstic acid. The pollen wall in A. antarctica is compared with that in filiform grains of other seagrasses.     

The verification of cytochemical tests for ATP-ase activity in plant cells using X-ray microanalysis

Summary Electron microprobe analysis (EMMA 4) was carried out on two types of electron opaque deposit found in thin sections of barley root tips as a result of cytochemical tests for ATP-ase activity.The granular type of deposit, which mainly occurs in radial and tangential cell walls of epidermal and sub-epidermal cells, was shown to contain lead, whereas lead was absent from the opaque globular deposits, which are much more generally distributed and always associated with membrane structures. Thus the latter deposits, which contain osmium and have often been interpreted as ATP-ase reaction products, should be regarded as artifacts of the fixation and rinsing procedures. It is suggested that calcium may play a role in the formation of the osmiophilic deposits.     

Germination of the resting sporangia ofPhysoderma maydis,the causal agent ofPhysoderma disease of maize

Summary The structural and developmental characteristics of the resting sporangium in uniflagellate phycomycetes, together with the type of zoospore, are of high taxonomic value. Among these fungi, however, only a few electron microscopic investigations have been published on this topic, mainly due to technical problems. In the present study ofPhysoderma maydis (Blastocladiales) these problems were overcome as the resting sporangia in this species are formed synchronously, in large numbers, the germination is readily induced and the impermeability of the resting sporangium wall can be circumvented by shaking the prefixed sporangia with glass beads.The germination of the resting sporangia ofP. maydis is described by correlative light and electron microscopic studies and discussed in relation to related investigations on sporogenesis: The germination process starts by a breakdown of large electron-dense accretions found in the resting stage. Simultaneously, the peripheral location of the lipid bodies is lost. The large operculum is pushed open by a protrusion of the inner sporangial wall; an additional wall layer is formed during this process. Synaptonemal complexes are found in the nuclei at this stage, as are nuclear division figures which suggests anEuallomyces type of life cycle for this fungus. Cleavage vesicles, formed from dictyosomes or endoplasmic reticulum, ultimately separate the sporangial content into meiospores. The sequential assembly of organelles into the side body complex is described. Sequestering of the ribosomes into a nuclear cap is interpreted as taking place immediately prior to zoospore discharge.     

The ultrastructure of zoospores ofAphanomyces euteiches and of their encystment and subsequent germination

Summary The ultrastructure ofAphanomyces euteiches during the periods of zoospore motility, encystment, and germination has been studied. The motile spore has two heterokont flagella inserted laterally into the groove of the zoospore body where each is attached to a kinetosome. The kinetosomes and flagella are anchored into the zoospore body by rootlets comprised of two rows of microtubules with up to 12 microtubules in the outer row and are attached by fine threads to a striate fiber bundle. Secondary microtubules are attached at right angles at regular intervals along the rootlets. An unidentified body, 1.25m in diameter, containing helical fibers 16 nm in diameter is present in each zoospore. This body is situated near the two kinetosomes on the side of the pyriform nucleus opposite the contractile vacuole. The Golgi complex is between the nucleus and the contractile vacuole. The latter is surrounded by a 0.5–1.0m wide zone of Golgi proliferated vesicles. Ribosomes are generally absent from this region. Endoplasmic reticulum containing tubules within the expanded cisternae are also present. Vesicles with striated electron opaque inclusions and vesicles containing a granular cortex and center that developed in previous stages of zoosporogenesis were also present. During encystment of the zoospore the latter vesicles disappear. The two flagella are shed at this time leaving a membrane-bounded granular knob protruding from each of the kinetosome terminal plates. The contractile vacuole becomes disorganized and the zoospore assumes a spherical shape. Cyst wall deposition begins immediately and is completed in 30 minutes. The spore begins to germinate 1 hour following initiation of encystment with the appearance of a bulge in the cyst wall which elongates into a germ tube. Mitotic nuclear division follows.Research supported by the College of Agricultural and Life Sciences Station Project No. 1281.Research assistant and Professor. The advice and assistance of G. A. deZoeten, G. R.Gaard, and S.Vicen are most gratefully acknowledged.     

Reproduction by zoospores inOedogonium

Summary Emergence of zoospores ofOedogonium and their subsequent developmental changes have been studied using live material and sections prepared for light and electron microscopy. Release commences with rupture of the cell wall at its pre-weakened site near the apical caps. The pliable protoplast of the zoospore becomes completely spherical once free of the wall; it is enclosed within the hyaline vesicle which expands continuously and then disappears. Meanwhile, as the flagella become active, the zoospore begins to elongate and its dome starts to protrude from a circular constriction where the flagella are inserted. Once free of the hyaline vesicle, it is actively motile for a variable period, during which elongation continues. The motile phase ceases when the zoospore begins to vibrate, whereupon the flagella are all violently shed. Soon after this, the constriction disappears from around the dome which becomes more pointed; the immobile cell now elongates further, increasing in volume. The cell periphery contains numerous contractile vacuoles. Zoospore elongation may be associated with a proliferation of longitudinal microtubules, and once the flagella are shed, the flagellar rootlet system disintegrates, probably releasing the rootlet microtubules. Mechanisms involved in the release of the zoospore are also discussed.     

Ultrastructural Studies on the Development of Oil Cells in Litsea pungens

The developmental process of oil cells in the shoot of Litsea pungens Hemsl. has been studied with transmission electron microscopy. According to the development of the three layers of cell wall, the developmental process could be divided into 4 stages. In stage 1, the cell wall consisted only of a primary (the outmost) cellulose layer, which might further be divided into two substages, the oil cell initial, and the vacuolizing oil cell. During this stage, there were some small electron translucent vesicles and dark osmiophilic droplets of variant sizes in the different-shaped plastids. It was observed that some dark and gray osmiophilic materials coalesced to vacuoles in the cytoplasm. In stage 2, a lamellated suberin layer accumulated inside the primary cellulose layer. In stage 3, a thicker and looser inner cellulose wall layer was formed gradually inside the suberin layer. Some dark osmiophilic droplets have been observed in this loose inner cellulose wall layer. The plasmodesmata were blocked up and became a special structure. Then, the big vacuole, which is the oil sac, was full of osmiophilic oil. In stage 4, the oil cell became matured and the cytoplasm disintegrated. The oil sac enveloped from plasmalemma was attached to the cupule, which was formed by the protuberance of the inner cellulose wall layer into the lumen. After the maturity of oil cell, the ground cytoplasm began to disintegrate and became electron opaque or exhibited in a disordered state, and the osmiophilic oil appeared light gray.     

木姜子油细胞发育的超微结构研究

利用超薄切片法和透射电镜研究了木姜子（Litsea pungens Hemsl.）油细胞的发育过程。油细胞3层细胞壁的发育可分为4个阶段,阶段1：油细胞仅有初生纤维素壁层,又可分为原始细胞和细胞 泡化两个时期。此阶段质体具透明小泡和黑色嗜锇物质,并与液泡融合。阶段2：木栓质化壁层的形成,片层状木栓质不断叠加在初生纤维素壁内侧,其细胞结构与前期相似,阶段3：内纤维素壁层的形成,较厚而松散的内纤维素壁层叠加在木栓质化壁层的内侧,在内纤维素壁层中可见黑色嗜锇物质,胞间连丝成为被阻塞的特化结构,此时大液泡被嗜锇油脂充满,成为油囊。阶段4：油细胞成熟及细胞质解体,杯形构造由内纤维素壁层向细胞腔内突起形成,油囊由液泡膜包被连接到杯形构造上,油呈浅灰色嗜锇状态,其细胞质和细胞器解体,变得电子不透明或呈杂乱状态。     

Glandular scale development and essential oil secretion in Origanum dictamnus L.

Glandular scales of Origanum dictamnus L. originate from a single protodermal cell. They are composed of a 12-celled head and an unicellular stalk and foot. During the early stages of gland differentiation, the head cells possess a small number of plastids which contain globular inclusions. Similar inclusions are also observed in the plastids of the stalk and the foot cell. The lateral walls of the stalk cell progressively undergo cutinization which does not extend to the upper and lower periclinal walls. At the onset of secretion the electron density of the plasmalemma region lining the apical walls of the head cells remarkably increases. These walls are impregnated with an osmiophilic substance identical in appearance to the content of the subcuticular space. In a following stage of the secretory process osmiophilic droplets of various size arise in the cytoplasm of the secretory cells which undergoes simultaneously a reduction of its initial density. After secretion has been concluded the protoplast of the head cells becomes gradually degenerated. The chlorenchyma cells of the mesophyll possess numerous microbodies closely associated with various organelles. In the cytoplasm of these cells crystalloids occasionally occur.     

Surface coat transformation and capsule formation by Leuconostoc mesenteroides NCDO 523 in the presence of sucrose

When Leuconostoc mesenteroides NCDO 523 was grown in MRS browth, electron microscopy of cells fixed in the presence of ruthenium red showed that the cell wall was covered with a thin layer of filamentous material. When MRS-grown cells were resuspended in the same medium supplemented with 3.6% sucrose, this surface coat doubled in thickness and a number of radial thickenings appeared within it. After 3 h the filamentous component of the surface coat had disappeared leaving only the radial projections. The progressive accumulation of polymer to produce a capsule visible by light microscopy was observed in only about 20% of the population. In this minority of cells, a dense globular dextran composed of fibrillar and particulate elements was always produced in the initial stages of synthesis. After 18 h, the dextran capsule was generally composed of an inner globular and outer fibrillar layer. It appeared that the outer layer was derived from the globular dextran of the capsule by a process of dispersion.     

Localization of α-galactomannan and of wheat germ agglutinin receptors inSchizosaccharomyces pombe

The location of galactomannan on the surface ofSchizosaccharomyces pombe cells was reexamined by scanning electron microscopy by an indirect but specific method using gold markers. The polysaccharide was found on the cell surface and at the end beginning to grow but not on the wall established by division. Galactomannan was also localized onS. pombe thin sections by transmission electron microscopy using the same method. The polysaccharide was found deposited in two layers in the cell wall, i.e. at the periphery of the wall and near the plasmalemma. The septum was also marked but mainly near the plasmalemma. These results indicated that the polysaccharide is elaborated onto the outside of the wall during extension but not during septum formation. When thin sections ofS. pombe were marked with gold granules labeled with wheat germ agglutinin, marking was found in vacuoles but not in the cell wall. This confirmed thatS. pombe cell wall is devoid of chitin.Non-Standard Abbreviations Au gold colloid - RCA_I Ricinus communis lectin - SEM scanning electron microscopy - TEM transmission electron microscopy - WGA wheat germ agglutinin     

Apomixis in plants: structural and functional aspects of diplospory inPoa nemoralis andP. palustris

Summary Data on structural and functional aspects of mitotic diplospory and later stages of apomictic seed formation are reported forPoa palustris andP. nemoralis. In this study, the plant material of two Russian populations ofP. nemoralis andP. palustris were used for transmission electron microscope observations. Seed formation was investigated by phase contrast microscopy in two populations ofP. nemoralis collected in The Netherlands. The processes of transformation of the megasporocytes to the megaspore mother cells of diplosporous embryo sacs, and thereafter to one- and two-nucleate diplosporous embryo sacs (Antennaria type) were characterized by an increase of cell size, structural and functional reorganization of the nucleus, nucleolus, and cytoplasm, and cell isolation as a result of thickening of the cell wall. These were accompanied by an increase in the cell metabolic activity inferred from visual evidence of the activation of nucleus, nucleolus, endoplasmic reticulum, dictyosomes, mitochondria, and from the appearance of a dense population of ribosomes and polysomes. The diplosporous embryo sac of the Antennaria type was characteristic for bothP. nemoralis andP. palustris. No signs of the presence of synaptonemal complexes were observed during the process of diplosporous-embryo-sac megaspore mother cell differentiation and division. About 90–95% of the diploid egg cells of diplosporous embryo sacs were able to produce apomictic embryos. These embryos developed before anthesis. However, many of them degenerated at the globular stage because of lack of endosperm. The ultrastructural events occurring during the process of diplospory of apomictic species, and meiosis and megagametogenesis of sexually reproduced plants are discussed.Abbreviations DMC megaspore mother cell of diplosporous embryo sac - TEM transmission electron microscopy - ER endoplasmic reticulum     

Expression of Polarity during early Development of Microspore-derived and Zygotic Embryos of Brassica napus L. cv. Topas

Microspore derived (MS-)embryogenesis and zygotic embryogenesis of Brassica napus L. cv. Topas were investigated by light and scanning electron microscopy to reveal the expression of polarity during the transition phase from globular to heart and torpedo shape. During the first 5 days of MS-embryo formation, the cell wall of the former microspores remained intact and a globular mass of cells developed within. Pollen walls ruptured after 5 days of culture; embryos proceeded through heart-shape and torpedo-shape stages within 15 days in a way comparable to, but faster than observed during zygotic embryogenesis. Expression of polarity in globular and elongating MS-embryos was analyzed by detection of the distribution of activated calmodulin as well as of free cytosolic calcium by using confocal scanning laser microscopy, and by the detection of starch. Calmodulin was evenly distributed in globular embryos and only exhibited clear polar distribution in elongated embryos. Free cytosolic Ca²⁺ accumulated in the protoderm of globular embryos and in the central cylinder of torpedo shaped embryos, but never showed polar distribution. Accumulation of starch granules at the root poles of both sexual as well as MS-embryos, however, indicated polar distribution before the transition from globular to heart shape stage. Since the local rupture of the pollen wall of 6-day-old MS-embryos was never preceded by the decrease of starch at that site, it is likely that the rupture of the pollen wall plays an important role in the local activation of cell metabolism and thus in the determination of the polarity axis in MS-embryos.     

Actin filaments predominate in morphogenic cell stages, whereas plaques predominate in non-morphogenic cell stages in Peronosporomycetes

We investigated the structural distribution of both types of actin arrays, filaments and plaques, in a soil-borne phytopathogenic peronosporomycete (oomycete), Aphanomyces cochlioides, under standardized host-free bioassays. The phenomenon was monitored during progression through all the asexual developmental processes of the organism. It was noted that the filamentous-form of actin was predominant during the morphogenic (morphologically active) stages of development. Conversely, during non-morphogenic (morphologically quiescent) stages, plaques dominated. From these analyses, we proposed a criterion that predominance of an actin form relates to, and precedes the morphological behaviour of a cellular stage in Peronosporomycetes. A decrease in the quantity of plaques in the encysted zoospore (non-morphogenic stage) during its developmental progression into morphogenic stages, both in germination and regeneration processes, asserted the notion that plaques function as the organization centres and are related to the reorganization of cell structure and the transition of the cell into a new stage. Furthermore, polymerization of filamentous-form during emergence stages in zoospore regeneration process revealed that filaments render motility to a developing zoospore. This unprecedented function of filaments in the developing zoospores was demonstrated using nicotinamide (0.8 × 10⁻⁶ m), which did not cause actin disruption, but could induce zoospore encystment, and its further replacement with water triggered the zoospore emergence process. Additionally, by using latrunculin B, an actin polymerization inhibitor, we also demonstrated the functional necessity of actin during various developmental processes in Aphanomyces.     

Ultrastructure of the Eyespot and its Possible Significance in Phototaxis of Tetracystis excentrica*†

SYNOPSIS. The eyespot of the zoospore of Tetracystis excentrica (a green alga) has been studied by light and electron microscopy. In Tetracystis the eyespot consists of about 110 osmiophilic granules which form a plate in the anterior third of the cell. The granules are about 80 Å in diameter and are found in the outermost portion of the chloroplast; they commonly show hexagonal close packing and a hexagonal shape. The granules are confined positionally by the chloroplast envelope and an inner thylakoid. The plasmalemma over the eyespot is thickened and is separated from the chloroplast envelope by a 50 mμ space. The eyespot of Tetracystis is compared with others reported in the literature and the possible functional significance of these studies is discussed. The possibility that the eyespot plate in Tetracystis serves as a shading device rather than the primary photoreceptor is considered.     

CONJUGATION IN THE DESMID CLOSTERIUM LITTORALE1

Conjugation in Closterium has been studied using sectioned material prepared for light and electron microscopy. Prior to conjugation, cells become dense and accumulate lipid droplets. Conjugation commences in paired cells with the formation through a point of mutual contact of a circumferential strip of papilla wall material which is not necessarily centrally situated in each cell. Some microtubules are initially present, near this specialized wall which grows all around each cell but asymmetrically so that the area of papillae in contact increases and each papilla balloons out toward the other, kinking the semicells. Vesicles probably contribute material to this wall. Microtubules disappear from near the papilla as it expands; they are then often found around the older semicell wall immediately adjacent to the papilla. An enlarging vacuole is always formed in each papilla. Cytoplasmic shrinkage is first evident from an accumulation of mucilage between the plasmalemma and wall at the tip of each semicell; the terminal vacuoles collapse and disappear. During further shrinkage of the protoplast, alveolate structures are common on the plasmalemma, and the papilla wall material separating the protoplasts thins out and disappears. Meanwhile, a profound general change renders the cytoplasm far less osmiophilic and stainable for both light and electron microscopy, as it becomes relatively homogeneous and granular, revealing a considerable loss in minor cytoplasmic structures; grana in the chloroplasts become bloated. These changes are not considered processing artefacts, but may result from a breakdown of vegetative cell structures no longer needed for zygote formation. After fusion, the zygote protoplast tends to round up. Zygote maturation commences with deposition of a multilayered wall and shrinkage of the protoplast. Then the very thick zygote wall proper is slowly secreted; meanwhile, very considerable cytoplasmic condensation now renders the cell increasingly osmiophilic and dense until ultrastructural detail becomes totally obscured. Fairly consistent changes in the appearance and probably in the function of the golgi bodies were noted. The results are discussed in terms of the possible function of cell organelles and their role in the mechanics of conjugation.     

Globuli ossei in the long limb bones of Pleurodeles waltl (Amphibia,Urodela, Salamandridae)

To date, little is known about the structure of the cells and the fibrillar matrix of the globuli ossei, globular structures showing histochemical properties of an osseous tissue, sometimes found in the resorption front of the hypertrophied cartilage in many tetrapods, and easily observed in the long bones of the Urodele Pleurodeles waltl. Here, we present the results obtained from the appendicular long bones of metamorphosed juveniles and subadults using histological and histochemical methods and transmission electron microscopy. The distal part of the cone‐shaped cartilage contains a heterogeneous cell population composed of the typical “light” hypertrophic chondrocytes and scarce “dark” hypertrophic chondrocytes. The “dark” chondrocytes display ultrastructural characteristics suggesting that they probably undergo degeneration through chondroptosis. However, in the hypertrophic, calcified cartilage close to the erosion front by the marrow, several noninvaded chondrocytic lacunae retained cells that do not show any morphological characteristics of degeneration and that cannot be identified as regular chondrocytes or osteocytes. These modified chondrocytes that have lost their regular morphology, appear to be active in the terminal cartilage and synthesize collagen fibrils of a peculiar diameter intermediate between the Type I collagen found in bone and the Type II collagen characteristic of cartilage. It is suggested that the local occurrence of globuli ossei is linked to a low rate of longitudinal growth as is the case in the long bones of postmetamorphic urodeles. J. Morphol. 275:1226–1237, 2014. © 2014 Wiley Periodicals, Inc.